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Mathey-Prevot, Bernard

Overview:

The central focus of the lab is to understand how signaling pathway architecture and integration result in specific cell fates and how these properties have been hijacked in cancer cells. In particular, we are interested in assessing the extent to which cell-to-cell heterogeneity can affect the temporal dynamics and regulation of signaling pathways. We are focusing on the E2F/Rb network and have established a platform to follow in real time the activation and expression of E2F1 at the single cell level. In collaboration with the You lab (Duke Biomedical Engineering), which has developed mathematical approaches to model the behavior of the Rb/E2F network, we wish to identify functional characteristics in the signaling dynamics that may underlie disparate cell fates in response to a given external cue. Ultimately, our goal is to build an integrative understanding of the logic and structure in aberrant signaling network(s) known to drive tumor development and metastasis. 

Positions:

Research Professor of Pharmacology & Cancer Biology

Pharmacology & Cancer Biology
School of Medicine

Professor in Pediatrics

Pediatrics, Hematology-Oncology
School of Medicine

Member of the Duke Cancer Institute

Duke Cancer Institute
School of Medicine

Affiliate of the Regeneration Next Initiative

Regeneration Next Initiative
School of Medicine

Education:

Ph.D. 1983

Ph.D. — Rockefeller University

Research Fellow In Biology, Biology

Massachusetts Institute of Technology

Grants:

Temporal E2F Dynamics and Cell-Fate Decisions in Single Mammalian Cells

Administered By
Biomedical Engineering
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
September 15, 2014
End Date
June 30, 2018

Publications:

Division of labour between Myc and G1 cyclins in cell cycle commitment and pace control.

A body of evidence has shown that the control of E2F transcription factor activity is critical for determining cell cycle entry and cell proliferation. However, an understanding of the precise determinants of this control, including the role of other cell-cycle regulatory activities, has not been clearly defined. Here, recognizing that the contributions of individual regulatory components could be masked by heterogeneity in populations of cells, we model the potential roles of individual components together with the use of an integrated system to follow E2F dynamics at the single-cell level and in real time. These analyses reveal that crossing a threshold amplitude of E2F accumulation determines cell cycle commitment. Importantly, we find that Myc is critical in modulating the amplitude, whereas cyclin D/E activities have little effect on amplitude but do contribute to the modulation of duration of E2F activation, thereby affecting the pace of cell cycle progression.

Authors
Dong, P; Maddali, MV; Srimani, JK; Thélot, F; Nevins, JR; Mathey-Prevot, B; You, L
MLA Citation
Dong, P, Maddali, MV, Srimani, JK, Thélot, F, Nevins, JR, Mathey-Prevot, B, and You, L. "Division of labour between Myc and G1 cyclins in cell cycle commitment and pace control." Nature communications 5 (September 2014): 4750-.
PMID
25175461
Source
epmc
Published In
Nature Communications
Volume
5
Publish Date
2014
Start Page
4750
DOI
10.1038/ncomms5750

Cross-platform prediction of gene expression signatures.

Gene expression signatures can predict the activation of oncogenic pathways and other phenotypes of interest via quantitative models that combine the expression levels of multiple genes. However, as the number of platforms to measure genome-wide gene expression proliferates, there is an increasing need to develop models that can be ported across diverse platforms. Because of the range of technologies that measure gene expression, the resulting signal values can vary greatly. To understand how this variation can affect the prediction of gene expression signatures, we have investigated the ability of gene expression signatures to predict pathway activation across Affymetrix and Illumina microarrays. We hybridized the same RNA samples to both platforms and compared the resultant gene expression readings, as well as the signature predictions. Using a new approach to map probes across platforms, we found that the genes in the signatures from the two platforms were highly similar, and that the predictions they generated were also strongly correlated. This demonstrates that our method can map probes from Affymetrix and Illumina microarrays, and that this mapping can be used to predict gene expression signatures across platforms.

Authors
Lin, S-H; Beane, L; Chasse, D; Zhu, KW; Mathey-Prevot, B; Chang, JT
MLA Citation
Lin, S-H, Beane, L, Chasse, D, Zhu, KW, Mathey-Prevot, B, and Chang, JT. "Cross-platform prediction of gene expression signatures. (Published online)" PLoS One 8.11 (2013): e79228-.
PMID
24244455
Source
pubmed
Published In
PloS one
Volume
8
Issue
11
Publish Date
2013
Start Page
e79228
DOI
10.1371/journal.pone.0079228

FlyRNAi.org--the database of the Drosophila RNAi screening center: 2012 update.

FlyRNAi (http://www.flyrnai.org), the database and website of the Drosophila RNAi Screening Center (DRSC) at Harvard Medical School, serves a dual role, tracking both production of reagents for RNA interference (RNAi) screening in Drosophila cells and RNAi screen results. The database and website is used as a platform for community availability of protocols, tools, and other resources useful to researchers planning, conducting, analyzing or interpreting the results of Drosophila RNAi screens. Based on our own experience and user feedback, we have made several changes. Specifically, we have restructured the database to accommodate new types of reagents; added information about new RNAi libraries and other reagents; updated the user interface and website; and added new tools of use to the Drosophila community and others. Overall, the result is a more useful, flexible and comprehensive website and database.

Authors
Flockhart, IT; Booker, M; Hu, Y; McElvany, B; Gilly, Q; Mathey-Prevot, B; Perrimon, N; Mohr, SE
MLA Citation
Flockhart, IT, Booker, M, Hu, Y, McElvany, B, Gilly, Q, Mathey-Prevot, B, Perrimon, N, and Mohr, SE. "FlyRNAi.org--the database of the Drosophila RNAi screening center: 2012 update." Nucleic Acids Res 40.Database issue (January 2012): D715-D719.
PMID
22067456
Source
pubmed
Published In
Nucleic Acids Research
Volume
40
Issue
Database issue
Publish Date
2012
Start Page
D715
End Page
D719
DOI
10.1093/nar/gkr953

Network calisthenics: control of E2F dynamics in cell cycle entry.

Stimulation of quiescent mammalian cells with mitogens induces an abrupt increase in E2F1-3 expression just prior to the onset of DNA synthesis, followed by a rapid decline as replication ceases. This temporal adaptation in E2F facilitates a transient pattern of gene expression that reflects the ordered nature of DNA replication. The challenge to understand how E2F dynamics coordinate molecular events required for high-fidelity DNA replication has great biological implications. Indeed, precocious, prolonged, elevated or reduced accumulation of E2F can generate replication stress that culminates in either arrest or death. Accordingly, temporal characteristics of E2F are regulated by several network modules that include feedforward and autoregulatory loops. In this review, we discuss how these network modules contribute to "shaping" E2F dynamics in the context of mammalian cell cycle entry.

Authors
Wong, JV; Dong, P; Nevins, JR; Mathey-Prevot, B; You, L
MLA Citation
Wong, JV, Dong, P, Nevins, JR, Mathey-Prevot, B, and You, L. "Network calisthenics: control of E2F dynamics in cell cycle entry." Cell Cycle 10.18 (September 15, 2011): 3086-3094. (Review)
PMID
21900750
Source
pubmed
Published In
Cell Cycle
Volume
10
Issue
18
Publish Date
2011
Start Page
3086
End Page
3094
DOI
10.4161/cc.10.18.17350

A pathway-based classification of human breast cancer.

The hallmark of human cancer is heterogeneity, reflecting the complexity and variability of the vast array of somatic mutations acquired during oncogenesis. An ability to dissect this heterogeneity, to identify subgroups that represent common mechanisms of disease, will be critical to understanding the complexities of genetic alterations and to provide a framework to develop rational therapeutic strategies. Here, we describe a classification scheme for human breast cancer making use of patterns of pathway activity to build on previous subtype characterizations using intrinsic gene expression signatures, to provide a functional interpretation of the gene expression data that can be linked to therapeutic options. We show that the identified subgroups provide a robust mechanism for classifying independent samples, identifying tumors that share patterns of pathway activity and exhibit similar clinical and biological properties, including distinct patterns of chromosomal alterations that were not evident in the heterogeneous total population of tumors. We propose that this classification scheme provides a basis for understanding the complex mechanisms of oncogenesis that give rise to these tumors and to identify rational opportunities for combination therapies.

Authors
Gatza, ML; Lucas, JE; Barry, WT; Kim, JW; Wang, Q; Crawford, MD; Datto, MB; Kelley, M; Mathey-Prevot, B; Potti, A; Nevins, JR
MLA Citation
Gatza, ML, Lucas, JE, Barry, WT, Kim, JW, Wang, Q, Crawford, MD, Datto, MB, Kelley, M, Mathey-Prevot, B, Potti, A, and Nevins, JR. "A pathway-based classification of human breast cancer." Proc Natl Acad Sci U S A 107.15 (April 13, 2010): 6994-6999.
PMID
20335537
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
107
Issue
15
Publish Date
2010
Start Page
6994
End Page
6999
DOI
10.1073/pnas.0912708107

Do-it-yourself RNAi made easy?

Authors
Mathey-Prevot, B; Perrimon, N
MLA Citation
Mathey-Prevot, B, and Perrimon, N. "Do-it-yourself RNAi made easy?." Nat Methods 4.4 (April 2007): 308-309.
PMID
17396126
Source
pubmed
Published In
Nature Methods
Volume
4
Issue
4
Publish Date
2007
Start Page
308
End Page
309
DOI
10.1038/nmeth0407-308

Matter arising: off-targets and genome-scale RNAi screens in Drosophila.

Recently, the issue of off-target effects (OTEs) associated with long double stranded RNAs (dsRNAs) used in RNAi screens, such as those performed at the Drosophila RNAi Screening Center and other laboratories, has become a focus of great interest and some concern. Although OTEs have been recognized as an important source of false positives in mammalian studies (where short siRNAs are used as triggers), they were generally thought to be inconsequential in Drosophila RNAi experiments because of the use of long dsRNAs. Two recent papers have disputed this contention and show that significant off-target effects can take place with the use of some long dsRNAs in Drosophila cells. Together, these studies provide evidence that OTEs mediated by short homology stretches of 19nt or greater within long dsRNAs can contribute to false positives in Drosophila RNAi screens. Here, we address how widespread the occurrence of OTE is in Drosophila screens, focusing on the DRSC dsRNA collections, and we discuss the implication for the interpretation of results reported in RNAi screens to-date. Lastly, we summarize steps taken by the DRSC to redress that situation and include a set of recommendations to observe in future RNAi screens.

Authors
Perrimon, N; Mathey-Prevot, B
MLA Citation
Perrimon, N, and Mathey-Prevot, B. "Matter arising: off-targets and genome-scale RNAi screens in Drosophila." Fly (Austin) 1.1 (January 2007): 1-5.
PMID
18705022
Source
pubmed
Published In
Fly
Volume
1
Issue
1
Publish Date
2007
Start Page
1
End Page
5

Applications of high-throughput RNA interference screens to problems in cell and developmental biology.

RNA interference (RNAi) in tissue culture cells has emerged as an excellent methodology for identifying gene functions systematically and in an unbiased manner. Here, we describe how RNAi high-throughput screening (HTS) in Drosophila cells are currently being performed and emphasize the strengths and weaknesses of the approach. Further, to demonstrate the versatility of the technology, we provide examples of the various applications of the method to problems in signal transduction and cell and developmental biology. Finally, we discuss emerging technological advances that will extend RNAi-based screening methods.

Authors
Perrimon, N; Mathey-Prevot, B
MLA Citation
Perrimon, N, and Mathey-Prevot, B. "Applications of high-throughput RNA interference screens to problems in cell and developmental biology." Genetics 175.1 (January 2007): 7-16. (Review)
PMID
17244760
Source
pubmed
Published In
Genetics
Volume
175
Issue
1
Publish Date
2007
Start Page
7
End Page
16
DOI
10.1534/genetics.106.069963

Drug-target identification in Drosophila cells: combining high-throughout RNAi and small-molecule screens.

RNA interference (RNAi) and small-molecule approaches are synergistic on multiple levels, from technology and high-throughput screen development to target identification and functional studies. Here, we describe the RNAi screening platform that we have established and made available to the community through the Drosophila RNAi Screening Center at Harvard Medical School. We then illustrate how the combination of RNAi and small-molecule HTS can lead to effective identification of targets in drug discovery.

Authors
Perrimon, N; Friedman, A; Mathey-Prevot, B; Eggert, US
MLA Citation
Perrimon, N, Friedman, A, Mathey-Prevot, B, and Eggert, US. "Drug-target identification in Drosophila cells: combining high-throughout RNAi and small-molecule screens." Drug Discov Today 12.1-2 (January 2007): 28-33. (Review)
PMID
17198970
Source
pubmed
Published In
Drug Discovery Today
Volume
12
Issue
1-2
Publish Date
2007
Start Page
28
End Page
33
DOI
10.1016/j.drudis.2006.10.006

A case study of the reproducibility of transcriptional reporter cell-based RNAi screens in Drosophila.

Off-target effects have been demonstrated to be a major source of false-positives in RNA interference (RNAi) high-throughput screens. In this study, we re-assess the previously published transcriptional reporter-based whole-genome RNAi screens for the Wingless and Hedgehog signaling pathways using second generation double-stranded RNA libraries. Furthermore, we investigate other factors that may influence the outcome of such screens, including cell-type specificity, robustness of reporters, and assay normalization, which determine the efficacy of RNAi-knockdown of target genes.

Authors
DasGupta, R; Nybakken, K; Booker, M; Mathey-Prevot, B; Gonsalves, F; Changkakoty, B; Perrimon, N
MLA Citation
DasGupta, R, Nybakken, K, Booker, M, Mathey-Prevot, B, Gonsalves, F, Changkakoty, B, and Perrimon, N. "A case study of the reproducibility of transcriptional reporter cell-based RNAi screens in Drosophila." Genome Biol 8.9 (2007): R203-.
PMID
17903264
Source
pubmed
Published In
Genome Biology
Volume
8
Issue
9
Publish Date
2007
Start Page
R203
DOI
10.1186/gb-2007-8-9-r203

Design and implementation of high-throughput RNAi screens in cultured Drosophila cells.

This protocol describes the various steps and considerations involved in planning and carrying out RNA interference (RNAi) genome-wide screens in cultured Drosophila cells. We focus largely on the procedures that have been modified as a result of our experience over the past 3 years and of our better understanding of the underlying technology. Specifically, our protocol offers a set of suggestions and considerations for screen optimization and a step-by-step description of the procedures successfully used at the Drosophila RNAi Screening Center for screen implementation, data collection and analysis to identify potential hits. In addition, this protocol briefly covers postscreen analysis approaches that are often needed to finalize the hit list. Depending on the scope of the screen and subsequent analysis and validation involved, the full protocol can take anywhere from 3 months to 2 years to complete.

Authors
Ramadan, N; Flockhart, I; Booker, M; Perrimon, N; Mathey-Prevot, B
MLA Citation
Ramadan, N, Flockhart, I, Booker, M, Perrimon, N, and Mathey-Prevot, B. "Design and implementation of high-throughput RNAi screens in cultured Drosophila cells." Nat Protoc 2.9 (2007): 2245-2264.
PMID
17853882
Source
pubmed
Published In
Nature Protocols
Volume
2
Issue
9
Publish Date
2007
Start Page
2245
End Page
2264
DOI
10.1038/nprot.2007.250

Evidence of off-target effects associated with long dsRNAs in Drosophila melanogaster cell-based assays.

To evaluate the specificity of long dsRNAs used in high-throughput RNA interference (RNAi) screens performed at the Drosophila RNAi Screening Center (DRSC), we performed a global analysis of their activity in 30 genome-wide screens completed at our facility. Notably, our analysis predicts that dsRNAs containing > or = 19-nucleotide perfect matches identified in silico to unintended targets may contribute to a significant false positive error rate arising from off-target effects. We confirmed experimentally that such sequences in dsRNAs lead to false positives and to efficient knockdown of a cross-hybridizing transcript, raising a cautionary note about interpreting results based on the use of a single dsRNA per gene. Although a full appreciation of all causes of false positive errors remains to be determined, we suggest simple guidelines to help ensure high-quality information from RNAi high-throughput screens.

Authors
Kulkarni, MM; Booker, M; Silver, SJ; Friedman, A; Hong, P; Perrimon, N; Mathey-Prevot, B
MLA Citation
Kulkarni, MM, Booker, M, Silver, SJ, Friedman, A, Hong, P, Perrimon, N, and Mathey-Prevot, B. "Evidence of off-target effects associated with long dsRNAs in Drosophila melanogaster cell-based assays." Nat Methods 3.10 (October 2006): 833-838.
PMID
16964256
Source
pubmed
Published In
Nature Methods
Volume
3
Issue
10
Publish Date
2006
Start Page
833
End Page
838
DOI
10.1038/nmeth935

Minimizing the risk of reporting false positives in large-scale RNAi screens.

Large-scale RNA interference (RNAi)-based analyses, very much as other 'omic' approaches, have inherent rates of false positives and negatives. The variability in the standards of care applied to validate results from these studies, if left unchecked, could eventually begin to undermine the credibility of RNAi as a powerful functional approach. This Commentary is an invitation to an open discussion started among various users of RNAi to set forth accepted standards that would insure the quality and accuracy of information in the large datasets coming out of genome-scale screens.

Authors
Echeverri, CJ; Beachy, PA; Baum, B; Boutros, M; Buchholz, F; Chanda, SK; Downward, J; Ellenberg, J; Fraser, AG; Hacohen, N; Hahn, WC; Jackson, AL; Kiger, A; Linsley, PS; Lum, L; Ma, Y; Mathey-Prévôt, B; Root, DE; Sabatini, DM; Taipale, J; Perrimon, N; Bernards, R
MLA Citation
Echeverri, CJ, Beachy, PA, Baum, B, Boutros, M, Buchholz, F, Chanda, SK, Downward, J, Ellenberg, J, Fraser, AG, Hacohen, N, Hahn, WC, Jackson, AL, Kiger, A, Linsley, PS, Lum, L, Ma, Y, Mathey-Prévôt, B, Root, DE, Sabatini, DM, Taipale, J, Perrimon, N, and Bernards, R. "Minimizing the risk of reporting false positives in large-scale RNAi screens." Nat Methods 3.10 (October 2006): 777-779.
PMID
16990807
Source
pubmed
Published In
Nature Methods
Volume
3
Issue
10
Publish Date
2006
Start Page
777
End Page
779
DOI
10.1038/nmeth1006-777

FlyRNAi: the Drosophila RNAi screening center database.

RNA interference (RNAi) has become a powerful tool for genetic screening in Drosophila. At the Drosophila RNAi Screening Center (DRSC), we are using a library of over 21,000 double-stranded RNAs targeting known and predicted genes in Drosophila. This library is available for the use of visiting scientists wishing to perform full-genome RNAi screens. The data generated from these screens are collected in the DRSC database (http://flyRNAi.org/cgi-bin/RNAi_screens.pl) in a flexible format for the convenience of the scientist and for archiving data. The long-term goal of this database is to provide annotations for as many of the uncharacterized genes in Drosophila as possible. Data from published screens are available to the public through a highly configurable interface that allows detailed examination of the data and provides access to a number of other databases and bioinformatics tools.

Authors
Flockhart, I; Booker, M; Kiger, A; Boutros, M; Armknecht, S; Ramadan, N; Richardson, K; Xu, A; Perrimon, N; Mathey-Prevot, B
MLA Citation
Flockhart, I, Booker, M, Kiger, A, Boutros, M, Armknecht, S, Ramadan, N, Richardson, K, Xu, A, Perrimon, N, and Mathey-Prevot, B. "FlyRNAi: the Drosophila RNAi screening center database." Nucleic Acids Res 34.Database issue (January 1, 2006): D489-D494.
PMID
16381918
Source
pubmed
Published In
Nucleic Acids Research
Volume
34
Issue
Database issue
Publish Date
2006
Start Page
D489
End Page
D494
DOI
10.1093/nar/gkj114

Drosophila genome-wide RNAi screens: are they delivering the promise?

The emergence of RNA interference (RNAi) on the heels of the successful completion of the Drosophila genome project was seen by many as the ace in functional genomics: Its application would quickly assign a function to all genes in this organism and help delineate the complex web of interactions or networks linking them at the systemic level. A few years wiser and a number of genome-wide Drosophila RNAi screens later, we reflect on the state of high-throughput RNAi screens in Drosophila and ask whether the initial promise was fulfilled. We review the impact that this approach has had in the field of Drosophila research and chart out strategies to extract maximal benefit from the application of RNAi to gene discovery and pursuit of systems biology.

Authors
Mathey-Prevot, B; Perrimon, N
MLA Citation
Mathey-Prevot, B, and Perrimon, N. "Drosophila genome-wide RNAi screens: are they delivering the promise?." Cold Spring Harb Symp Quant Biol 71 (2006): 141-148. (Review)
PMID
17381290
Source
pubmed
Published In
Cold Spring Harbor Laboratory: Symposia on Quantitative Biology
Volume
71
Publish Date
2006
Start Page
141
End Page
148
DOI
10.1101/sqb.2006.71.027

High-throughput RNA interference screens in Drosophila tissue culture cells.

This chapter describes the method used to conduct high-throughput screening (HTs) by RNA interference in Drosophila tissue culture cells. It covers four main topics: (1) a brief description of the existing platforms to conduct RNAi-screens in cell-based assays; (2) a table of the Drosophila cell lines available for these screens and a brief mention of the need to establish other cell lines as well as cultures of primary cells; (3) a discussion of the considerations and protocols involved in establishing assays suitable for HTS in a 384-well format; and (A) a summary of the various ways of handling raw data from an ongoing screen, with special emphasis on how to apply normalization for experimental variation and statistical filters to sort out noise from signals.

Authors
Armknecht, S; Boutros, M; Kiger, A; Nybakken, K; Mathey-Prevot, B; Perrimon, N
MLA Citation
Armknecht, S, Boutros, M, Kiger, A, Nybakken, K, Mathey-Prevot, B, and Perrimon, N. "High-throughput RNA interference screens in Drosophila tissue culture cells." Methods Enzymol 392 (2005): 55-73.
PMID
15644175
Source
pubmed
Published In
Methods in Enzymology
Volume
392
Publish Date
2005
Start Page
55
End Page
73
DOI
10.1016/S0076-6879(04)92004-6

Increased expression of Drosophila tetraspanin, Tsp68C, suppresses the abnormal proliferation of ytr-deficient and Ras/Raf-activated hemocytes.

Tetraspanins are evolutionary conserved transmembrane proteins thought to facilitate cell proliferation, movement or fusion by acting as organizers of different signaling events. Despite their prevalence and conservation, their specific role and functions remain largely elusive, as their redundancy in various organisms has hindered loss of function studies. Here, we take a gain of function approach to study Drosophila tetraspanin Tsp68C and its effect on larval hemocytes. We recently characterized a lethal mutation in ytr, a conserved gene that encodes a nuclear arginine-rich protein of unknown function, which is accompanied by abnormal differentiation and proliferation of the larval hematopoietic tissue in flies. A hemolectin (hml)-Gal4 construct carried by hml-Gal4 transgenic flies was sufficient by itself to abrogate the hematopoietic defects in ytr mutant larvae. This rescue correlated with the overexpression of tsp68C, a tetraspanin gene nested in the hml promoter. The suppression of abnormal proliferation by the hml-Gal4 construct was not restricted to ytr-deficient hemocytes, but was also observed in hemocytes expressing the oncogenic forms of Raf or Ras proteins. However, it had no effect on overproliferation mediated by a constitutively active form of Jak. New hml-Gal4 lines, in which the tsp68C gene was silenced or deleted from the promoter, no longer rescued the hematopoietic defect in ytr mutants nor suppressed the activated Raf-induced overproliferation. Therefore, change in tetraspanin Tsp68C expression has a strong suppressor effect on abnormal proliferation and differentiation of hemocytes in the context of specific lesions, such as overactivation of the Ras/Raf/MAPK pathway.

Authors
Sinenko, SA; Mathey-Prevot, B
MLA Citation
Sinenko, SA, and Mathey-Prevot, B. "Increased expression of Drosophila tetraspanin, Tsp68C, suppresses the abnormal proliferation of ytr-deficient and Ras/Raf-activated hemocytes." Oncogene 23.56 (December 2, 2004): 9120-9128.
PMID
15480416
Source
pubmed
Published In
Oncogene: Including Oncogene Reviews
Volume
23
Issue
56
Publish Date
2004
Start Page
9120
End Page
9128
DOI
10.1038/sj.onc.1208156

Yantar, a conserved arginine-rich protein is involved in Drosophila hemocyte development.

To identify novel factors involved in Drosophila hematopoiesis, we screened a collection of lethal recessive mutations that also affected normal hemocyte composition in larvae. We present the characterization of the gene yantar (ytr) for which we isolated null and hypomorphic mutations that were associated with severe defects in hemocyte differentiation and proliferation; ytr is predominantly expressed in the hematopoietic tissue during larval development and encodes an evolutionary conserved protein which is predominantly localized in the nucleus. The hematopoietic phenotype in ytr mutants is consistent with a defect or block in differentiation of precursor hemocytes: mutant larvae have enlarged lymph glands (LGs) and have an excess of circulating hemocytes. In addition, many cells exhibit both lamellocyte and crystal cell markers. Ytr function has been preserved in evolution as hematopoietic specific expression of the Drosophila or mouse Ytr proteins rescue the differentiation defects in mutant hemocytes.

Authors
Sinenko, SA; Kim, EK; Wynn, R; Manfruelli, P; Ando, I; Wharton, KA; Perrimon, N; Mathey-Prevot, B
MLA Citation
Sinenko, SA, Kim, EK, Wynn, R, Manfruelli, P, Ando, I, Wharton, KA, Perrimon, N, and Mathey-Prevot, B. "Yantar, a conserved arginine-rich protein is involved in Drosophila hemocyte development." Dev Biol 273.1 (September 1, 2004): 48-62.
PMID
15302597
Source
pubmed
Published In
Developmental Biology
Volume
273
Issue
1
Publish Date
2004
Start Page
48
End Page
62
DOI
10.1016/j.ydbio.2004.05.022

Signaling role of hemocytes in Drosophila JAK/STAT-dependent response to septic injury.

To characterize the features of JAK/STAT signaling in Drosophila immune response, we have identified totA as a gene that is regulated by the JAK/STAT pathway in response to septic injury. We show that septic injury triggers the hemocyte-specific expression of upd3, a gene encoding a novel Upd-like cytokine that is necessary for the JAK/STAT-dependent activation of totA in the Drosophila counterpart of the mammalian liver, the fat body. In addition, we demonstrate that totA activation also requires the NF-KB-like Relish pathway, indicating that fat body cells integrate the activity of NF-KB and JAK/STAT signaling pathways upon immune response. This study reveals that, in addition to the pattern recognition receptor-mediated NF-KB-dependent immune response, Drosophila undergoes a complex systemic response that is mediated by the production of cytokines in blood cells, a process that is similar to the acute phase response in mammals.

Authors
Agaisse, H; Petersen, UM; Boutros, M; Mathey-Prevot, B; Perrimon, N
MLA Citation
Agaisse, H, Petersen, UM, Boutros, M, Mathey-Prevot, B, and Perrimon, N. "Signaling role of hemocytes in Drosophila JAK/STAT-dependent response to septic injury." Dev Cell 5.3 (September 2003): 441-450.
PMID
12967563
Source
pubmed
Published In
Developmental Cell
Volume
5
Issue
3
Publish Date
2003
Start Page
441
End Page
450

Differential requirement for STAT by gain-of-function and wild-type receptor tyrosine kinase Torso in Drosophila.

Malignant transformation frequently involves aberrant signaling from receptor tyrosine kinases (RTKs). These receptors commonly activate Ras/Raf/MEK/MAPK signaling but when overactivated can also induce the JAK/STAT pathway, originally identified as the signaling cascade downstream of cytokine receptors. Inappropriate activation of STAT has been found in many human cancers. However, the contribution of the JAK/STAT pathway in RTK signaling remains unclear. We have investigated the requirement of the JAK/STAT pathway for signaling by wild-type and mutant forms of the RTK Torso (Tor) using a genetic approach in Drosophila. Our results indicate that the JAK/STAT pathway plays little or no role in signaling by wild-type Tor. In contrast, we find that STAT, encoded by marelle (mrl; DStat92E), is essential for the gain-of-function mutant Tor (Tor(GOF)) to activate ectopic gene expression. Our findings indicate that the Ras/Raf/MEK/MAPK signaling pathway is sufficient to mediate the normal functions of wild-type RTK, whereas the effects of gain-of-function mutant RTK additionally require STAT activation.

Authors
Li, WX; Agaisse, H; Mathey-Prevot, B; Perrimon, N
MLA Citation
Li, WX, Agaisse, H, Mathey-Prevot, B, and Perrimon, N. "Differential requirement for STAT by gain-of-function and wild-type receptor tyrosine kinase Torso in Drosophila." Development 129.18 (September 2002): 4241-4248.
PMID
12183376
Source
pubmed
Published In
Development (Cambridge)
Volume
129
Issue
18
Publish Date
2002
Start Page
4241
End Page
4248

SOCS36E, a novel Drosophila SOCS protein, suppresses JAK/STAT and EGF-R signalling in the imaginal wing disc.

We have cloned a novel SOCS gene from Drosophila, socs36E, which is most homologous to the mammalian socs-5 gene. Socs36E is expressed zygotically, predominantly during embryogenesis, in a highly dynamic pattern. In vivo expression of SOCS36E in transgenic flies results in several adult phenotypes. Engrailed-GAL4 directed expression causes loss of the wing anterior cross vein, humeral outgrowths, absence of halteres and eye pigmentation defects. Expression of SOCS36E under apterous-GAL4 control resulted in outstretched wings. Full penetrance of these phenotypes required the presence of the SH2 and SOCS-box domains of SOCS36E. The observed phenotypes were consistent with defects in JAK/STAT or EGF-R signalling and were exacerbated in flies heterozygous for either the d-jak (hopscotch), d-stat (stat92E) or d-egf-r (der) genes. Conversely, inactivating one copy of the d-cbl gene, a negative regulator of the d-EGF-R, partially rescued the wing phenotypes. These genetic interactions imply that SOCS36E can suppress activities of the JAK/STAT and EGF-R signalling pathways in the wing disc and suggest that SOCS36E interacts with multiple pathways in vivo.

Authors
Callus, BA; Mathey-Prevot, B
MLA Citation
Callus, BA, and Mathey-Prevot, B. "SOCS36E, a novel Drosophila SOCS protein, suppresses JAK/STAT and EGF-R signalling in the imaginal wing disc." Oncogene 21.31 (July 18, 2002): 4812-4821.
PMID
12101419
Source
pubmed
Published In
Oncogene: Including Oncogene Reviews
Volume
21
Issue
31
Publish Date
2002
Start Page
4812
End Page
4821
DOI
10.1038/sj.onc.1205618

Functional genomic analysis of phagocytosis and identification of a Drosophila receptor for E. coli.

The recognition and phagocytosis of microbes by macrophages is a principal aspect of innate immunity that is conserved from insects to humans. Drosophila melanogaster has circulating macrophages that phagocytose microbes similarly to mammalian macrophages, suggesting that insect macrophages can be used as a model to study cell-mediated innate immunity. We devised a double-stranded RNA interference-based screen in macrophage-like Drosophila S2 cells, and have defined 34 gene products involved in phagocytosis. These include proteins that participate in haemocyte development, vesicle transport, actin cytoskeleton regulation and a cell surface receptor. This receptor, Peptidoglycan recognition protein LC (PGRP-LC), is involved in phagocytosis of Gram-negative but not Gram-positive bacteria. Drosophila humoral immunity also distinguishes between Gram-negative and Gram-positive bacteria through the Imd and Toll pathways, respectively; however, a receptor for the Imd pathway has not been identified. Here we show that PGRP-LC is important for antibacterial peptide synthesis induced by Escherichia coli both in vitro and in vivo. Furthermore, totem mutants, which fail to express PGRP-LC, are susceptible to Gram-negative (E. coli), but not Gram-positive, bacterial infection. Our results demonstrate that PGRP-LC is an essential component for recognition and signalling of Gram-negative bacteria. Furthermore, this functional genomic approach is likely to have applications beyond phagocytosis.

Authors
Rämet, M; Manfruelli, P; Pearson, A; Mathey-Prevot, B; Ezekowitz, RAB
MLA Citation
Rämet, M, Manfruelli, P, Pearson, A, Mathey-Prevot, B, and Ezekowitz, RAB. "Functional genomic analysis of phagocytosis and identification of a Drosophila receptor for E. coli." Nature 416.6881 (April 11, 2002): 644-648.
PMID
11912489
Source
pubmed
Published In
Nature
Volume
416
Issue
6881
Publish Date
2002
Start Page
644
End Page
648
DOI
10.1038/nature735

Hydrophobic residues Phe751 and Leu753 are essential for STAT5 transcriptional activity.

One facet of cytokine signaling is relayed to the nucleus by the activation, through tyrosine phosphorylation, of latent cytoplasmic signal transducers and activators of transcription (STAT) family members. It has been demonstrated that the C termini of STATs contain the transactivation domain and are essential for the transactivation of target genes. To better understand the function of the STAT C terminus, we have generated a series of C-terminal mutants in STAT5a and examined their effects on transactivation, tyrosine phosphorylation, and DNA binding. Using GAL4 chimerae with the C terminus of STAT5, we have defined a 12-amino acid region essential for STAT5 transactivation. Surprisingly, deletion of these 12 amino acids in the context of the native STAT5 backbone preserved the overall transcriptional activity of the protein. Further analysis revealed that deletion of this region resulted in hyper-DNA binding activity, thus compensating for the weakened transactivation domain. Using site-directed mutagenesis, we show that within this 12-amino acid region the acidic residues were non-essential for transactivation. In contrast, the non-acidic residues were crucial for transactivation. Mutating either Phe(751) or Leu(753) to alanine abolished transactivation suggesting that these residues were essential for connecting STAT5 to the basal transcriptional machinery.

Authors
Callus, BA; Mathey-Prevot, B
MLA Citation
Callus, BA, and Mathey-Prevot, B. "Hydrophobic residues Phe751 and Leu753 are essential for STAT5 transcriptional activity." J Biol Chem 275.22 (June 2, 2000): 16954-16962.
PMID
10748177
Source
pubmed
Published In
The Journal of biological chemistry
Volume
275
Issue
22
Publish Date
2000
Start Page
16954
End Page
16962
DOI
10.1074/jbc.M909976199

Rapid selection of tetracycline-controlled inducible cell lines using a green fluorescent-transactivator fusion protein.

We describe a modification of the tetracycline-controlled expression system that facilitates the rapid identification of tetracycline-sensitive clones. The TetR/VP16 transactivator protein was tagged with the green fluorescent protein (GFP) at its N-terminus. This results in a functional transactivator which allows cells expressing high levels of the modified transactivator to be selected by fluorescent-activated cell sorting. After expansion, single cell clones that maintain a high level of GFP fluorescence can be tested for their ability to transactivate a luciferase gene under control of the Tet operator, leading to the rapid identification of clones with strong inducibility.

Authors
Callus, BA; Mathey-Prevot, B
MLA Citation
Callus, BA, and Mathey-Prevot, B. "Rapid selection of tetracycline-controlled inducible cell lines using a green fluorescent-transactivator fusion protein." Biochem Biophys Res Commun 257.3 (April 21, 1999): 874-878.
PMID
10208877
Source
pubmed
Published In
Biochemical and Biophysical Research Communications
Volume
257
Issue
3
Publish Date
1999
Start Page
874
End Page
878
DOI
10.1006/bbrc.1999.0558

Interleukin-3-induced activation of the JAK/STAT pathway is prolonged by proteasome inhibitors.

One facet of cytokine receptor signaling involves the activation of signal transducers and activators of transcription (STATs). STATs are rapidly activated via tyrosine phosphorylation by Janus kinase (JAK) family members and subsequently inactivated within a short period. We investigated the effect of proteasome inhibition on interleukin-3 (IL-3) activation of the JAK/STAT pathway following stimulation of Ba/F3 cells. Treatment of Ba/F3 cells with the proteasome inhibitor, N-acetyl-L-leucinyl-L-leucinyl-norleucinal (LLnL), led to stable tyrosine phosphorylation of the IL-3 receptor, beta common (betac), and STAT5 following stimulation. The effects of LLnL were not restricted to the JAK/STAT pathway, as Shc and mitogen-activated protein kinase (MAPK) phosphorylation were also prolonged in LLnL-treated cells. Further investigation showed these stable phosphorylation events were the result of prolonged activation of JAK2 and JAK1. These observations were confirmed using pharmacologic inhibitors. In the presence of LLnL, stable phosphorylation of STAT5 and betac was abrogated if the tyrosine kinase inhibitor, staurosporine, was added. The effect of staurosporine on STAT5 phosphorylation could be overcome if the phosphatase inhibitor, vanadate, was also added, suggesting phosphorylated STAT5 could be stabilized by phosphatase, but not by proteasome inhibition per se. These observations are consistent with the hypothesis that proteasome-mediated protein degradation can modulate the activity of the JAK/STAT pathway by regulating the deactivation of JAK.

Authors
Callus, BA; Mathey-Prevot, B
MLA Citation
Callus, BA, and Mathey-Prevot, B. "Interleukin-3-induced activation of the JAK/STAT pathway is prolonged by proteasome inhibitors." Blood 91.9 (May 1, 1998): 3182-3192.
PMID
9558373
Source
pubmed
Published In
Blood
Volume
91
Issue
9
Publish Date
1998
Start Page
3182
End Page
3192

Mammalian and Drosophila blood: JAK of all trades?

Authors
Mathey-Prevot, B; Perrimon, N
MLA Citation
Mathey-Prevot, B, and Perrimon, N. "Mammalian and Drosophila blood: JAK of all trades?." Cell 92.6 (March 20, 1998): 697-700. (Review)
PMID
9529244
Source
pubmed
Published In
Cell
Volume
92
Issue
6
Publish Date
1998
Start Page
697
End Page
700

JAK2 is required for induction of the murine DUB-1 gene.

Cytokine receptors activate multiple signal transduction pathways, resulting in the induction of specific target genes. We have recently identified a hematopoietic cell-specific immediate-early gene, DUB-1, that encodes a growth-regulatory deubiquitinating enzyme. The DUB-1 gene contains a 112-bp enhancer element that is specifically induced by the beta c subunit of the interleukin-3 (IL-3) receptor. To investigate the mechanism of DUB-1 induction, we examined the effects of dominant-negative forms of JAK kinases, STAT transcription factors, and Raf-1 in transient transfection assays. In Ba/F3 cells, IL-3 induced a dose-dependent activation of DUB-1-luciferase (luc) and GAS-luc reporter constructs. A dominant-negative form of JAK2 (truncated at amino acid 829) inhibited the induction of DUB-1-luc and GAS-luc by IL-3. A dominant-negative form of STAT5 (truncated at amino acid 650) inhibited the induction of GAS-luc but not DUB-1-luc. A dominant-negative form of Raf-1 inhibited the induction of DUB-1-luc but had no effect on the induction of GAS-luc by IL-3. The requirement for JAK2 in the stimulation of the DUB-1 enhancer was further supported by the suppression of DUB-1 induction in Ba/F3 cells stably expressing the dominant-negative JAK2 polypeptide. We hypothesize that IL-3 activates a JAK2/Raf-1 signaling pathway that is required for DUB-1 induction and is independent of STAT5.

Authors
Jaster, R; Zhu, Y; Pless, M; Bhattacharya, S; Mathey-Prevot, B; D'Andrea, AD
MLA Citation
Jaster, R, Zhu, Y, Pless, M, Bhattacharya, S, Mathey-Prevot, B, and D'Andrea, AD. "JAK2 is required for induction of the murine DUB-1 gene." Mol Cell Biol 17.6 (June 1997): 3364-3372.
PMID
9154835
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
17
Issue
6
Publish Date
1997
Start Page
3364
End Page
3372

Receptors that induce erythroid differentiation of Ba/F3 cells: structural requirements and effect on STAT5 binding.

Ectopic expression of the erythropoietin receptor (EpoR) in the interleukin-3 (IL-3)-dependent cell line Ba/F3 results in growth and partial erythroid differentiation in Epo. In contrast, introduction and activation of the interleukin-5 receptor (IL-5R) or of the granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) results in proliferation only. As this effect is specific to the EpoR, the role of its extracellular or cytoplasmic domain in differentiation was tested after construction of two chimeric receptors. One receptor contained the extracellular domain of EpoR fused to the endodomain of IL-3R beta-chain (E/beta), while the other contained the EpoR cytoplasmic region fused to the extracellular domain of GM-CSFR alpha-chain (GMER). Surprisingly, both receptors induced differentiation ruling out a strict specificity of the extracellular or cytoplasmic region of EpoR in this process. Instead the ability to signal differentiation correlated with structural features shared by the EpoR, GMER, and E/beta receptors. Dimerization of all three receptors results in the pairing of two signal transducing chains in the cytoplasm, in contrast to the mitogenic receptors IL-3R, IL-5R, GM-CSFR, which assemble as alphabeta heterodimers. Two new chimeric receptors that fulfilled the structural requirement exemplified by EpoR, but lacked any part of EpoR, were designed to consolidate this model. They consisted of the ectodomains of the GMR-alpha and IL-5R alpha, respectively, fused to the endodomain of IL-3R beta-chain. Both receptors were as effective as EpoR in signaling differentiation in response to their cognate ligand. Another property of receptors fulfilling these structural requirements is that they cause a marked delay in signal transducers and activators of transcription 5 (STAT5) activation on ligand stimulation. Taken together our studies show that structural assembly of receptors dictates their potential to signal erythroid differentiation in Ba/F3 cells, that differentiation can take place in the absence of Epo and that a delay in STAT5 activation is highly predictive of this process.

Authors
Pless, M; Norga, K; Carroll, M; Heim, MH; D'Andrea, AD; Mathey-Prevot, B
MLA Citation
Pless, M, Norga, K, Carroll, M, Heim, MH, D'Andrea, AD, and Mathey-Prevot, B. "Receptors that induce erythroid differentiation of Ba/F3 cells: structural requirements and effect on STAT5 binding." Blood 89.9 (May 1, 1997): 3175-3185.
PMID
9129020
Source
pubmed
Published In
Blood
Volume
89
Issue
9
Publish Date
1997
Start Page
3175
End Page
3185

The murine DUB-1 gene is specifically induced by the betac subunit of interleukin-3 receptor.

Cytokines regulate cell growth and differentiation by inducing the expression of specific target genes. We have recently isolated a cytokine-inducible, immediate-early cDNA, DUB-1, that encodes a deubiquitinating enzyme. The DUB-1 mRNA was specifically induced by the receptors for interleukin-3, granulocyte-macrophage colony-stimulating factor, and interleukin-5, suggesting a role for the beta common (betac subunit known to be shared by these receptors. In order to identify the mechanism of cytokine induction, we isolated a murine genomic clone for DUB-1 containing a functional promoter region. The DUB-1 gene contains two exons, and the nucleotide sequence of its coding region is identical to the sequence of DUB-1 cDNA. Various regions of the 5' flanking region of the DUB-1 gene were assayed for cytokine-inducible activity. An enhancer region that retains the beta c-specific inducible activity of the DUB-1 gene was identified. Enhancer activity was localized to a 112-bp fragment located 1.4 kb upstream from the ATG start codon. Gel mobility shift assays revealed two specific protein complexes that bound to this minimal enhancer region. One complex was induced by betac signaling, while the other was noninducible. Finally, the membrane-proximal region of human betac was required for DUB-1 induction. In conclusion, DUB-1 is the first example of an immediate-early gene that is induced by a specific subunit of a cytokine receptor. Further analysis of the DUB-1 enhancer element may reveal specific determinants of a betac-specific signaling pathway.

Authors
Zhu, Y; Pless, M; Inhorn, R; Mathey-Prevot, B; D'Andrea, AD
MLA Citation
Zhu, Y, Pless, M, Inhorn, R, Mathey-Prevot, B, and D'Andrea, AD. "The murine DUB-1 gene is specifically induced by the betac subunit of interleukin-3 receptor." Mol Cell Biol 16.9 (September 1996): 4808-4817.
PMID
8756639
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
16
Issue
9
Publish Date
1996
Start Page
4808
End Page
4817

Cooperation between core binding factor and adjacent promoter elements contributes to the tissue-specific expression of interleukin-3.

Tissue-specific expression of interleukin-3 (IL-3) is mediated via cis-acting elements located within 315 base pairs of the transcription start. This is achieved in part through the positive activities of the AP-1 and Elf-1 sites in the IL-3 promoter. The contribution to T cell-specific expression by other promoter sites was assessed in a transient expression assay with IL-3 promoter constructs linked to a luciferase gene, focusing initially on the core binding factor (CBF) site, which is footprinted in vivo upon T cell activation. Activity of the CBF site is shown to be critically dependent on the adjacent activator site Act-1. Together the Act-1 and CBF sites form a functional unit (AC unit) with dual activity. The AC unit is demonstrated to enhance basal activity of promoters both in fibroblasts and T cells. This activity is further inducible in activated T cells, but not in fibroblasts. In addition to the already identified NIP repressor site, evidence is presented for a second repressor region that restricts promoter activity in fibroblasts. Finally, a novel positive regulatory element has been mapped in the IL-3 promoter between nucleotide -180 and -210 that leads to increased expression in T cells. Together these results demonstrate that T cell expression of IL-3 is not specified by the activity of a single tissue-specific element, but instead involves multiple interacting elements that provide both specific positive regulation in T cells and specific negative regulation in fibroblasts.

Authors
Taylor, DS; Laubach, JP; Nathan, DG; Mathey-Prevot, B
MLA Citation
Taylor, DS, Laubach, JP, Nathan, DG, and Mathey-Prevot, B. "Cooperation between core binding factor and adjacent promoter elements contributes to the tissue-specific expression of interleukin-3." J Biol Chem 271.24 (June 14, 1996): 14020-14027.
PMID
8662845
Source
pubmed
Published In
The Journal of biological chemistry
Volume
271
Issue
24
Publish Date
1996
Start Page
14020
End Page
14027

Multiple proteins interact with the nuclear inhibitory protein repressor element in the human interleukin-3 promoter.

T cell expression of interleukin 3 (IL-3) is directed by positive and negative cis-acting DNA elements clustered within 300 base pairs of the transcriptional start site. A strong repressor element, termed nuclear inhibitory protein (NIP), was previously mapped to a segment of the IL-3 promoter between nucleotides -271 and -250. Functional characterization of this element demonstrates that it can mediate repression when linked in cis to a heterologous promoter. DNA binding experiments were carried out to characterize the repressor activity. Using varying conditions, three distinct complexes were shown to interact specifically with the NIP region, although only one correlates with repressor activity. Complex 1 results from binding of a ubiquitous polypeptide that recognizes the 3' portion of this sequence and is not required for repression. Complex 2 corresponds to binding of transcription factor (upstream stimulatory factor) to an E-box motif in the 5' portion of the NIP region. DNA binding specificity of complex 3 overlaps with that of upstream stimulatory factor but is clearly distinct. To determine which of the latter two complexes represents NIP activity, we incorporated small alterations into the NIP site of an IL-3 promoter-linked reporter construct and examined their effects on NIP-mediated repression. Functional specificity for repression matches the DNA binding specificity of complex 3; both repressor activity and complex 3 binding require the consensus sequence CTCACNTNC.

Authors
Engeland, K; Andrews, NC; Mathey-Prevot, B
MLA Citation
Engeland, K, Andrews, NC, and Mathey-Prevot, B. "Multiple proteins interact with the nuclear inhibitory protein repressor element in the human interleukin-3 promoter." J Biol Chem 270.41 (October 13, 1995): 24572-24579.
PMID
7592676
Source
pubmed
Published In
The Journal of biological chemistry
Volume
270
Issue
41
Publish Date
1995
Start Page
24572
End Page
24579

Multiple transcription start sites and 5' alternate splicing of murine IL-3 receptor beta-chain transcripts.

The murine interleukin-3 receptor beta-chain genes, IL-3R beta IL-3 and IL-3R beta c, encode the signal transducing chains of the high affinity receptors for IL-3 and IL-3, GM-CSF and IL-5 respectively. Little is known about the regulation of their expression. To enable the study of the promotors of IL-3R beta IL-3 and IL-3R beta c, we have characterized their respective 5' untranslated regions using a modified 5' RACE protocol. Four classes of alternatively spliced transcripts were isolated that initiate in a 400 nt region upstream from a previously reported start site(1). The initially reported partial IL-3R beta IL-3 clone belongs to the first class of transcripts(2). The second class starts in the middle of an intron as defined by the first class. The 3rd and 4th class establish 2 novel splice donor sites. These results were confirmed by RNAse-protection assay. The complex organization as evident from our data establishes an experimental framework for future experiments aimed at the study of the promoters for the murine IL-3R beta genes.

Authors
Norga, K; Pless, M; Stanulla, M; TePas, EC; Mathey-Prevot, B
MLA Citation
Norga, K, Pless, M, Stanulla, M, TePas, EC, and Mathey-Prevot, B. "Multiple transcription start sites and 5' alternate splicing of murine IL-3 receptor beta-chain transcripts." Biochem Biophys Res Commun 205.1 (November 30, 1994): 886-892.
PMID
7999127
Source
pubmed
Published In
Biochemical and Biophysical Research Communications
Volume
205
Issue
1
Publish Date
1994
Start Page
886
End Page
892

Differentiation domains of the erythropoietin receptor.

Ectopic expression of the erythropoietin receptor (Epo-R) in Ba/F3, an interleukin-3 (IL-3)-dependent progenitor cell line, confers both Epo-dependent cell growth and Epo-dependent induction of beta-globin mRNA. We have used this system of limited erythroid differentiation to characterize the role of the Epo-R in differentiation. In particular, we have been interested in identifying a differentiation domain of the Epo-R. We have studied three chimeras encoding regions of the extracellular region of the Epo-R and the intracellular region of the IL-3R beta IL-3. After transfection into Ba/F3 cells, all three chimeras conferred Epo-dependent growth and induced the expression of beta-globin, suggesting that the extracellular region of the Epo-R plays a critical role in differentiation. However, a truncated Epo-R containing only the extracellular region of the Epo-R and a 15 amino acid cytoplasmic tail does not induced beta-globin expression, although it is processed to the cell surface and binds Epo. These experiments show that the extracytoplasmic region of the Epo-R is necessary but not sufficient to induce erythroid-specific differentiation in this system.

Authors
Carroll, M; Mathey-Prevot, B; D'Andrea, A
MLA Citation
Carroll, M, Mathey-Prevot, B, and D'Andrea, A. "Differentiation domains of the erythropoietin receptor." Proc Soc Exp Biol Med 206.3 (July 1994): 289-294.
PMID
8016166
Source
pubmed
Published In
Experimental Biology and Medicine
Volume
206
Issue
3
Publish Date
1994
Start Page
289
End Page
294

Identification of a critical regulatory site in the human interleukin-3 promoter by in vivo footprinting.

Interleukin-3 (IL-3) is involved in proliferation and differentiation of hematopoietic progenitor cells. Its expression is subject to precise, tissue-specific regulation, and has been studied extensively in the gibbon T-cell line MLA 144 by a combination of functional assays and DNA binding experiments. To extend these studies, the gibbon IL-3 promoter was cloned and in vivo footprinting of the gibbon and human IL-3 proximal promoters was performed. The gibbon IL-3 promoter was found to be highly homologous to its human counterpart and both promoters yielded identical in vivo footprints after induction of IL-3 synthesis. In particular, we observed specific protection of three guanines over a core sequence TGTGGTTT (IF-1IL3) that had not been recognized in previous studies. IF-1IL3 is not found in other cytokine promoters, but it is conserved in the IL-3 promoter of several species and is similar to a recurring motif in viral and T-cell-specific cellular enhancers. IF-1IL3 binds a specific complex in MLA 144 and Jurkat nuclear extracts in vitro, which shares the same specificity as the complex bound by the polyoma virus and T-cell receptor delta enhancers. Mutation of the three guanines in IF-1IL3 core sequence disrupts binding in vitro and abrogates the ability of the IL-3 promoter to mediate inducible expression in T cells. Although IF-1IL3 is necessary for IL-3 expression, it is not sufficient: a truncated IL-3 promoter with an intact IF-1IL3 site but no other activator sites is transcriptionally silent. These studies describe a new regulatory element within the IL-3 promoter that is essential for expression and conserved between species.

Authors
Cameron, S; Taylor, DS; TePas, EC; Speck, NA; Mathey-Prevot, B
MLA Citation
Cameron, S, Taylor, DS, TePas, EC, Speck, NA, and Mathey-Prevot, B. "Identification of a critical regulatory site in the human interleukin-3 promoter by in vivo footprinting." Blood 83.10 (May 15, 1994): 2851-2859.
PMID
8180380
Source
pubmed
Published In
Blood
Volume
83
Issue
10
Publish Date
1994
Start Page
2851
End Page
2859

Erythropoietin receptor contains both growth-promoting activity and differentiation-promoting activity.

Authors
DeMartino, J; Carroll, M; Mathey-Prevot, B; D'Andrea, AD
MLA Citation
DeMartino, J, Carroll, M, Mathey-Prevot, B, and D'Andrea, AD. "Erythropoietin receptor contains both growth-promoting activity and differentiation-promoting activity." Ann N Y Acad Sci 718 (April 15, 1994): 213-221. (Review)
PMID
8185230
Source
pubmed
Published In
Annals of the New York Academy of Sciences
Volume
718
Publish Date
1994
Start Page
213
End Page
221

Erythropoietin receptor signals both proliferation and erythroid-specific differentiation.

Ectopic expression of the erythropoietin receptor (EPO-R) in Ba/F3, an interleukin 3-dependent progenitor cell line, confers EPO-dependent cell growth. To examine whether the introduced EPO-R could affect differentiation, we isolated Ba/F3-EPO-R subclones in interleukin 3 and assayed for the induction of beta-globin mRNA synthesis after exposure to EPO. Detection of beta-globin mRNA was observed within 3 days of EPO treatment, with peak levels accumulating after 10 days. When EPO was withdrawn, expression of beta-globin mRNA persisted in most clones, suggesting that commitment to erythroid differentiation had occurred. Although EPO-R expression also supports EPO-dependent proliferation of CTLL-2, a mature T-cell line, those cells did not produce globin transcripts, presumably because they lack requisite cellular factors involved in erythrocyte differentiation. We conclude that the EPO-R transmits signals important for both proliferation and differentiation along the erythroid lineage.

Authors
Liboi, E; Carroll, M; D'Andrea, AD; Mathey-Prevot, B
MLA Citation
Liboi, E, Carroll, M, D'Andrea, AD, and Mathey-Prevot, B. "Erythropoietin receptor signals both proliferation and erythroid-specific differentiation." Proc Natl Acad Sci U S A 90.23 (December 1, 1993): 11351-11355.
PMID
8248252
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
90
Issue
23
Publish Date
1993
Start Page
11351
End Page
11355

Interleukin-3 expression by activated T cells involves an inducible, T-cell-specific factor and an octamer binding protein.

Interleukin-3 (IL-3) is exclusively expressed by activated T and natural killer cells, a function that is tightly controlled both in a lineage-specific and in a stimulation-dependent manner. We have investigated the protein binding characteristics and functional importance of the ACT-1-activating region of the IL-3 promoter. This region binds an inducible, T-cell-specific factor over its 5' end, a site that is necessary for the expression of IL-3 in the absence of other upstream elements. Over its 3' end, it binds a factor that is ubiquitously and constitutively expressed. This factor is Oct-1 or an immunologically related octamer-binding protein, and it plays a role in coordinating the activity of several regulatory elements. These characteristics make the ACT-1 site analogous to the activating ARRE-1 site in the IL-2 promoter. Furthermore, and despite a lack of sequence homology, the promoters of IL-3 and IL-2 share an organizational pattern of regulatory elements that is likely to be important for the T-cell-specific expression of these genes.

Authors
Davies, K; TePas, EC; Nathan, DG; Mathey-Prevot, B
MLA Citation
Davies, K, TePas, EC, Nathan, DG, and Mathey-Prevot, B. "Interleukin-3 expression by activated T cells involves an inducible, T-cell-specific factor and an octamer binding protein." Blood 81.4 (February 15, 1993): 928-934.
PMID
8428000
Source
pubmed
Published In
Blood
Volume
81
Issue
4
Publish Date
1993
Start Page
928
End Page
934

Enhanced expression of interleukin-3 and granulocyte-macrophage colony-stimulating factor receptor subunits in murine hematopoietic cells stimulated with hematopoietic growth factors.

AIC2A and AIC2B are closely related genes encoding components of the receptors for murine interleukin-3 (IL-3) (AIC2A) and granulocyte-macrophage colony-stimulating factor (GM-CSF) and IL-5 (AIC2B). We have studied the parallel regulation of expression of these genes in erythroid and myeloid progenitor cell lines. AIC2A and AIC2B transcription was transiently induced in these cells in response to a variety of hematopoietic growth factors, including erythropoietin (EPO), monocyte-CSF, IL-3, GM-CSF, and stem cell factor (SCF or kit ligand). Run-on assays established that the increase occurred mainly at the transcriptional level. Immunoprecipitation experiments confirmed that the increase in messenger RNA expression resulted in augmented synthesis of both AIC2A and AIC2B proteins, and binding studies further showed these proteins to be functional. We observed a fourfold increase in low-affinity IL-3 sites in an erythroid precursor cell line stimulated with EPO, and a threefold increase in GM-CSF high-affinity sites in a myeloid cell line stimulated with IL-3. In addition, we showed that the increase in the IL-3 receptor chain AIC2A in the erythroid precursor cell line correlated with the ability of IL-3 to exert a cooperative effect with EPO in the induction of beta-globin in these cells.

Authors
Liboi, E; Jubinsky, P; Andrews, NC; Nathan, DG; Mathey-Prevot, B
MLA Citation
Liboi, E, Jubinsky, P, Andrews, NC, Nathan, DG, and Mathey-Prevot, B. "Enhanced expression of interleukin-3 and granulocyte-macrophage colony-stimulating factor receptor subunits in murine hematopoietic cells stimulated with hematopoietic growth factors." Blood 80.5 (September 1, 1992): 1183-1189.
PMID
1387562
Source
pubmed
Published In
Blood
Volume
80
Issue
5
Publish Date
1992
Start Page
1183
End Page
1189

The erythropoietin receptor transmembrane region is necessary for activation by the Friend spleen focus-forming virus gp55 glycoprotein.

The erythropoietin receptor (EPO-R), a member of the cytokine receptor superfamily, can be activated by binding either erythropoietin (EPO) or gp55, the Friend spleen focus-forming virus glycoprotein. The highly specific interaction between gp55 and EPO-R triggers cell proliferation and thereby causes the first stage of Friend virus-induced erythroleukemia. We have generated functional chimeric receptors containing regions of the EPO-R and the interleukin-3 receptor (AIC2A polypeptide), a related cytokine receptor which does not interact with gp55. All chimeric receptors were expressed at similar levels, had similar binding affinities for EPO, and conferred EPO-dependent cell growth. Only those chimeric receptors which contained the EPO-R transmembrane region were activated by gp55. These results demonstrate that the transmembrane region of the EPO-R is critical for activation by gp55. In addition, analysis of a soluble, secreted EPO-R and cysteine point mutants of the EPO-R show that the extracytoplasmic region of the EPO-R specifically interacts with gp55.

Authors
Zon, LI; Moreau, JF; Koo, JW; Mathey-Prevot, B; D'Andrea, AD
MLA Citation
Zon, LI, Moreau, JF, Koo, JW, Mathey-Prevot, B, and D'Andrea, AD. "The erythropoietin receptor transmembrane region is necessary for activation by the Friend spleen focus-forming virus gp55 glycoprotein." Mol Cell Biol 12.7 (July 1992): 2949-2957.
PMID
1320192
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
12
Issue
7
Publish Date
1992
Start Page
2949
End Page
2957

A functional isoform of the human granulocyte/macrophage colony-stimulating factor receptor has an unusual cytoplasmic domain.

The granulocyte/macrophage colony-stimulating factor (GM-CSF) receptor (GMR) transduces a signal that results in the proliferation, differentiation, and functional activation of hematopoietic cells. This study sought to determine whether functional isoforms of the receptor exist that may be important in generating this diversity of cellular response. We have isolated a cDNA encoding an isoform of the low-affinity human GMR that is a product of alternative splicing of the GMR gene and results in a predicted 410-amino acid protein with a cytoplasmic domain that is rich in serine residues, a feature of regions critical in signal transduction for other receptors of the hematopoietin receptor superfamily. This receptor bound ligand and was functionally active when introduced into a murine factor-dependent cell line; mRNA transcripts representative of this isoform were coexpressed with those for a previously isolated 400-amino acid isoform of the GMR in normal hematopoietic and leukemic cells. In view of the recent isolation of a cDNA, designated GM-CSF R beta, that confers high-affinity binding of GM-CSF in cotransfection experiments with the low-affinity receptor, we suggest that the previously isolated low-affinity receptor be designated GM-CSF R alpha 1 and the one described in this report be designated GM-CSF R alpha 2.

Authors
Crosier, KE; Wong, GG; Mathey-Prevot, B; Nathan, DG; Sieff, CA
MLA Citation
Crosier, KE, Wong, GG, Mathey-Prevot, B, Nathan, DG, and Sieff, CA. "A functional isoform of the human granulocyte/macrophage colony-stimulating factor receptor has an unusual cytoplasmic domain." Proc Natl Acad Sci U S A 88.17 (September 1, 1991): 7744-7748.
PMID
1715577
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
88
Issue
17
Publish Date
1991
Start Page
7744
End Page
7748

Preneoplastic lesions induced by myc and src oncogenes in a heterotopic rat colon.

With the use of viral vectors harboring myc and src oncogenes, we have assessed the potential contribution of these different elements to colonic neoplasia using a transplantation technique resulting in the formation of a heterotopic colon in Wistar Furth rats. While myc alone induced atypia and some dysplasia, src induced focal dysplastic lesions throughout the colon mucosa with evidence of metaplasia. In contrast, lesions induced by myc and src acting cooperatively, were highly dysplastic with evidence of tumor formation after protracted periods. These results indicated the formation of histologically distinct preneoplastic lesions elicited by the action of a single oncogene in colon implants with the production of adenocarcinomas when such oncogenic elements act cooperatively. This model provides an opportunity for studies of the action of different oncogenes, acting singly or in combination, in the multi-step progression to colon tumor formation.

Authors
D'Emilia, JC; Mathey-Prevot, B; Jaros, K; Wolf, B; Steele, G; Summerhayes, IC
MLA Citation
D'Emilia, JC, Mathey-Prevot, B, Jaros, K, Wolf, B, Steele, G, and Summerhayes, IC. "Preneoplastic lesions induced by myc and src oncogenes in a heterotopic rat colon." Oncogene 6.2 (February 1991): 303-309.
PMID
2000223
Source
pubmed
Published In
Oncogene: Including Oncogene Reviews
Volume
6
Issue
2
Publish Date
1991
Start Page
303
End Page
309

Positive and negative elements regulate human interleukin 3 expression.

The human interleukin 3 (IL-3) promoter is comprised of several cis-acting DNA sequences that modulate T-cell expression of IL-3. These are located within 315 nucleotides upstream of the mRNA start site. Transient expression of reporter genes linked to serially deleted sequences of the IL-3 promoter has allowed mapping of two activator sequences and an interposed repressor sequence. The proximal regulatory region is specific to IL-3 and prerequisite for efficient transcription. Its effect is enhanced by a second, more distal activating sequence consisting of an AP-1 binding site. Between the two activators lies a transcriptional silencer, which is a potent repressor in the absence of the AP-1 site. DNA-nuclear protein binding experiments demonstrate specific complex formation within each of these functional regions. Thus, both positive and negative regulatory elements appear to control expression of the human IL-3 gene in activated T cells.

Authors
Mathey-Prevot, B; Andrews, NC; Murphy, HS; Kreissman, SG; Nathan, DG
MLA Citation
Mathey-Prevot, B, Andrews, NC, Murphy, HS, Kreissman, SG, and Nathan, DG. "Positive and negative elements regulate human interleukin 3 expression." Proc Natl Acad Sci U S A 87.13 (July 1990): 5046-5050.
PMID
1695008
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
87
Issue
13
Publish Date
1990
Start Page
5046
End Page
5050

Granulocyte-macrophage colony-stimulating factor and interleukin-3 mRNAs are produced by a small fraction of blood mononuclear cells.

Northern blot analysis has identified granulocyte macrophage colony stimulating factor (GM-CSF) mRNA in monocytes and both GM-CSF and interleukin-3 (IL-3) mRNA in lymphocytes. However, these results have not addressed whether all cells or a subset of the population is capable of hematopoietic growth factor (HGF) production. To resolve this question, we applied in situ hybridization of radiolabeled antisense RNA probes to centrifuged preparations of total blood mononuclear cells (BMCs) and fractionated lymphocyte subpopulations. Without stimulation, no circulating cells expressed detectable levels of GM-CSF or IL-3 mRNA. On stimulation of BMCs with phorbol myristate acetate (PMA) and phytohemagglutinin or PMA and the calcium ionophore ionomycin, approximately 5% expressed GM-CSF mRNA and approximately 1% IL-3 mRNA. Control sense probes produced no labeled cells. To determine the subsets of lymphocytes capable of GM-CSF and IL-3 expression, BMCs were fractionated by FACS into CD8+ and CD4+ lymphocyte subsets and CD16+ (NK) cells. The unfractionated cells and cell fractions were then stimulated with PMA and ionomycin. Results demonstrated that 3% to 5% of the CD16+, CD8+, and CD4+ lymphocytes produced GM-CSF mRNA. However, the number of IL-3 mRNA-positive cells in the FACS-sorted subsets was greatly reduced (0.02% to 0.05%) as compared with the unseparated cells (1%). Treatment of BMCs with high-dose interleukin-2 (IL-2) for 1 week followed by PMA plus ionomycin resulted in a lymphocyte population in which 50% and 3% of cells expressed GM-CSF and IL-3 mRNA, respectively. Thus, GM-CSF and IL-3 mRNA expression in T cells and NK cells is restricted to a small fraction of cells that can be greatly expanded by IL-2 stimulation. These results suggest a possible physiologic mechanism for increasing HGF production by circulating lymphocytes.

Authors
Wimperis, JZ; Niemeyer, CM; Sieff, CA; Mathey-Prevot, B; Nathan, DG; Arceci, RJ
MLA Citation
Wimperis, JZ, Niemeyer, CM, Sieff, CA, Mathey-Prevot, B, Nathan, DG, and Arceci, RJ. "Granulocyte-macrophage colony-stimulating factor and interleukin-3 mRNAs are produced by a small fraction of blood mononuclear cells." Blood 74.5 (October 1989): 1525-1530.
PMID
2676015
Source
pubmed
Published In
Blood
Volume
74
Issue
5
Publish Date
1989
Start Page
1525
End Page
1530

Expression of human interleukin-3 (multi-CSF) is restricted to human lymphocytes and T-cell tumor lines.

While the cellular sources for granulocyte-macrophage colony-stimulating factor (GM-CSF) are known to be widely distributed among several cell types, interleukin-3 (IL-3) gene expression has been demonstrated in only certain T-cell clones and in blood mononuclear cells stimulated with phytohemagglutinin (PHA) and phorbol-myristate-acetate (PMA). To determine which blood cells were responsible for this expression, we fractionated PHA/PMA-stimulated mononuclear cells and identified T lymphocytes as the source of IL-3 mRNA. Low-level IL-3 expression was detected as well in several stimulated human T-cell lines. Hematopoietic stromal cells such as fibroblasts and endothelial cells could not be induced to express IL-3 mRNA. The kinetics of IL-3 mRNA induction in mononuclear cells and lymphocytes stimulated with PHA/PMA or anti-CD3 monoclonal antibody (MoAb) and interleukin-1 (IL-1) were similar to those observed for GM-CSF expression.

Authors
Niemeyer, CM; Sieff, CA; Mathey-Prevot, B; Wimperis, JZ; Bierer, BE; Clark, SC; Nathan, DG
MLA Citation
Niemeyer, CM, Sieff, CA, Mathey-Prevot, B, Wimperis, JZ, Bierer, BE, Clark, SC, and Nathan, DG. "Expression of human interleukin-3 (multi-CSF) is restricted to human lymphocytes and T-cell tumor lines." Blood 73.4 (March 1989): 945-951.
PMID
2645952
Source
pubmed
Published In
Blood
Volume
73
Issue
4
Publish Date
1989
Start Page
945
End Page
951

Regulation of the human interleukin-3 gene.

Authors
Andrews, NC; Mathey-Prevot, B; Murphy, H; Nathan, DG
MLA Citation
Andrews, NC, Mathey-Prevot, B, Murphy, H, and Nathan, DG. "Regulation of the human interleukin-3 gene." Trans Assoc Am Physicians 102 (1989): 240-251.
PMID
2638528
Source
pubmed
Published In
Transactions of the Association of American Physicians
Volume
102
Publish Date
1989
Start Page
240
End Page
251

A human immunoglobulin gene reduces the incidence of lymphomas in c-Myc-bearing transgenic mice.

Transgenic mice carrying an immunoglobulin enhancer-driven c-myc oncogene develop rapid-onset pre-B cell lymphomas. The incidence of these malignancies is greatly reduced when an additional transgene encoding the membrane-bound form (but not the secreted form) of human Ig mu is bred into the susceptible strain. This suppressive effect correlates with a subtle alteration in B-cell development induced by the immunoglobulin transgene.

Authors
Nussenzweig, MC; Schmidt, EV; Shaw, AC; Sinn, E; Campos-Torres, J; Mathey-Prevot, B; Pattengale, PK; Leder, P
MLA Citation
Nussenzweig, MC, Schmidt, EV, Shaw, AC, Sinn, E, Campos-Torres, J, Mathey-Prevot, B, Pattengale, PK, and Leder, P. "A human immunoglobulin gene reduces the incidence of lymphomas in c-Myc-bearing transgenic mice." Nature 336.6198 (December 1, 1988): 446-450.
PMID
3143076
Source
pubmed
Published In
Nature
Volume
336
Issue
6198
Publish Date
1988
Start Page
446
End Page
450
DOI
10.1038/336446a0

Recombinants within the tyrosine kinase region of v-abl and v-src identify a v-abl segment that confers lymphoid specificity.

The v-abl and v-src oncogenes encode protein-tyrosine kinases that possess different biological properties in spite of their high degree of amino acid conservation. To correlate functional differences with structural domains of the two oncogenes, we recombined v-abl and v-src just downstream of the lysines in their ATP-binding sites, within the kinase domain. The biological activity of the chimeric genes was studied and compared with that of v-src and v-abl. The v-src/v-abl recombinant shared with v-src and v-abl the ability to transform fibroblasts. In addition, like v-abl, it transformed lymphoid cells and relieved a hematopoietic cell line of its interleukin 3 requirement. In contrast, the reciprocal construct, v-abl/v-src, was transformation defective. Lack of biological activity correlated with formation of a stable complex between the chimeric protein and two cellular proteins and with low kinase activity. We conclude that the specificity within the kinase domain determines the particular biological behavior of protein-tyrosine kinase oncogenes.

Authors
Mathey-Prevot, B; Baltimore, D
MLA Citation
Mathey-Prevot, B, and Baltimore, D. "Recombinants within the tyrosine kinase region of v-abl and v-src identify a v-abl segment that confers lymphoid specificity." Mol Cell Biol 8.1 (January 1988): 234-240.
PMID
3122023
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
8
Issue
1
Publish Date
1988
Start Page
234
End Page
240

Neuronal pp60c-src contains a six-amino acid insertion relative to its non-neuronal counterpart.

Neuronal cells express a pp60c-src variant that displays an altered electrophoretic mobility and a different V8 peptide pattern relative to pp60c-src expressed in tissues of non-neuronal origin. To determine whether the neuronal form of pp60c-src is encoded by a brain-specific messenger RNA, a mouse brain complementary DNA (cDNA) library was screened with a chicken c-src probe and a 3.8-kilobase c-src cDNA clone was isolated. This clone encodes a 60-kilodalton protein that differs from chicken or human pp60c-src primarily in having six extra amino acids (Arg-Lys-Val-Asp-Val-Arg) within the NH2-terminal 16 kilodaltons of the molecule. S1 nuclease protection analysis confirmed that brain c-src RNA contains an 18-nucleotide insertion at the position of the extra six amino acids. This insertion occurs at a position that corresponds to a splice junction in the chicken and human c-src genes. The isolated c-src cDNA clone encodes a protein that displays an identical V8 peptide pattern to that observed in pp60c-src isolated from tissues of neuronal origin.

Authors
Martinez, R; Mathey-Prevot, B; Bernards, A; Baltimore, D
MLA Citation
Martinez, R, Mathey-Prevot, B, Bernards, A, and Baltimore, D. "Neuronal pp60c-src contains a six-amino acid insertion relative to its non-neuronal counterpart." Science 237.4813 (July 24, 1987): 411-415.
PMID
2440106
Source
pubmed
Published In
Science
Volume
237
Issue
4813
Publish Date
1987
Start Page
411
End Page
415

Abelson virus abrogation of interleukin-3 dependence in a lymphoid cell line.

Among several tyrosine-protein kinases, only v-abl could abrogate interleukin 3 dependence of a lymphoblastoid cell line; v-src and v-fps proteins gave partial or no interleukin 3 independence, respectively. Lymphokine independence was achieved via a nonautocrine mechanism. Direct involvement of c-myc in this process was not evident.

Authors
Mathey-Prevot, B; Nabel, G; Palacios, R; Baltimore, D
MLA Citation
Mathey-Prevot, B, Nabel, G, Palacios, R, and Baltimore, D. "Abelson virus abrogation of interleukin-3 dependence in a lymphoid cell line." Mol Cell Biol 6.11 (November 1986): 4133-4135.
PMID
3025637
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
6
Issue
11
Publish Date
1986
Start Page
4133
End Page
4135

Specific transforming potential of oncogenes encoding protein-tyrosine kinases.

Several chimeric murine retroviruses were constructed to test whether the gag sequence of Abelson murine leukemia virus (A-MuLV) could influence the in vitro specificity of two sarcoma-inducing oncogenes: src of Rous sarcoma virus and fps of Fujinami sarcoma virus. Although the src- or fps- containing chimerae could transform fibroblasts, they were unable to mimic the action of A-MuLV in causing lymphoid transformation in vitro. A-MuLV-derived gag sequences could, however, functionally replace the 5' end of src and restore the transformation potential of a 5'-truncated src gene. To investigate this functional similarity, we replaced the gag sequence of an A-MuLV virus with the 5' end of src. This recombinant virus behaved like the A-MuLV virus from which it was derived: it transformed both fibroblasts and lymphoid cells in vitro. Taken together, these results suggest that lymphoid transformation in vitro is a specific property of abl and not of src or fps. Furthermore, it shows that a functional homology exists between the gag sequence of A-MuLV and the 5' end of src.

Authors
Mathey-Prevot, B; Baltimore, D
MLA Citation
Mathey-Prevot, B, and Baltimore, D. "Specific transforming potential of oncogenes encoding protein-tyrosine kinases." EMBO J 4.7 (July 1985): 1769-1774.
PMID
2992940
Source
pubmed
Published In
EMBO Journal
Volume
4
Issue
7
Publish Date
1985
Start Page
1769
End Page
1774

Preferential expression of the c-fps protein in chicken macrophages and granulocytic cells.

We have studied the expression of the protein kinase activity of NCP98, the c-fps gene product, in several hemopoietic tissues of chickens as a function of the developmental stage of these organs. We found that in bone marrow, spleen, and bursa, maximum NCP98 kinase activity on a per-cell basis correlates with the peak of granulopoiesis in these organs. Furthermore, in a bovine serum albumin density gradient fractionation of bone marrow cells, granulocytic cells appeared to account for most of the NCP98 kinase activity. No correlation was found between the distribution of erythrocytic, lymphocytic, or thrombocytic cells and the distribution of the expression of NCP98 kinase activity. However, NCP98 protein and kinase activity were 10-fold higher in macrophages than in bone marrow. In addition, depletion by complement-mediated lysis of erythrocytic cells in bone marrow did not significantly reduce the total recovery of NCP98 kinase activity. These results argue for the specific expression of the c-fps gene product in granulocytic cells and macrophages.

Authors
Samarut, J; Mathey-Prevot, B; Hanafusa, H
MLA Citation
Samarut, J, Mathey-Prevot, B, and Hanafusa, H. "Preferential expression of the c-fps protein in chicken macrophages and granulocytic cells." Mol Cell Biol 5.5 (May 1985): 1067-1072.
PMID
2987674
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
5
Issue
5
Publish Date
1985
Start Page
1067
End Page
1072

Revertants and partial transformants of rat fibroblasts infected with Fujinami sarcoma virus.

Fifteen revertants were isolated from three independent clones of rat fibroblasts transformed by Fujinami sarcoma virus (FSV). Three revertant clones resulted from the deletion of the one copy of the FSV provirus, and one encoded an enzymatically inactive, transformation-defective protein. The remaining revertant clones were characterized by a transcriptional block of the provirus. Digestion of chromosomal DNA with MspI and HpaII revealed that the FSV provirus was hypermethylated in these revertants, whereas proviral DNA of their spontaneous retransformants was hypomethylated. Furthermore, the revertants had lost DNase I-hypersensitive sites in and around the FSV provirus. The effect of transcriptional regulation of the FSV provirus was further analyzed in clones showing various degrees of phenotypic transformation. We quantitated v-fps mRNA levels in these cells by liquid hybridization and found that increasing levels of viral RNA correlated with a more pronounced transformed phenotype. These results suggest that transcription of FSV proviral DNA is under both viral and cellular control and that transformation by FSV is a function of the dosage of v-fps mRNA.

Authors
Mathey-Prevot, B; Shibuya, M; Samarut, J; Hanafusa, H
MLA Citation
Mathey-Prevot, B, Shibuya, M, Samarut, J, and Hanafusa, H. "Revertants and partial transformants of rat fibroblasts infected with Fujinami sarcoma virus." J Virol 50.2 (May 1984): 325-334.
PMID
6323733
Source
pubmed
Published In
Journal of virology
Volume
50
Issue
2
Publish Date
1984
Start Page
325
End Page
334

A cellular protein is immunologically crossreactive with and functionally homologous to the Fujinami sarcoma virus transforming protein.

We obtained a regressing-tumor antiserum specific for the unique sequence of the transforming protein P140 of Fujinami sarcoma virus by injecting Fischer rats with syngeneic embryo cells transformed with Fujinami sarcoma virus. This serum is capable of immunoprecipitating a protein of 98,000 daltons from cell extracts of normal, uninfected chicken bone marrow cells. This normal cellular protein (NCP98) was shown to be structurally related to P140, sharing the majority of 35S-methionine-labeled tryptic peptides with the viral gene product P140. NCP98 is a phosphoprotein in vivo, with an associated in vitro protein kinase activity, capable of phosphorylating specifically at tyrosine residues of NCP98 itself and alpha-casein, an externally added substrate. This kinase activity is biochemically indistinguishable from the kinase activity associated with P140 by all criteria tested. Moreover, in vitro-phosphorylated NCP98 and P140 shared the same phosphopeptides. The expression of NCP98 is tissue-specific. It is readily detectable in bone marrow cells and detectable to a lesser extent in liver and lung cells from 6--18 day old chickens.

Authors
Mathey-Prevot, B; Hanafusa, H; Kawai, S
MLA Citation
Mathey-Prevot, B, Hanafusa, H, and Kawai, S. "A cellular protein is immunologically crossreactive with and functionally homologous to the Fujinami sarcoma virus transforming protein." Cell 28.4 (April 1982): 897-906.
PMID
7094017
Source
pubmed
Published In
Cell
Volume
28
Issue
4
Publish Date
1982
Start Page
897
End Page
906

Isolation of 16L virus: A rapidly transforming sarcoma virus from an avian leukosis virus-induced sarcoma

We have isolated a replication-defective rapidly transforming sarcoma virus (designated 16L virus) from a fibrosarcoma in a chicken infected with td107A, a transformation-defective deletion mutant of subgroup A Schmidt-Ruppin Rous sarcoma virus. 16L virus transforms fibroblasts and causes sarcomas in infected chickens within 2 wk. Its genomic RNA is 6.0 kilobases and contains sequences homologous to the transforming gene (fps) of Fujinami sarcoma virus (FSV). RNase T1 oligonucleotide analysis shows that the 5' and 3' terminal sequences of 16L virus are indistinguishable from (and presumably derived from) td107A RNA. The central part of 16L viral RNA consists of fps-related sequences. These oligonucleotides fall into four classes: oligonucleotides common to the putative transforming regions of FSV and another fps-containing avian sarcoma virus, URI; an oligonucleotide also present in FSV but not in UR1; an oligonucleotide also present in UR1 but not in FSV; and an oligonucleotide not present in either FSV, UR1, or td107A. Cells infected with 16L virus synthesize a protein of M(r) 142,000 that is immunoprecipitated with anti-gag antiserum. This protein has protein kinase activity. These results suggest that 16L virus arose by recombination between td107A and the cellular fps gene.

Authors
Neel, BG; Wang, LH; Mathey-Prevot, B; Hanafusa, T; Hanafusa, H; Hayward, WS
MLA Citation
Neel, BG, Wang, LH, Mathey-Prevot, B, Hanafusa, T, Hanafusa, H, and Hayward, WS. "Isolation of 16L virus: A rapidly transforming sarcoma virus from an avian leukosis virus-induced sarcoma." Proceedings of the National Academy of Sciences of the United States of America 79.16 I (1982): 5088-5092.
Source
scival
Published In
Proceedings of the National Academy of Sciences of USA
Volume
79
Issue
16 I
Publish Date
1982
Start Page
5088
End Page
5092
DOI
10.1073/pnas.79.16.5088

Mutants of Fujinami sarcoma virus which are temperature sensitive for cellular transformation and protein kinase activity.

Two temperature-sensitive mutants of Fujinami sarcoma virus were isolated and characterized. Cells infected with the mutants were temperature sensitive in focus formation, colony formation, increased sugar uptake, and synthesis of plasminogen activator. The changes between transformed and nontransformed states of cultures were completely reversible by shifting the temperature. A Fujinami sarcoma virus-specific protein of 130,000 daltons, p130, was synthesized in mutant-infected cells regardless of the temperature, but the immunoprecipitates of p130 from extracts of infected cells were active in protein kinase only when cells had been incubated at the permissive temperature. These results appear to indicate that p130 is the transforming protein of Fujinami sarcoma virus, and that its protein kinase activity plays a crucial role in cell transformation by this virus.

Authors
Hanafusa, T; Mathey-Prevot, B; Feldman, RA; Hanafusa, H
MLA Citation
Hanafusa, T, Mathey-Prevot, B, Feldman, RA, and Hanafusa, H. "Mutants of Fujinami sarcoma virus which are temperature sensitive for cellular transformation and protein kinase activity." J Virol 38.1 (April 1981): 347-355.
PMID
6264108
Source
pubmed
Published In
Journal of virology
Volume
38
Issue
1
Publish Date
1981
Start Page
347
End Page
355

Purification and chemical characterization of melittin and acetylated derivatives.

Melittin, the main basic and hydrophobic peptide of bee venom, displays marked detergent-like properties. At high peptide concentration, and depending on salt and pH, it forms a tetramer. This is prevented by using urea. A purification procedure in presence of 4.0 M urea was developed to prepare melittin in its monomeric form, free of other venom constituents such as N alpha-formyl melittin, degradation products of peptides and phospholipase A2. NH2-residues on the melittin molecule were modified by reaction with acetic anhydride to alter the asymmetrical charge distribution supposed to confer detergent-like properties to the molecule. This gave rise to di- and mono acetyl derivatives which could be used, once isolated, to study further the melittin structure-activity relationship.

Authors
Maulet, Y; Mathey-Prevot, B; Kaiser, G; Rüegg, UT; Fulpius, BW
MLA Citation
Maulet, Y, Mathey-Prevot, B, Kaiser, G, Rüegg, UT, and Fulpius, BW. "Purification and chemical characterization of melittin and acetylated derivatives." Biochim Biophys Acta 625.2 (October 21, 1980): 274-280.
PMID
7437462
Source
pubmed
Published In
Biochimica et Biophysica Acta: international journal of biochemistry and biophysics
Volume
625
Issue
2
Publish Date
1980
Start Page
274
End Page
280
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