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Bagnat, Michel

Positions:

Associate Professor of Cell Biology

Cell Biology
School of Medicine

Member of the Duke Cancer Institute

Duke Cancer Institute
School of Medicine

Education:

Ph.D. 2002

Ph.D. — University of Madrid (Spain)

News:

Grants:

Mechanisms protecting cell surface integrity in giant vacuolated cells of the notochord

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
June 01, 2017
End Date
May 31, 2020

Regulation of the Epicardial Injury Response During Heart Regeneration in Zebrafish

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Co Investigator
Start Date
January 01, 2016
End Date
December 31, 2019

Uncovering mechanisms controlling notochord vacuole and spine morphogenesis

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
September 17, 2013
End Date
August 31, 2018

Epigenetic Control of Intestinal Inflammation

Administered By
Cell Biology
AwardedBy
Kenneth Rainin Foundation
Role
Principal Investigator
Start Date
October 01, 2016
End Date
September 30, 2017

Duke Training Grant in Digestive Diseases and Nutrition

Administered By
Medicine, Gastroenterology
AwardedBy
National Institutes of Health
Role
Mentor
Start Date
July 01, 1988
End Date
June 30, 2017

Lightsheet Imaging System

Administered By
Biology
AwardedBy
National Institutes of Health
Role
Major User
Start Date
March 15, 2016
End Date
March 14, 2017

Epigenetic control of intestinal inflammation

Administered By
Cell Biology
AwardedBy
Kenneth Rainin Foundation
Role
Principal Investigator
Start Date
October 01, 2015
End Date
September 30, 2016

Epigenetic Regulation of Intestinal Inflammation

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
December 01, 2013
End Date
June 30, 2015

Discovering New Regulators of CFTR and Fluid Secretion in Zebrafish

Administered By
Cell Biology
AwardedBy
National Institutes of Health
Role
Principal Investigator
Start Date
September 30, 2009
End Date
June 30, 2014
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Publications:

Single epicardial cell transcriptome sequencing identifies Caveolin 1 as an essential factor in zebrafish heart regeneration.

In contrast to mammals, adult zebrafish have a high capacity to regenerate damaged or lost myocardium through proliferation of cardiomyocytes spared from damage. The epicardial sheet covering the heart is activated by injury and aids muscle regeneration through paracrine effects and as a multipotent cell source, and has received recent attention as a target in cardiac repair strategies. Although it is recognized that epicardium is required for muscle regeneration and itself has high regenerative potential, the extent of cellular heterogeneity within epicardial tissue is largely unexplored. Here, we performed transcriptome analysis on dozens of epicardial lineage cells purified from zebrafish harboring a transgenic reporter for the pan-epicardial gene tcf21. Hierarchical clustering analysis suggested the presence of at least three epicardial cell subsets defined by expression signatures. We validated many new pan-epicardial and epicardial markers by alternative expression assays. Additionally, we explored the function of the scaffolding protein and main component of caveolae, caveolin 1 (cav1), which was present in each epicardial subset. In BAC transgenic zebrafish, cav1 regulatory sequences drove strong expression in ostensibly all epicardial cells and in coronary vascular endothelial cells. Moreover, cav1 mutant zebrafish generated by genome editing showed grossly normal heart development and adult cardiac anatomy, but displayed profound defects in injury-induced cardiomyocyte proliferation and heart regeneration. Our study defines a new platform for the discovery of epicardial lineage markers, genetic tools, and mechanisms of heart regeneration.

Authors
Cao, J; Navis, A; Cox, BD; Dickson, AL; Gemberling, M; Karra, R; Bagnat, M; Poss, KD
MLA Citation
Cao, J, Navis, A, Cox, BD, Dickson, AL, Gemberling, M, Karra, R, Bagnat, M, and Poss, KD. "Single epicardial cell transcriptome sequencing identifies Caveolin 1 as an essential factor in zebrafish heart regeneration." Development (Cambridge, England) 143.2 (January 2016): 232-243.
PMID
26657776
Source
epmc
Published In
Development (Cambridge)
Volume
143
Issue
2
Publish Date
2016
Start Page
232
End Page
243
DOI
10.1242/dev.130534

Developing pressures: fluid forces driving morphogenesis.

Over several decades genetic studies have unraveled many molecular mechanisms that underlie the signaling networks guiding morphogenesis, but the mechanical forces at work remain much less well understood. Accumulation of fluid within a luminal space can generate outward hydrostatic pressure capable of shaping morphogenesis at several scales, ranging from individual organs to the entire vertebrate body-plan. Here, we focus on recent work that uncovered mechanical roles for fluid secretion during morphogenesis. Identifying the roles and regulation of fluid secretion will be instrumental for understanding the mechanics of morphogenesis as well as many human diseases of complex genetic and environmental origin including secretory diarrheas and scoliosis.

Authors
Navis, A; Bagnat, M
MLA Citation
Navis, A, and Bagnat, M. "Developing pressures: fluid forces driving morphogenesis." Current opinion in genetics & development 32 (June 2015): 24-30. (Review)
PMID
25698116
Source
epmc
Published In
Current Opinion in Genetics & Development
Volume
32
Publish Date
2015
Start Page
24
End Page
30
DOI
10.1016/j.gde.2015.01.010

Infection, Inflammation and Healing in Zebrafish: Intestinal Inflammation.

Inflammatory bowel diseases (IBD), which include Crohn's disease and ulcerative colitis, contribute to significant morbidity and mortality globally. Despite an increase in incidence, IBD onset is still poorly understood. Mouse models of IBD recapitulate several aspects of human disease, but limited accessibility for live imaging and the lack of forward genetics highlight the need for new model systems for disease onset characterization. Zebrafish represent a powerful platform to model IBD using forward and reverse genetics, live imaging of transgenic lines and physiological assays. In this review, we address current models of IBD in zebrafish and newly developed reagents available for future studies.

Authors
Marjoram, L; Bagnat, M
MLA Citation
Marjoram, L, and Bagnat, M. "Infection, Inflammation and Healing in Zebrafish: Intestinal Inflammation." Current pathobiology reports 3.2 (June 2015): 147-153.
PMID
26236567
Source
epmc
Published In
Current Pathobiology Reports
Volume
3
Issue
2
Publish Date
2015
Start Page
147
End Page
153
DOI
10.1007/s40139-015-0079-x

Loss of cftr function leads to pancreatic destruction in larval zebrafish.

The development and function of many internal organs requires precisely regulated fluid secretion. A key regulator of vertebrate fluid secretion is an anion channel, the cystic fibrosis transmembrane conductance regulator (CFTR). Loss of CFTR function leads to defects in fluid transport and cystic fibrosis (CF), a complex disease characterized by a loss of fluid secretion and mucus buildup in many organs including the lungs, liver, and pancreas. Several animal models including mouse, ferret and pig have been generated to investigate the pathophysiology of CF. However, these models have limited accessibility to early processes in the development of CF and are not amenable for forward genetic or chemical screens. Here, we show that Cftr is expressed and localized to the apical membrane of the zebrafish pancreatic duct and that loss of cftr function leads to destruction of the exocrine pancreas and a cystic fibrosis phenotype that mirrors human disease. Our analyses reveal that the cftr mutant pancreas initially develops normally, then rapidly loses pancreatic tissue during larval life, reflecting pancreatic disease in CF. Altogether, we demonstrate that the cftr mutant zebrafish is a powerful new model for pancreatitis and pancreatic destruction in CF. This accessible model will allow more detailed investigation into the mechanisms that drive CF of the pancreas and facilitate development of new therapies to treat the disease.

Authors
Navis, A; Bagnat, M
MLA Citation
Navis, A, and Bagnat, M. "Loss of cftr function leads to pancreatic destruction in larval zebrafish." Developmental biology 399.2 (March 2015): 237-248.
PMID
25592226
Source
epmc
Published In
Developmental Biology
Volume
399
Issue
2
Publish Date
2015
Start Page
237
End Page
248
DOI
10.1016/j.ydbio.2014.12.034

Developmental regulation of apical endocytosis controls epithelial patterning in vertebrate tubular organs.

Epithelial organs develop through tightly coordinated events of cell proliferation and differentiation in which endocytosis plays a major role. Despite recent advances, how endocytosis regulates the development of vertebrate organs is still unknown. Here we describe a mechanism that facilitates the apical availability of endosomal SNARE receptors for epithelial morphogenesis through the developmental upregulation of plasmolipin (pllp) in a highly endocytic segment of the zebrafish posterior midgut. The protein PLLP (Pllp in fish) recruits the clathrin adaptor EpsinR to sort the SNARE machinery of the endolysosomal pathway into the subapical compartment, which is a switch for polarized endocytosis. Furthermore, PLLP expression induces apical Crumbs internalization and the activation of the Notch signalling pathway, both crucial steps in the acquisition of cell polarity and differentiation of epithelial cells. We thus postulate that differential apical endosomal SNARE sorting is a mechanism that regulates epithelial patterning.

Authors
Rodríguez-Fraticelli, AE; Bagwell, J; Bosch-Fortea, M; Boncompain, G; Reglero-Real, N; García-León, MJ; Andrés, G; Toribio, ML; Alonso, MA; Millán, J; Perez, F; Bagnat, M; Martín-Belmonte, F
MLA Citation
Rodríguez-Fraticelli, AE, Bagwell, J, Bosch-Fortea, M, Boncompain, G, Reglero-Real, N, García-León, MJ, Andrés, G, Toribio, ML, Alonso, MA, Millán, J, Perez, F, Bagnat, M, and Martín-Belmonte, F. "Developmental regulation of apical endocytosis controls epithelial patterning in vertebrate tubular organs." Nature cell biology 17.3 (March 2015): 241-250.
PMID
25706235
Source
epmc
Published In
Nature Cell Biology
Volume
17
Issue
3
Publish Date
2015
Start Page
241
End Page
250
DOI
10.1038/ncb3106

Epigenetic control of intestinal barrier function and inflammation in zebrafish.

The intestinal epithelium forms a barrier protecting the organism from microbes and other proinflammatory stimuli. The integrity of this barrier and the proper response to infection requires precise regulation of powerful immune homing signals such as tumor necrosis factor (TNF). Dysregulation of TNF leads to inflammatory bowel diseases (IBD), but the mechanism controlling the expression of this potent cytokine and the events that trigger the onset of chronic inflammation are unknown. Here, we show that loss of function of the epigenetic regulator ubiquitin-like protein containing PHD and RING finger domains 1 (uhrf1) in zebrafish leads to a reduction in tnfa promoter methylation and the induction of tnfa expression in intestinal epithelial cells (IECs). The increase in IEC tnfa levels is microbe-dependent and results in IEC shedding and apoptosis, immune cell recruitment, and barrier dysfunction, consistent with chronic inflammation. Importantly, tnfa knockdown in uhrf1 mutants restores IEC morphology, reduces cell shedding, and improves barrier function. We propose that loss of epigenetic repression and TNF induction in the intestinal epithelium can lead to IBD onset.

Authors
Marjoram, L; Alvers, A; Deerhake, ME; Bagwell, J; Mankiewicz, J; Cocchiaro, JL; Beerman, RW; Willer, J; Sumigray, KD; Katsanis, N; Tobin, DM; Rawls, JF; Goll, MG; Bagnat, M
MLA Citation
Marjoram, L, Alvers, A, Deerhake, ME, Bagwell, J, Mankiewicz, J, Cocchiaro, JL, Beerman, RW, Willer, J, Sumigray, KD, Katsanis, N, Tobin, DM, Rawls, JF, Goll, MG, and Bagnat, M. "Epigenetic control of intestinal barrier function and inflammation in zebrafish." Proceedings of the National Academy of Sciences of the United States of America 112.9 (March 2015): 2770-2775.
PMID
25730872
Source
epmc
Published In
Proceedings of the National Academy of Sciences of USA
Volume
112
Issue
9
Publish Date
2015
Start Page
2770
End Page
2775
DOI
10.1073/pnas.1424089112

Loss of cftr function leads to pancreatic destruction in larval zebrafish

© 2015 Elsevier Inc.The development and function of many internal organs requires precisely regulated fluid secretion. A key regulator of vertebrate fluid secretion is an anion channel, the cystic fibrosis transmembrane conductance regulator (CFTR). Loss of CFTR function leads to defects in fluid transport and cystic fibrosis (CF), a complex disease characterized by a loss of fluid secretion and mucus buildup in many organs including the lungs, liver, and pancreas. Several animal models including mouse, ferret and pig have been generated to investigate the pathophysiology of CF. However, these models have limited accessibility to early processes in the development of CF and are not amenable for forward genetic or chemical screens. Here, we show that Cftr is expressed and localized to the apical membrane of the zebrafish pancreatic duct and that loss of cftr function leads to destruction of the exocrine pancreas and a cystic fibrosis phenotype that mirrors human disease. Our analyses reveal that the cftr mutant pancreas initially develops normally, then rapidly loses pancreatic tissue during larval life, reflecting pancreatic disease in CF. Altogether, we demonstrate that the cftr mutant zebrafish is a powerful new model for pancreatitis and pancreatic destruction in CF. This accessible model will allow more detailed investigation into the mechanisms that drive CF of the pancreas and facilitate development of new therapies to treat the disease.

Authors
Navis, A; Bagnat, M
MLA Citation
Navis, A, and Bagnat, M. "Loss of cftr function leads to pancreatic destruction in larval zebrafish." Developmental Biology 399.2 (January 1, 2015): 237-248.
Source
scopus
Published In
Developmental Biology
Volume
399
Issue
2
Publish Date
2015
Start Page
237
End Page
248
DOI
10.1016/j.ydbio.2014.12.034

Apicobasal polarity and lumen formation during development

© Springer International Publishing Switzerland 2015.Networks of interconnected tubes form the basic structural element of many organs. Tubes are composed of polarized epithelia that enclose a lumen. During organogenesis, lumens form by several distinct mechanisms, ranging from wrapping of an epithelial sheet to generation of a lumen de novo within a rod of cells. Nevertheless, all tube formation processes share some basic common principles that result in the generation of a single, continuous lumen. Interactions with the surrounding environment direct epithelial cell polarization that governs physiological regulators of lumen formation including the actin cytoskeleton, adhesion, vesicular transport, and fluid secretion. Polarized physiological processes mediate the mechanical interactions between epithelial cells and their environment. Polarity within actin cytoskeleton generates contractile forces generating morphogenetic movements. Secretion of fluid or matrix into the lumen generates outward forces driving lumen opening and expansion. Thus, cell polarity is crucial for vectorial transport processes and structural asymmetries during lumen formation. Here we focus on recent discoveries illuminating the relationship between lumen formation and cell polarity in vivo.

Authors
Bagnat, M; Navis, A
MLA Citation
Bagnat, M, and Navis, A. "Apicobasal polarity and lumen formation during development." Cell Polarity 2: Role in Development and Disease. January 1, 2015. 67-93.
Source
scopus
Publish Date
2015
Start Page
67
End Page
93
DOI
10.1007/978-3-319-14466-5_3

Single continuous lumen formation in the zebrafish gut is mediated by smoothened-dependent tissue remodeling.

The formation of a single lumen during tubulogenesis is crucial for the development and function of many organs. Although 3D cell culture models have identified molecular mechanisms controlling lumen formation in vitro, their function during vertebrate organogenesis is poorly understood. Using light sheet microscopy and genetic approaches we have investigated single lumen formation in the zebrafish gut. Here we show that during gut development multiple lumens open and enlarge to generate a distinct intermediate, which consists of two adjacent unfused lumens separated by basolateral contacts. We observed that these lumens arise independently from each other along the length of the gut and do not share a continuous apical surface. Resolution of this intermediate into a single, continuous lumen requires the remodeling of contacts between adjacent lumens and subsequent lumen fusion. We show that lumen resolution, but not lumen opening, is impaired in smoothened (smo) mutants, indicating that fluid-driven lumen enlargement and resolution are two distinct processes. Furthermore, we show that smo mutants exhibit perturbations in the Rab11 trafficking pathway and demonstrate that Rab11-mediated trafficking is necessary for single lumen formation. Thus, lumen resolution is a distinct genetically controlled process crucial for single, continuous lumen formation in the zebrafish gut.

Authors
Alvers, AL; Ryan, S; Scherz, PJ; Huisken, J; Bagnat, M
MLA Citation
Alvers, AL, Ryan, S, Scherz, PJ, Huisken, J, and Bagnat, M. "Single continuous lumen formation in the zebrafish gut is mediated by smoothened-dependent tissue remodeling." Development (Cambridge, England) 141.5 (March 2014): 1110-1119.
PMID
24504339
Source
epmc
Published In
Development (Cambridge)
Volume
141
Issue
5
Publish Date
2014
Start Page
1110
End Page
1119
DOI
10.1242/dev.100313

Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations.

Congenital vertebral malformations (CVM) occur in 1 in 1000 live births and in many cases can cause spinal deformities, such as scoliosis, and result in disability and distress of affected individuals. Many severe forms of the disease, such as spondylocostal dystostosis, are recessive monogenic traits affecting somitogenesis, however the etiologies of the majority of CVM cases remain undetermined. Here we demonstrate that morphological defects of the notochord in zebrafish can generate congenital-type spine defects. We characterize three recessive zebrafish leviathan/col8a1a mutant alleles ((m531, vu41, vu105)) that disrupt collagen type VIII alpha1a (col8a1a), and cause folding of the embryonic notochord and consequently adult vertebral column malformations. Furthermore, we provide evidence that a transient loss of col8a1a function or inhibition of Lysyl oxidases with drugs during embryogenesis was sufficient to generate vertebral fusions and scoliosis in the adult spine. Using periodic imaging of individual zebrafish, we correlate focal notochord defects of the embryo with vertebral malformations (VM) in the adult. Finally, we show that bends and kinks in the notochord can lead to aberrant apposition of osteoblasts normally confined to well-segmented areas of the developing vertebral bodies. Our results afford a novel mechanism for the formation of VM, independent of defects of somitogenesis, resulting from aberrant bone deposition at regions of misshapen notochord tissue.

Authors
Gray, RS; Wilm, TP; Smith, J; Bagnat, M; Dale, RM; Topczewski, J; Johnson, SL; Solnica-Krezel, L
MLA Citation
Gray, RS, Wilm, TP, Smith, J, Bagnat, M, Dale, RM, Topczewski, J, Johnson, SL, and Solnica-Krezel, L. "Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations." Dev Biol 386.1 (February 1, 2014): 72-85.
PMID
24333517
Source
pubmed
Published In
Developmental Biology
Volume
386
Issue
1
Publish Date
2014
Start Page
72
End Page
85
DOI
10.1016/j.ydbio.2013.11.028

Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations

Congenital vertebral malformations (CVM) occur in 1 in 1000 live births and in many cases can cause spinal deformities, such as scoliosis, and result in disability and distress of affected individuals. Many severe forms of the disease, such as spondylocostal dystostosis, are recessive monogenic traits affecting somitogenesis, however the etiologies of the majority of CVM cases remain undetermined. Here we demonstrate that morphological defects of the notochord in zebrafish can generate congenital-type spine defects. We characterize three recessive zebrafish leviathan/col8a1a mutant alleles (m531, vu41, vu105) that disrupt collagen type VIII alpha1a (col8a1a), and cause folding of the embryonic notochord and consequently adult vertebral column malformations. Furthermore, we provide evidence that a transient loss of col8a1a function or inhibition of Lysyl oxidases with drugs during embryogenesis was sufficient to generate vertebral fusions and scoliosis in the adult spine. Using periodic imaging of individual zebrafish, we correlate focal notochord defects of the embryo with vertebral malformations (VM) in the adult. Finally, we show that bends and kinks in the notochord can lead to aberrant apposition of osteoblasts normally confined to well-segmented areas of the developing vertebral bodies. Our results afford a novel mechanism for the formation of VM, independent of defects of somitogenesis, resulting from aberrant bone deposition at regions of misshapen notochord tissue. © 2013 Elsevier Inc.

Authors
Gray, RS; Wilm, TP; Smith, J; Bagnat, M; Dale, RM; Topczewski, J; Johnson, SL; Solnica-Krezel, L
MLA Citation
Gray, RS, Wilm, TP, Smith, J, Bagnat, M, Dale, RM, Topczewski, J, Johnson, SL, and Solnica-Krezel, L. "Loss of col8a1a function during zebrafish embryogenesis results in congenital vertebral malformations." Developmental Biology 386.1 (February 1, 2014): 72-85.
Source
scopus
Published In
Developmental Biology
Volume
386
Issue
1
Publish Date
2014
Start Page
72
End Page
85
DOI
10.1016/j.ydbio.2013.11.028

Directing traffic into the future.

Authors
Antonny, B; Audhya, J; l Bagnat, M; von Blume, J; Briggs, JA; Giraudo, C; Kaeser, PS; Miller, E; Reinisch, K; Sbalzarini, IF; Schuldiner, M; Shen, J; Takamori, S; Verstreken, P; Walther, T
MLA Citation
Antonny, B, Audhya, J, l Bagnat, M, von Blume, J, Briggs, JA, Giraudo, C, Kaeser, PS, Miller, E, Reinisch, K, Sbalzarini, IF, Schuldiner, M, Shen, J, Takamori, S, Verstreken, P, and Walther, T. "Directing traffic into the future." Developmental cell 27.5 (December 2013): 480-484.
PMID
24482802
Source
epmc
Published In
Developmental Cell
Volume
27
Issue
5
Publish Date
2013
Start Page
480
End Page
484
DOI
10.1016/j.devcel.2013.11.017

Rapid identification of kidney cyst mutations by whole exome sequencing in zebrafish.

Forward genetic approaches in zebrafish have provided invaluable information about developmental processes. However, the relative difficulty of mapping and isolating mutations has limited the number of new genetic screens. Recent improvements in the annotation of the zebrafish genome coupled to a reduction in sequencing costs prompted the development of whole genome and RNA sequencing approaches for gene discovery. Here we describe a whole exome sequencing (WES) approach that allows rapid and cost-effective identification of mutations. We used our WES methodology to isolate four mutations that cause kidney cysts; we identified novel alleles in two ciliary genes as well as two novel mutants. The WES approach described here does not require specialized infrastructure or training and is therefore widely accessible. This methodology should thus help facilitate genetic screens and expedite the identification of mutants that can inform basic biological processes and the causality of genetic disorders in humans.

Authors
Ryan, S; Willer, J; Marjoram, L; Bagwell, J; Mankiewicz, J; Leshchiner, I; Goessling, W; Bagnat, M; Katsanis, N
MLA Citation
Ryan, S, Willer, J, Marjoram, L, Bagwell, J, Mankiewicz, J, Leshchiner, I, Goessling, W, Bagnat, M, and Katsanis, N. "Rapid identification of kidney cyst mutations by whole exome sequencing in zebrafish." Development 140.21 (November 2013): 4445-4451.
PMID
24130329
Source
pubmed
Published In
Development (Cambridge)
Volume
140
Issue
21
Publish Date
2013
Start Page
4445
End Page
4451
DOI
10.1242/dev.101170

The vacuole within: how cellular organization dictates notochord function.

Authors
Ellis, K; Hoffman, BD; Bagnat, M
MLA Citation
Ellis, K, Hoffman, BD, and Bagnat, M. "The vacuole within: how cellular organization dictates notochord function." Bioarchitecture 3.3 (May 2013): 64-68. (Review)
PMID
23887209
Source
pubmed
Published In
Bioarchitecture
Volume
3
Issue
3
Publish Date
2013
Start Page
64
End Page
68
DOI
10.4161/bioa.25503

Cftr controls lumen expansion and function of Kupffer's vesicle in zebrafish.

Regulated fluid secretion is crucial for the function of most organs. In vertebrates, the chloride channel cystic fibrosis transmembrane conductance regulator (CFTR) is a master regulator of fluid secretion. Although the biophysical properties of CFTR have been well characterized in vitro, little is known about its in vivo role during development. Here, we investigated the function of Cftr during zebrafish development by generating several cftr mutant alleles using TAL effector nucleases. We found that loss of cftr function leads to organ laterality defects. In zebrafish, left-right (LR) asymmetry requires cilia-driven fluid flow within the lumen of Kupffer's vesicle (KV). Using live imaging we found that KV morphogenesis is disrupted in cftr mutants. Loss of Cftr-mediated fluid secretion impairs KV lumen expansion leading to defects in organ laterality. Using bacterial artificial chromosome recombineering, we generated transgenic fish expressing functional Cftr fusion proteins with fluorescent tags under the control of the cftr promoter. The transgenes completely rescued the cftr mutant phenotype. Live imaging of these transgenic lines showed that Cftr is localized to the apical membrane of the epithelial cells in KV during lumen formation. Pharmacological stimulation of Cftr-dependent fluid secretion led to an expansion of the KV lumen. Conversely, inhibition of ion gradient formation impaired KV lumen inflation. Interestingly, cilia formation and motility in KV were not affected, suggesting that fluid secretion and flow are independently controlled in KV. These findings uncover a new role for cftr in KV morphogenesis and function during zebrafish development.

Authors
Navis, A; Marjoram, L; Bagnat, M
MLA Citation
Navis, A, Marjoram, L, and Bagnat, M. "Cftr controls lumen expansion and function of Kupffer's vesicle in zebrafish." Development 140.8 (April 2013): 1703-1712.
PMID
23487313
Source
pubmed
Published In
Development (Cambridge)
Volume
140
Issue
8
Publish Date
2013
Start Page
1703
End Page
1712
DOI
10.1242/dev.091819

Zebrafish as a model to analyze macromolecule absorption in intestinal enterocytes

Authors
Cocchiaro, JL; Navis, A; Bagnat, M; Rawls, JF
MLA Citation
Cocchiaro, JL, Navis, A, Bagnat, M, and Rawls, JF. "Zebrafish as a model to analyze macromolecule absorption in intestinal enterocytes." April 2013.
Source
wos-lite
Published In
The FASEB journal : official publication of the Federation of American Societies for Experimental Biology
Volume
27
Publish Date
2013

Notochord vacuoles are lysosome-related organelles that function in axis and spine morphogenesis.

The notochord plays critical structural and signaling roles during vertebrate development. At the center of the vertebrate notochord is a large fluid-filled organelle, the notochord vacuole. Although these highly conserved intracellular structures have been described for decades, little is known about the molecular mechanisms involved in their biogenesis and maintenance. Here we show that zebrafish notochord vacuoles are specialized lysosome-related organelles whose formation and maintenance requires late endosomal trafficking regulated by the vacuole-specific Rab32a and H(+)-ATPase-dependent acidification. We establish that notochord vacuoles are required for body axis elongation during embryonic development and identify a novel role in spine morphogenesis. Thus, the vertebrate notochord plays important structural roles beyond early development.

Authors
Ellis, K; Bagwell, J; Bagnat, M
MLA Citation
Ellis, K, Bagwell, J, and Bagnat, M. "Notochord vacuoles are lysosome-related organelles that function in axis and spine morphogenesis." J Cell Biol 200.5 (March 4, 2013): 667-679.
PMID
23460678
Source
pubmed
Published In
The Journal of Cell Biology
Volume
200
Issue
5
Publish Date
2013
Start Page
667
End Page
679
DOI
10.1083/jcb.201212095

IDENTIFICATION OF CFTR AND FLUID SECRETION REGULATORS IN ZEBRAFISH

Authors
Marjoram, LT; Bagnat, M
MLA Citation
Marjoram, LT, and Bagnat, M. "IDENTIFICATION OF CFTR AND FLUID SECRETION REGULATORS IN ZEBRAFISH." PEDIATRIC PULMONOLOGY 47 (September 2012): 282-282.
Source
wos-lite
Published In
Pediatric Pulmonology
Volume
47
Publish Date
2012
Start Page
282
End Page
282

CFTR IS REQUIRED DURING DEVELOPMENT OF THE ZEBRAFISH LIVER AND PANCREAS

Authors
Navis, A; Bagnat, M
MLA Citation
Navis, A, and Bagnat, M. "CFTR IS REQUIRED DURING DEVELOPMENT OF THE ZEBRAFISH LIVER AND PANCREAS." PEDIATRIC PULMONOLOGY 47 (September 2012): 288-288.
Source
wos-lite
Published In
Pediatric Pulmonology
Volume
47
Publish Date
2012
Start Page
288
End Page
288

Regulation of intrahepatic biliary duct morphogenesis by Claudin 15-like b.

The intrahepatic biliary ducts transport bile produced by the hepatocytes out of the liver. Defects in biliary cell differentiation and biliary duct remodeling cause a variety of congenital diseases including Alagille Syndrome and polycystic liver disease. While the molecular pathways regulating biliary cell differentiation have received increasing attention (Lemaigre, 2010), less is known about the cellular behavior underlying biliary duct remodeling. Here, we have identified a novel gene, claudin 15-like b (cldn15lb), which exhibits a unique and dynamic expression pattern in the hepatocytes and biliary epithelial cells in zebrafish. Claudins are tight junction proteins that have been implicated in maintaining epithelial polarity, regulating paracellular transport, and providing barrier function. In zebrafish cldn15lb mutant livers, tight junctions are observed between hepatocytes, but these cells show polarization defects as well as canalicular malformations. Furthermore, cldn15lb mutants show abnormalities in biliary duct morphogenesis whereby biliary epithelial cells remain clustered together and form a disorganized network. Our data suggest that Cldn15lb plays an important role in the remodeling process during biliary duct morphogenesis. Thus, cldn15lb mutants provide a novel in vivo model to study the role of tight junction proteins in the remodeling of the biliary network and hereditary cholestasis.

Authors
Cheung, ID; Bagnat, M; Ma, TP; Datta, A; Evason, K; Moore, JC; Lawson, ND; Mostov, KE; Moens, CB; Stainier, DY
MLA Citation
Cheung, ID, Bagnat, M, Ma, TP, Datta, A, Evason, K, Moore, JC, Lawson, ND, Mostov, KE, Moens, CB, and Stainier, DY. "Regulation of intrahepatic biliary duct morphogenesis by Claudin 15-like b." Dev Biol 361.1 (January 1, 2012): 68-78.
PMID
22020048
Source
pubmed
Published In
Developmental Biology
Volume
361
Issue
1
Publish Date
2012
Start Page
68
End Page
78
DOI
10.1016/j.ydbio.2011.10.004

Microbial colonization induces dynamic temporal and spatial patterns of NF-κB activation in the zebrafish digestive tract

Background & Aims: The nuclear factor κ-light-chain enhancer of activated B cells (NF-κB) transcription factor pathway is activated in response to diverse microbial stimuli to regulate expression of genes involved in immune responses and tissue homeostasis. However, the temporal and spatial activation of NF-κB in response to microbial signals have not been determined in whole living organisms, and the molecular and cellular details of these responses are not well understood. We used in vivo imaging and molecular approaches to analyze NF-κB activation in response to the commensal microbiota in transparent gnotobiotic zebrafish. Methods: We used DNA microarrays, in situ hybridization, and quantitative reverse transcription polymerase chain reaction analyses to study the effects of the commensal microbiota on gene expression in gnotobiotic zebrafish. Zebrafish PAC2 and ZFL cells were used to study the NF-κB signaling pathway in response to bacterial stimuli. We generated transgenic zebrafish that express enhanced green fluorescent protein under transcriptional control of NF-κB, and used them to study patterns of NF-κB activation during development and microbial colonization. Results: Bacterial stimulation induced canonical activation of the NF-κB pathway in zebrafish cells. Colonization of germ-free transgenic zebrafish with a commensal microbiota activated NF-κB and led to up-regulation of its target genes in intestinal and extraintestinal tissues of the digestive tract. Colonization with the bacterium Pseudomonas aeruginosa was sufficient to activate NF-κB, and this activation required a functional flagellar apparatus. Conclusions: In zebrafish, transcriptional activity of NF-κB is spatially and temporally regulated by specific microbial factors. The observed patterns of NF-κB-dependent responses to microbial colonization indicate that cells in the gastrointestinal tract respond robustly to the microbial environment. © 2011 AGA Institute.

Authors
Kanther, M; Sun, X; Mhlbauer, M; MacKey, LC; III, EJF; Bagnat, M; Jobin, C; Rawls, JF
MLA Citation
Kanther, M, Sun, X, Mhlbauer, M, MacKey, LC, III, EJF, Bagnat, M, Jobin, C, and Rawls, JF. "Microbial colonization induces dynamic temporal and spatial patterns of NF-κB activation in the zebrafish digestive tract." Gastroenterology 141.1 (2011): 197-207.
PMID
21439961
Source
scival
Published In
Gastroenterology
Volume
141
Issue
1
Publish Date
2011
Start Page
197
End Page
207
DOI
10.1053/j.gastro.2011.03.042

Cse1l is a negative regulator of CFTR-dependent fluid secretion.

Transport of chloride through the cystic fibrosis transmembrane conductance regulator (CFTR) channel is a key step in regulating fluid secretion in vertebrates [1, 2]. Loss of CFTR function leads to cystic fibrosis [1, 3, 4], a disease that affects the lungs, pancreas, liver, intestine, and vas deferens. Conversely, uncontrolled activation of the channel leads to increased fluid secretion and plays a major role in several diseases and conditions including cholera [5, 6] and other secretory diarrheas [7] as well as polycystic kidney disease [8-10]. Understanding how CFTR activity is regulated in vivo has been limited by the lack of a genetic model. Here, we used a forward genetic approach in zebrafish to uncover CFTR regulators. We report the identification, isolation, and characterization of a mutation in the zebrafish cse1l gene that leads to the sudden and dramatic expansion of the gut tube. We show that this phenotype results from a rapid accumulation of fluid due to the uncontrolled activation of the CFTR channel. Analyses in zebrafish larvae and mammalian cells indicate that Cse1l is a negative regulator of CFTR-dependent fluid secretion. This work demonstrates the importance of fluid homeostasis in development and establishes the zebrafish as a much-needed model system to study CFTR regulation in vivo.

Authors
Bagnat, M; Navis, A; Herbstreith, S; Brand-Arzamendi, K; Curado, S; Gabriel, S; Mostov, K; Huisken, J; Stainier, DY
MLA Citation
Bagnat, M, Navis, A, Herbstreith, S, Brand-Arzamendi, K, Curado, S, Gabriel, S, Mostov, K, Huisken, J, and Stainier, DY. "Cse1l is a negative regulator of CFTR-dependent fluid secretion." Curr Biol 20.20 (October 26, 2010): 1840-1845.
PMID
20933420
Source
pubmed
Published In
Current Biology
Volume
20
Issue
20
Publish Date
2010
Start Page
1840
End Page
1845
DOI
10.1016/j.cub.2010.09.012

Erratum: Cse1l is a negative regulator of CFTR-dependent fluid secretion (Current Biology (2010) 20 (1840-1845))

Authors
Bagnat, M; Navis, A; Herbstreith, S; Brand-Arzamendi, K; Curado, S; Gabriel, S; Mostov, K; Huisken, J; Stainier, DYR
MLA Citation
Bagnat, M, Navis, A, Herbstreith, S, Brand-Arzamendi, K, Curado, S, Gabriel, S, Mostov, K, Huisken, J, and Stainier, DYR. "Erratum: Cse1l is a negative regulator of CFTR-dependent fluid secretion (Current Biology (2010) 20 (1840-1845))." Current Biology 20.23 (2010): 2157--.
Source
scival
Published In
Current Biology
Volume
20
Issue
23
Publish Date
2010
Start Page
2157-
DOI
10.1016/j.cub.2010.11.031

IDENTIFICATION OF THREE CFTR REGULATORS USING FORWARD GENETICS IN ZEBRAFISH

Authors
Bagnat, M; Huisken, J; Herbstreith, S; Brand-Arzamendi, K; Curado, S; Satinier, DY
MLA Citation
Bagnat, M, Huisken, J, Herbstreith, S, Brand-Arzamendi, K, Curado, S, and Satinier, DY. "IDENTIFICATION OF THREE CFTR REGULATORS USING FORWARD GENETICS IN ZEBRAFISH." PEDIATRIC PULMONOLOGY (2009): 226-226.
Source
wos-lite
Published In
Pediatric Pulmonology
Publish Date
2009
Start Page
226
End Page
226

Genetic control of single lumen formation in the zebrafish gut.

Most organs consist of networks of interconnected tubes that serve as conduits to transport fluid and cells and act as physiological barriers between compartments. Biological tubes are assembled through very diverse developmental processes that generate structures of different shapes and sizes. Nevertheless, all biological tubes invariably possess one single lumen. The mechanisms responsible for single lumen specification are not known. Here we show that zebrafish mutants for the MODY5 and familial GCKD gene tcf2 (also known as vhnf1) fail to specify a single lumen in their gut tube and instead develop multiple lumens. We show that Tcf2 controls single lumen formation by regulating claudin15 and Na+/K+-ATPase expression. Our in vivo and in vitro results indicate that Claudin15 functions in paracellular ion transport to specify single lumen formation. This work shows that single lumen formation is genetically controlled and appears to be driven by the accumulation of fluid.

Authors
Bagnat, M; Cheung, ID; Mostov, KE; Stainier, DY
MLA Citation
Bagnat, M, Cheung, ID, Mostov, KE, and Stainier, DY. "Genetic control of single lumen formation in the zebrafish gut." Nat Cell Biol 9.8 (August 2007): 954-960.
PMID
17632505
Source
pubmed
Published In
Nature Cell Biology
Volume
9
Issue
8
Publish Date
2007
Start Page
954
End Page
960
DOI
10.1038/ncb1621

Plasma membrane polarization during mating in yeast cells.

The yeast mating cell provides a simple paradigm for analyzing mechanisms underlying the generation of surface polarity. Endocytic recycling and slow diffusion on the plasma membrane were shown to facilitate polarized surface distribution of Snc1p (Valdez-Taubas, J., and H.R. Pelham. 2003. Curr. Biol. 13:1636-1640). Here, we found that polarization of Fus1p, a raft-associated type I transmembrane protein involved in cell fusion, does not depend on endocytosis. Instead, Fus1p localization to the tip of the mating projection was determined by its cytosolic domain, which binds to peripheral proteins involved in mating tip polarization. Furthermore, we provide evidence that the lipid bilayer at the mating projection is more condensed than the plasma membrane enclosing the cell body, and that sphingolipids are required for this lipid organization.

Authors
Proszynski, TJ; Klemm, R; Bagnat, M; Gaus, K; Simons, K
MLA Citation
Proszynski, TJ, Klemm, R, Bagnat, M, Gaus, K, and Simons, K. "Plasma membrane polarization during mating in yeast cells." J Cell Biol 173.6 (June 19, 2006): 861-866.
PMID
16769822
Source
pubmed
Published In
The Journal of Cell Biology
Volume
173
Issue
6
Publish Date
2006
Start Page
861
End Page
866
DOI
10.1083/jcb.200602007

A genome-wide visual screen reveals a role for sphingolipids and ergosterol in cell surface delivery in yeast.

Recently synthesized proteins are sorted at the trans-Golgi network into specialized routes for exocytosis. Surprisingly little is known about the underlying molecular machinery. Here, we present a visual screen to search for proteins involved in cargo sorting and vesicle formation. We expressed a GFP-tagged plasma membrane protein in the yeast deletion library and identified mutants with altered marker localization. This screen revealed a requirement of several enzymes regulating the synthesis of sphingolipids and ergosterol in the correct and efficient delivery of the marker protein to the cell surface. Additionally, we identified mutants regulating the actin cytoskeleton (Rvs161p and Vrp1p), known membrane traffic regulators (Kes1p and Chs5p), and several unknown genes. This visual screening method can now be used for different cargo proteins to search in a genome-wide fashion for machinery involved in post-Golgi sorting.

Authors
Proszynski, TJ; Klemm, RW; Gravert, M; Hsu, PP; Gloor, Y; Wagner, J; Kozak, K; Grabner, H; Walzer, K; Bagnat, M; Simons, K; Walch-Solimena, C
MLA Citation
Proszynski, TJ, Klemm, RW, Gravert, M, Hsu, PP, Gloor, Y, Wagner, J, Kozak, K, Grabner, H, Walzer, K, Bagnat, M, Simons, K, and Walch-Solimena, C. "A genome-wide visual screen reveals a role for sphingolipids and ergosterol in cell surface delivery in yeast." Proc Natl Acad Sci U S A 102.50 (December 13, 2005): 17981-17986.
PMID
16330752
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
102
Issue
50
Publish Date
2005
Start Page
17981
End Page
17986
DOI
10.1073/pnas.0509107102

O-glycosylation as a sorting determinant for cell surface delivery in yeast.

Little is known about the mechanisms that determine localization of proteins to the plasma membrane in Saccharomyces cerevisiae. The length of the transmembrane domains and association of proteins with lipid rafts have been proposed to play a role in sorting to the cell surface. Here, we report that Fus1p, an O-glycosylated integral membrane protein involved in cell fusion during yeast mating, requires O-glycosylation for cell surface delivery. In cells lacking PMT4, encoding a mannosyltransferase involved in the initial step of O-glycosylation, Fus1p was not glycosylated and accumulated in late Golgi structures. A chimeric protein lacking O-glycosylation motif was missorted to the vacuole and accumulated in late Golgi in wild-type cells. Exocytosis of this protein could be restored by addition of a 33-amino acid portion of an O-glycosylated sequence from Fus1p. Our data suggest that O-glycosylation functions as a sorting determinant for cell surface delivery of Fus1p.

Authors
Proszynski, TJ; Simons, K; Bagnat, M
MLA Citation
Proszynski, TJ, Simons, K, and Bagnat, M. "O-glycosylation as a sorting determinant for cell surface delivery in yeast." Mol Biol Cell 15.4 (April 2004): 1533-1543.
PMID
14742720
Source
pubmed
Published In
Molecular Biology of the Cell
Volume
15
Issue
4
Publish Date
2004
Start Page
1533
End Page
1543
DOI
10.1091/mbc.E03-07-0511

Aberrant processing of the WSC family and Mid2p cell surface sensors results in cell death of Saccharomyces cerevisiae O-mannosylation mutants.

Protein O mannosylation is a crucial protein modification in uni- and multicellular eukaryotes. In humans, a lack of O-mannosyl glycans causes congenital muscular dystrophies that are associated with brain abnormalities. In yeast, protein O mannosylation is vital; however, it is not known why impaired O mannosylation results in cell death. To address this question, we analyzed the conditionally lethal Saccharomyces cerevisiae protein O-mannosyltransferase pmt2 pmt4Delta mutant. We found that pmt2 pmt4Delta cells lyse as small-budded cells in the absence of osmotic stabilization and that treatment with mating pheromone causes pheromone-induced cell death. These phenotypes are partially suppressed by overexpression of upstream elements of the protein kinase C (PKC1) cell integrity pathway, suggesting that the PKC1 pathway is defective in pmt2 pmt4Delta mutants. Congruently, induction of Mpk1p/Slt2p tyrosine phosphorylation does not occur in pmt2 pmt4Delta mutants during exposure to mating pheromone or elevated temperature. Detailed analyses of the plasma membrane sensors of the PKC1 pathway revealed that Wsc1p, Wsc2p, and Mid2p are aberrantly processed in pmt mutants. Our data suggest that in yeast, O mannosylation increases the activity of Wsc1p, Wsc2p, and Mid2p by enhancing their stability. Reduced O mannosylation leads to incorrect proteolytic processing of these proteins, which in turn results in impaired activation of the PKC1 pathway and finally causes cell death in the absence of osmotic stabilization.

Authors
Lommel, M; Bagnat, M; Strahl, S
MLA Citation
Lommel, M, Bagnat, M, and Strahl, S. "Aberrant processing of the WSC family and Mid2p cell surface sensors results in cell death of Saccharomyces cerevisiae O-mannosylation mutants." Mol Cell Biol 24.1 (January 2004): 46-57.
PMID
14673142
Source
pubmed
Published In
Molecular and Cellular Biology
Volume
24
Issue
1
Publish Date
2004
Start Page
46
End Page
57

Cell surface polarization during yeast mating

Authors
BAGNAT, M
MLA Citation
BAGNAT, M. "Cell surface polarization during yeast mating." Proc. Natl. Acad. Sci. USA 99 (2004): 14183-14188.
Source
cinii-english
Published In
Proc. Natl. Acad. Sci. USA
Volume
99
Publish Date
2004
Start Page
14183
End Page
14188
DOI
10.1073/pnas.172517799

Rvs161p and sphingolipids are required for actin repolarization following salt stress.

In Saccharomyces cerevisiae, the actin cytoskeleton is depolarized by NaCl stress. In this study, the response was maximal after 30 min, and then actin patches repolarized. Rvs161p was required for actin repolarization because the rvs161delta mutant did not repolarize actin patches after growth in a salt medium. Mutations suppressing the rvs161delta-related salt sensitivity all occurred in genes required for sphingolipid biosynthesis: FEN1, SUR4, SUR2, SUR1, and IPT1. These suppressors also suppressed act1-1-related salt sensitivity and the defect in actin repolarization of the rvs161delta mutant, providing a link between sphingolipids and actin polarization. Indeed, deletion of the suppressor genes suppressed the rvs161delta defect in actin repolarization in two ways: either actin was not depolarized at the wild-type level in a set of suppressor mutants, or actin was repolarized in the absence of Rvs161p in the other suppressor mutants. Rvs161p was localized as cortical patches that concentrated at polarization sites, i.e., bud emergence and septa, and was found to be associated with lipid rafts. An important link between sphingolipids and actin polarization is that Rvs161p was required for actin repolarization and was found to be located in lipid rafts.

Authors
Balguerie, A; Bagnat, M; Bonneu, M; Aigle, M; Breton, AM
MLA Citation
Balguerie, A, Bagnat, M, Bonneu, M, Aigle, M, and Breton, AM. "Rvs161p and sphingolipids are required for actin repolarization following salt stress." Eukaryot Cell 1.6 (December 2002): 1021-1031.
PMID
12477802
Source
pubmed
Published In
Eukaryotic cell
Volume
1
Issue
6
Publish Date
2002
Start Page
1021
End Page
1031

Cell surface polarization during yeast mating.

Exposure to mating pheromone in haploid Saccharomyces cerevisiae cells results in the arrest of the cell cycle, expression of mating-specific genes, and polarized growth toward the mating partner. Proteins involved in signaling, polarization, cell adhesion, and fusion are localized to the tip of the mating cell (shmoo) where fusion will eventually occur. The mechanisms ensuring the correct targeting and retention of these proteins are poorly understood. Here we show that in pheromone-treated cells, a reorganization of the plasma membrane involving lipid rafts results in the retention of proteins at the tip of the mating projection, segregated from the rest of the membrane. Sphingolipid and ergosterol biosynthetic mutants fail to polarize proteins to the tip of the shmoo and are deficient in mating. Our results show that membrane microdomain clustering at the mating projection is involved in the generation and maintenance of polarity during mating.

Authors
Bagnat, M; Simons, K
MLA Citation
Bagnat, M, and Simons, K. "Cell surface polarization during yeast mating." Proc Natl Acad Sci U S A 99.22 (October 29, 2002): 14183-14188.
PMID
12374868
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
99
Issue
22
Publish Date
2002
Start Page
14183
End Page
14188
DOI
10.1073/pnas.172517799

Lipid rafts in protein sorting and cell polarity in budding yeast Saccharomyces cerevisiae.

Cellular membranes contain many types and species of lipids. One of the most important functional consequences of this heterogeneity is the existence of microdomains within the plane of the membrane. Sphingolipid acyl chains have the ability of forming tightly packed platforms together with sterols. These platforms or lipid rafts constitute segregation and sorting devices into which proteins specifically associate. In budding yeast, Saccharomyces cerevisiae, lipid rafts serve as sorting platforms for proteins destined to the cell surface. The segregation capacity of rafts also provides the basis for the polarization of proteins at the cell surface during mating. Here we discuss some recent findings that stress the role of lipid rafts as key players in yeast protein sorting and cell polarity.

Authors
Bagnat, M; Simons, K
MLA Citation
Bagnat, M, and Simons, K. "Lipid rafts in protein sorting and cell polarity in budding yeast Saccharomyces cerevisiae." Biol Chem 383.10 (October 2002): 1475-1480.
PMID
12452424
Source
pubmed
Published In
Biological Chemistry
Volume
383
Issue
10
Publish Date
2002
Start Page
1475
End Page
1480
DOI
10.1515/BC.2002.169

Plasma membrane proton ATPase Pma1p requires raft association for surface delivery in yeast.

Correct sorting of proteins is essential to generate and maintain the identity and function of the different cellular compartments. In this study we demonstrate the role of lipid rafts in biosynthetic delivery of Pma1p, the major plasma membrane proton ATPase, to the cell surface. Disruption of rafts led to mistargeting of Pma1p to the vacuole. Conversely, Pma1-7, an ATPase mutant that is mistargeted to the vacuole, was shown to exhibit impaired raft association. One of the previously identified suppressors, multicopy AST1, not only restored surface delivery but also raft association of Pma1-7. Ast1p, which is a peripheral membrane protein, was found to directly interact with Pma1p inducing its clustering into a SDS/Triton X100-resistant oligomer. We suggest that clustering facilitates partition of Pma1p into rafts and transport to the cell surface.

Authors
Bagnat, M; Chang, A; Simons, K
MLA Citation
Bagnat, M, Chang, A, and Simons, K. "Plasma membrane proton ATPase Pma1p requires raft association for surface delivery in yeast." Mol Biol Cell 12.12 (December 2001): 4129-4138.
PMID
11739806
Source
pubmed
Published In
Molecular Biology of the Cell
Volume
12
Issue
12
Publish Date
2001
Start Page
4129
End Page
4138

Plasma membrane proton ATPase Pmalp requires raft association for surface delivery in yeast

Authors
BAGNAT, M
MLA Citation
BAGNAT, M. "Plasma membrane proton ATPase Pmalp requires raft association for surface delivery in yeast." Mol. Biol. Cell. 12 (2001): 4129-4138.
Source
cinii-english
Published In
Mol. Biol. Cell.
Volume
12
Publish Date
2001
Start Page
4129
End Page
4138

Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast.

Lipid rafts, formed by lateral association of sphingolipids and cholesterol, have been implicated in membrane traffic and cell signaling in mammalian cells. Sphingolipids also have been shown to play a role in protein sorting in yeast. Therefore, we wanted to investigate whether lipid rafts exist in yeast and whether these membrane microdomains have an analogous function to their mammalian counterparts. We first developed a protocol for isolating detergent-insoluble glycolipid-enriched complexes (DIGs) from yeast cells. Sequencing of the major protein components of the isolated DIGs by mass spectrometry allowed us to identify, among others, Gas1p, Pma1p, and Nce2p. Using lipid biosynthetic mutants we could demonstrate that conditions that impair the synthesis of sphingolipids and ergosterol also disrupt raft association of Gas1p and Pma1p but not the secretion of acid phosphatase. That endoplasmic reticulum (ER)-to-Golgi transport of Gas1p is blocked in the sphingolipid mutant lcb1-100 raised the question of whether proteins associate with lipid rafts in the ER or later as shown in mammalian cells. Using the sec18-1 mutant we found that DIGs are present already in the ER. Taken together, our results suggest that lipid rafts are involved in the biosynthetic delivery of proteins to the yeast plasma membrane.

Authors
Bagnat, M; Keränen, S; Shevchenko, A; Shevchenko, A; Simons, K
MLA Citation
Bagnat, M, Keränen, S, Shevchenko, A, Shevchenko, A, and Simons, K. "Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast." Proc Natl Acad Sci U S A 97.7 (March 28, 2000): 3254-3259.
PMID
10716729
Source
pubmed
Published In
Proceedings of the National Academy of Sciences of USA
Volume
97
Issue
7
Publish Date
2000
Start Page
3254
End Page
3259
DOI
10.1073/pnas.060034697

Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast

Authors
BAGNAT, M
MLA Citation
BAGNAT, M. "Lipid rafts function in biosynthetic delivery of proteins to the cell surface in yeast." Proc. Natl. Acad. Sci. U.S.A. 97 (2000): 3254-3259.
Source
cinii-english
Published In
Proc. Natl. Acad. Sci. U.S.A.
Volume
97
Publish Date
2000
Start Page
3254
End Page
3259
DOI
10.1073/pnas.060034697

OP18/stathmin binds near the C-terminus of tubulin and facilitates GTP binding.

It is has been previously suggested that the protein Op18/stathmin may interact with tubulin via the alpha-tubulin subunit [Larsson, N., Marklund, U., Melander Gradin, H., Brattsand, G. & Gullberg, M. (1997) Mol. Cell. Biol. 17, 5530-5539]. In this study we have used limited proteolysis and cross-linking analysis to localize further the stathmin-binding site on alpha-tubulin. Our results indicate that such a binding site is in a region close to the C-terminus of the molecule comprising residues 307 to the subtilisin-cleavage site on the alpha-tubulin subunit. Based on a recent model of the structure of tubulin [Nogales, E., Wolf, S.G. & Dowing, D.H. (1998) Nature (London) 391, 199-203], we found that this region contained the same areas that may be involved in longitudinal contacts of alpha-tubulin subunits within the microtubule. We also observed that the binding of stathmin to tubulin can modulate the binding of GTP to tubulin, as a consequence of a conformational change in the beta-tubulin subunit that occurs upon interaction of stathmin with tubulin.

Authors
Moreno, FJ; Bagnat, M; Lim, F; Avila, J
MLA Citation
Moreno, FJ, Bagnat, M, Lim, F, and Avila, J. "OP18/stathmin binds near the C-terminus of tubulin and facilitates GTP binding." Eur J Biochem 262.2 (June 1999): 557-562.
PMID
10336642
Source
pubmed
Published In
European journal of biochemistry / FEBS
Volume
262
Issue
2
Publish Date
1999
Start Page
557
End Page
562
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