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1.
Dev Cell ; 13(5): 743-751, 2007 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-17981141

RESUMO

An important problem in polarized morphogenesis is how polarized transport of membrane vesicles is spatiotemporally regulated. Here, we report that a local change in the transbilayer phospholipid distribution of the plasma membrane regulates the axis of polarized growth. Type 4 P-type ATPases Lem3p-Dnf1p and -Dnf2p are putative heteromeric phospholipid flippases in budding yeast that are localized to polarized sites on the plasma membrane. The lem3Delta mutant exhibits prolonged apical growth due to a defect in the switch to isotropic bud growth. In lem3Delta cells, the small GTPase Cdc42p remains polarized at the bud tip where phosphatidylethanolamine remains exposed on the outer leaflet. Intriguingly, phosphatidylethanolamine and phosphatidylserine stimulate GTPase-activating protein (GAP) activity of Rga1p and Rga2p toward Cdc42p, whereas PI(4,5)P(2) inhibits it. We propose that a redistribution of phospholipids to the inner leaflet of the plasma membrane triggers the dispersal of Cdc42p from the apical growth site, through activation of GAPs.


Assuntos
Subunidades alfa Gs de Proteínas de Ligação ao GTP/fisiologia , Proteínas Ativadoras de GTPase/fisiologia , Fosfolipídeos/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Transportadores de Cassetes de Ligação de ATP , Adenosina Trifosfatases/metabolismo , Membrana Celular/metabolismo , Polaridade Celular , Proliferação de Células , Proteínas de Membrana Transportadoras/metabolismo , Mutação , Fosfatidiletanolaminas/metabolismo , Fosfatidilserinas/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética
2.
Curr Biol ; 18(22): 1719-26, 2008 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-19013066

RESUMO

BACKGROUND: In 1952, Alan Turing suggested that spatial patterns could arise from homogeneous starting conditions by feedback amplification of stochastic fluctuations. One example of such self-organization, called symmetry breaking, involves spontaneous cell polarization in the absence of spatial cues. The conserved GTPase Cdc42p is essential for both guided and spontaneous polarization, and in budding yeast cells Cdc42p concentrates at a single site (the presumptive bud site) at the cortex. Cdc42p concentrates at a random cortical site during symmetry breaking in a manner that requires the scaffold protein Bem1p. The mechanism whereby Bem1p promotes this polarization was unknown. RESULTS: Here we show that Bem1p promotes symmetry breaking by assembling a complex in which both a Cdc42p-directed guanine nucleotide exchange factor (GEF) and a Cdc42p effector p21-activated kinase (PAK) associate with Bem1p. Analysis of Bem1p mutants indicates that both GEF and PAK must bind to the same molecule of Bem1p, and a protein fusion linking the yeast GEF and PAK bypasses the need for Bem1p. Although mammalian cells lack a Bem1p ortholog, they contain more complex multidomain GEFs that in some cases can directly interact with PAKs, and we show that yeast containing an artificial GEF with similar architecture can break symmetry even without Bem1p. CONCLUSIONS: Yeast symmetry-breaking polarization involves a GEF-PAK complex that binds GTP-Cdc42p via the PAK and promotes local Cdc42p GTP-loading via the GEF. By generating fresh GTP-Cdc42p near pre-existing GTP-Cdc42p, the complex amplifies clusters of GTP-Cdc42p at the cortex. Our findings provide mechanistic insight into an evolutionarily conserved pattern-forming positive-feedback pathway.


Assuntos
Polaridade Celular , Fatores de Troca do Nucleotídeo Guanina/fisiologia , Saccharomyces cerevisiae/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Quinases Ativadas por p21/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Fatores de Troca do Nucleotídeo Guanina/química , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Modelos Biológicos , Mutação , Estrutura Terciária de Proteína , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/citologia , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/química , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo , Quinases Ativadas por p21/química , Quinases Ativadas por p21/metabolismo
3.
Trends Cell Biol ; 13(8): 403-9, 2003 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-12888292

RESUMO

Septins are a conserved eukaryotic family of GTP-binding filament-forming proteins with functions in cytokinesis and other processes. In the budding yeast Saccharomyces cerevisiae, septins initially localize to the presumptive bud site and then to the cortex of the mother-bud neck as an hourglass structure. During cytokinesis, the septin hourglass splits and single septin rings partition with each of the resulting cells. Septins are thought to function in diverse processes in S. cerevisiae, mainly by acting as a scaffold to direct the neck localization of septin-associated proteins.


Assuntos
Proteínas do Citoesqueleto/fisiologia , Saccharomycetales/fisiologia , Proteínas de Ciclo Celular/fisiologia , Divisão Celular/fisiologia , Proteínas do Citoesqueleto/genética , Exocitose/fisiologia , Proteínas de Ligação ao GTP/fisiologia , Modelos Biológicos , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/fisiologia , Saccharomycetales/citologia , Saccharomycetales/genética , Fuso Acromático/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
4.
J Cell Biol ; 155(2): 261-70, 2001 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-11604421

RESUMO

The establishment of cell polarity in budding yeast involves assembly of actin filaments at specified cortical domains. Elucidation of the underlying mechanism requires an understanding of the machinery that controls actin polymerization and how this machinery is in turn controlled by signaling proteins that respond to polarity cues. We showed previously that the yeast orthologue of the Wiskott-Aldrich Syndrome protein, Bee1/Las17p, and the type I myosins are key regulators of cortical actin polymerization. Here, we demonstrate further that these proteins together with Vrp1p form a multivalent Arp2/3-activating complex. During cell polarization, a bifurcated signaling pathway downstream of the Rho-type GTPase Cdc42p recruits and activates this complex, leading to local assembly of actin filaments. One branch, which requires formin homologues, mediates the recruitment of the Bee1p complex to the cortical site where the activated Cdc42p resides. The other is mediated by the p21-activated kinases, which activate the motor activity of myosin-I through phosphorylation. Together, these findings provide insights into the essential processes leading to polarization of the actin cytoskeleton.


Assuntos
Actinas/metabolismo , Polaridade Celular , Proteínas do Citoesqueleto , Proteínas Fúngicas/metabolismo , Proteínas de Saccharomyces cerevisiae , Leveduras/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Proteína 2 Relacionada a Actina , Proteína 3 Relacionada a Actina , Citoesqueleto/metabolismo , Substâncias Macromoleculares , Proteínas dos Microfilamentos/metabolismo , Modelos Biológicos , Proteínas Motores Moleculares/metabolismo , Miosina Tipo I/metabolismo , Proteínas do Tecido Nervoso/metabolismo , Fosforilação , Transporte Proteico , Proteína da Síndrome de Wiskott-Aldrich
5.
J Cell Biol ; 155(4): 581-92, 2001 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-11706050

RESUMO

The Rho family GTPase Cdc42 is a key regulator of cell polarity and cytoskeletal organization in eukaryotic cells. In yeast, the role of Cdc42 in polarization of cell growth includes polarization of the actin cytoskeleton, which delivers secretory vesicles to growth sites at the plasma membrane. We now describe a novel temperature-sensitive mutant, cdc42-6, that reveals a role for Cdc42 in docking and fusion of secretory vesicles that is independent of its role in actin polarization. cdc42-6 mutants can polarize actin and deliver secretory vesicles to the bud, but fail to fuse those vesicles with the plasma membrane. This defect is manifested only during the early stages of bud formation when growth is most highly polarized, and appears to reflect a requirement for Cdc42 to maintain maximally active exocytic machinery at sites of high vesicle throughput. Extensive genetic interactions between cdc42-6 and mutations in exocytic components support this hypothesis, and indicate a functional overlap with Rho3, which also regulates both actin organization and exocytosis. Localization data suggest that the defect in cdc42-6 cells is not at the level of the localization of the exocytic apparatus. Rather, we suggest that Cdc42 acts as an allosteric regulator of the vesicle docking and fusion apparatus to provide maximal function at sites of polarized growth.


Assuntos
Exocitose/fisiologia , Proteínas de Saccharomyces cerevisiae , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Alelos , Ciclo Celular , Divisão Celular , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Genes Fúngicos , Glucana Endo-1,3-beta-D-Glucosidase/metabolismo , Complexo de Golgi/metabolismo , Mutação Puntual , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo , Proteínas rho de Ligação ao GTP/genética , Proteínas rho de Ligação ao GTP/metabolismo
6.
Mol Biol Cell ; 17(6): 2824-38, 2006 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-16571678

RESUMO

The highly conserved small Rho G-protein, Cdc42p plays a critical role in cell polarity and cytoskeleton organization in all eukaryotes. In the yeast Saccharomyces cerevisiae, Cdc42p is important for cell polarity establishment, septin ring assembly, and pheromone-dependent MAP-kinase signaling during the yeast mating process. In this study, we further investigated the role of Cdc42p in the mating process by screening for specific mating defective cdc42 alleles. We have identified and characterized novel mating defective cdc42 alleles that are unaffected in vegetative cell polarity. Replacement of the Cdc42p Val36 residue with Met resulted in a specific cell fusion defect. This cdc42[V36M] mutant responded to mating pheromone but was defective in cell fusion and in localization of the cell fusion protein Fus1p, similar to a previously isolated cdc24 (cdc24-m6) mutant. Overexpression of a fast cycling Cdc42p mutant suppressed the cdc24-m6 fusion defect and conversely, overexpression of Cdc24p suppressed the cdc42[V36M] fusion defect. Taken together, our results indicate that Cdc42p GDP-GTP cycling is critical for efficient cell fusion.


Assuntos
Guanosina Difosfato/metabolismo , Guanosina Trifosfato/metabolismo , Saccharomyces cerevisiae/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Sequência de Aminoácidos , Fusão Celular , Polaridade Celular , Genótipo , Dados de Sequência Molecular , Mutagênese , Plasmídeos , Saccharomyces cerevisiae/ultraestrutura , Proteínas de Saccharomyces cerevisiae/fisiologia
7.
Mol Biol Cell ; 11(1): 339-54, 2000 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-10637312

RESUMO

Cdc42p, a Rho family GTPase of the Ras superfamily, is a key regulator of cell polarity and morphogenesis in eukaryotes. Using 37 site-directed cdc42 mutants, we explored the functions and interactions of Cdc42p in the budding yeast Saccharomyces cerevisiae. Cytological and genetic analyses of these cdc42 mutants revealed novel and diverse phenotypes, showing that Cdc42p possesses at least two distinct essential functions and acts as a nodal point of cell polarity regulation in vivo. In addition, mapping the functional data for each cdc42 mutation onto a structural model of the protein revealed as functionally important a surface of Cdc42p that is distinct from the canonical protein-interacting domains (switch I, switch II, and the C terminus) identified previously in members of the Ras superfamily. This region overlaps with a region (alpha5-helix) recently predicted by structural models to be a specificity determinant for Cdc42p-protein interactions.


Assuntos
Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Sequência de Aminoácidos , Sítios de Ligação , Humanos , Modelos Moleculares , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Fenótipo , Conformação Proteica , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/química , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética
8.
Mol Biol Cell ; 11(2): 647-61, 2000 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-10679021

RESUMO

Aip3p/Bud6p is a regulator of cell and cytoskeletal polarity in Saccharomyces cerevisiae that was previously identified as an actin-interacting protein. Actin-interacting protein 3 (Aip3p) localizes at the cell cortex where cytoskeleton assembly must be achieved to execute polarized cell growth, and deletion of AIP3 causes gross defects in cell and cytoskeletal polarity. We have discovered that Aip3p localization is mediated by the secretory pathway. Mutations in early- or late-acting components of the secretory apparatus lead to Aip3p mislocalization. Biochemical data show that a pool of Aip3p is associated with post-Golgi secretory vesicles. An investigation of the sequences within Aip3p necessary for Aip3p localization has identified a sequence within the N terminus of Aip3p that is sufficient for directing Aip3p localization. Replacement of the N terminus of Aip3p with a homologous region from a Schizosaccharomyces pombe protein allows for normal Aip3p localization, indicating that the secretory pathway-mediated Aip3p localization pathway is conserved. Delivery of Aip3p also requires the type V myosin motor Myo2p and its regulatory light-chain calmodulin. These data suggest that one function of calmodulin is to activate Myo2p's activity in the secretory pathway; this function is likely the polarized movement of late secretory vesicles and associated Aip3p on actin cables.


Assuntos
Polaridade Celular , Proteínas Fúngicas/metabolismo , Proteínas dos Microfilamentos/metabolismo , Cadeias Pesadas de Miosina , Miosina Tipo II , Miosina Tipo V , Miosinas/metabolismo , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/metabolismo , Proteínas de Schizosaccharomyces pombe , Sequência de Aminoácidos , Transporte Biológico , Calmodulina/genética , Calmodulina/fisiologia , Proteínas de Transporte/genética , Proteínas de Transporte/fisiologia , Sequência Conservada/genética , Proteínas Fúngicas/química , Proteínas Fúngicas/genética , Proteínas Fúngicas/fisiologia , Genes Fúngicos/genética , Genes Fúngicos/fisiologia , Complexo de Golgi/fisiologia , Proteínas dos Microfilamentos/química , Proteínas dos Microfilamentos/genética , Microssomos/metabolismo , Dados de Sequência Molecular , Mutação/genética , Miosinas/química , Miosinas/fisiologia , Sinais Direcionadores de Proteínas/química , Sinais Direcionadores de Proteínas/genética , Sinais Direcionadores de Proteínas/metabolismo , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Proteínas Recombinantes de Fusão/metabolismo , Saccharomyces cerevisiae/genética , Schizosaccharomyces/genética , Alinhamento de Sequência , Temperatura , Fatores de Tempo , Vacúolos/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
10.
Curr Opin Microbiol ; 5(2): 179-86, 2002 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-11934615

RESUMO

Polarized growth involves a hierarchy of events such as selection of the growth site, polarization of the cytoskeleton to the selected growth site, and transport of secretory vesicles containing components required for growth. The budding yeast Saccharomyces cerevisiae is an excellent model system for the study of polarized cell growth. A large number of proteins have been found to be involved in these processes, although their mechanisms of action are not yet well-understood. Recent discoveries have helped elucidate many of the processes involved in cell polarity and bud-site selection in yeast and have modified the traditional view of cellular structures involved in these processes. This review focuses on recent advances on the roles of cortical tags, GTPases and the cytoskeleton in the generation and maintenance of cell polarity in yeast.


Assuntos
Polaridade Celular , Saccharomycetales/crescimento & desenvolvimento , Actinas/genética , Actinas/fisiologia , Citoesqueleto/química , Saccharomycetales/química , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
11.
Eur J Cell Biol ; 84(12): 939-49, 2005 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-16325503

RESUMO

Cdc42, a member of the Rho subfamily of small GTPases, is highly conserved in both sequence and function across eukaryotic species. In budding yeast, Cdc42 triggers polarized growth necessary for bud emergence via rearrangement of the actin cytoskeleton. It has been shown that the role of Cdc42 in bud emergence requires both Cdc28-Cln (G1) kinase and the passage through START. In this report, we show that Cdc42 also serves an essential function in the establishment of bud site prior to START by catalyzing the translocation of bud-site components such as Spa2 to the cell cortex. Our analysis of various conditional alleles of CDC42 suggests that these two functions (bud site establishment and bud emergence) are genetically separable. Surprisingly, the role of Cdc42 in the cortical localization of Spa2 appears to be independent of its well known GTP/GDP exchange factor Cdc24. We also provide evidence that this role of Cdc42 requires the function of the COPI coatomer complex.


Assuntos
Proteínas de Ciclo Celular/metabolismo , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Alelos , Ciclo Celular/fisiologia , Proteínas de Ciclo Celular/genética , Núcleo Celular/química , Polaridade Celular , Complexo I de Proteína do Envoltório/fisiologia , Proteínas do Citoesqueleto , Citoesqueleto/fisiologia , Regulação Fúngica da Expressão Gênica , Genes Fúngicos , Fatores de Troca do Nucleotídeo Guanina/genética , Microscopia Confocal , Mutação , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/crescimento & desenvolvimento , Proteínas de Saccharomyces cerevisiae/genética , Temperatura , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética
12.
J Cell Biol ; 211(1): 19-26, 2015 Oct 12.
Artigo em Inglês | MEDLINE | ID: mdl-26459595

RESUMO

Establishment of cell polarity in animal and fungal cells involves localization of the conserved Rho-family guanosine triphosphatase, Cdc42, to the cortical region destined to become the "front" of the cell. The high local concentration of active Cdc42 promotes cytoskeletal polarization through various effectors. Cdc42 accumulation at the front is thought to involve positive feedback, and studies in the budding yeast Saccharomyces cerevisiae have suggested distinct positive feedback mechanisms. One class of mechanisms involves localized activation of Cdc42 at the front, whereas another class involves localized delivery of Cdc42 to the front. Here we show that Cdc42 activation must be localized for successful polarity establishment, supporting local activation rather than local delivery as the dominant mechanism in this system.


Assuntos
Saccharomyces cerevisiae/enzimologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas de Ciclo Celular/metabolismo , Polaridade Celular , Ativação Enzimática , Fatores de Troca do Nucleotídeo Guanina/metabolismo , Transporte Proteico , Saccharomyces cerevisiae/citologia , Proteínas de Saccharomyces cerevisiae/metabolismo
13.
Elife ; 2: e01085, 2013 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-24286829

RESUMO

Septins are guanine nucleotide-binding proteins that polymerize into filamentous and higher-order structures. Cdc42 and its effector Gic1 are involved in septin recruitment, ring formation and dissociation. The regulatory mechanisms behind these processes are not well understood. Here, we have used electron microscopy and cryo electron tomography to elucidate the structural basis of the Gic1-septin and Gic1-Cdc42-septin interaction. We show that Gic1 acts as a scaffolding protein for septin filaments forming long and flexible filament cables. Cdc42 in its GTP-form binds to Gic1, which ultimately leads to the dissociation of Gic1 from the filament cables. Surprisingly, Cdc42-GDP is not inactive, but in the absence of Gic1 directly interacts with septin filaments resulting in their disassembly. We suggest that this unanticipated dual function of Cdc42 is crucial for the cell cycle. Based on our results we propose a novel regulatory mechanism for septin filament formation and dissociation. DOI: http://dx.doi.org/10.7554/eLife.01085.001.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Septinas/biossíntese , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Microscopia Eletrônica , Saccharomyces cerevisiae/fisiologia , Septinas/metabolismo
14.
Curr Biol ; 23(1): 42-7, 2013 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-23200991

RESUMO

How cells polarize in response to external cues is a fundamental biological problem. For mating, yeast cells orient growth toward the source of a pheromone gradient produced by cells of the opposite mating type. Polarized growth depends on the small GTPase Cdc42, a central eukaryotic polarity regulator that controls signaling, cytoskeleton polarization, and vesicle trafficking. However, the mechanisms of polarity establishment and mate selection in complex cellular environments are poorly understood. Here we show that, in fission yeast, low-level pheromone signaling promotes a novel polarization state, where active Cdc42, its GEF Scd1, and scaffold Scd2 form colocalizing dynamic zones that sample the periphery of the cell. Two direct Cdc42 effectors--actin cables marked by myosin V Myo52 and the exocyst complex labeled by Sec6 and Sec8--also dynamically colocalize with active Cdc42. However, these cells do not grow due to a block in the exocytosis of cell wall synthases Bgs1 and Bgs4. High-level pheromone stabilizes active Cdc42 zones and promotes cell wall synthase exocytosis and polarized growth. However, in the absence of prior low-level pheromone signaling, exploration fails, and cells polarize growth at cell poles by default. Consequently, these cells show altered partner choice, mating preferentially with sister rather than nonsister cells. Thus, Cdc42 exploration serves to orient growth for partner selection. This process may also promote genetic diversification.


Assuntos
Schizosaccharomyces/citologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Polaridade Celular , Feromônios/metabolismo , Feromônios/fisiologia , Schizosaccharomyces/fisiologia , Diferenciação Sexual , Transdução de Sinais , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo
15.
Curr Biol ; 23(1): 32-41, 2013 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-23200992

RESUMO

BACKGROUND: Many cells are remarkably proficient at tracking very shallow chemical gradients, despite considerable noise from stochastic receptor-ligand interactions. Motile cells appear to undergo a biased random walk: spatial noise in receptor activity may determine the instantaneous direction, but because noise is spatially unbiased, it is filtered out by time averaging, resulting in net movement upgradient. How nonmotile cells might filter out noise is unknown. RESULTS: Using yeast chemotropic mating as a model, we demonstrate that a polarized patch of polarity regulators "wanders" along the cortex during gradient tracking. Computational and experimental findings suggest that actin-directed membrane traffic contributes to wandering by diluting local polarity factors. The pheromone gradient appears to bias wandering via interactions between receptor-activated Gßγ and polarity regulators. Artificially blocking patch wandering impairs gradient tracking. CONCLUSIONS: We suggest that the polarity patch undergoes an intracellular biased random walk that enables noise filtering by time averaging, allowing nonmotile cells to track shallow gradients.


Assuntos
Polaridade Celular , Saccharomyces cerevisiae/citologia , Actinas/metabolismo , Actinas/fisiologia , Quimiotaxia , Feromônios/fisiologia , Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/fisiologia , Proteínas de Saccharomyces cerevisiae/análise , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/fisiologia , Fatores de Tempo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/análise , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
16.
Curr Biol ; 23(1): R10-2, 2013 Jan 07.
Artigo em Inglês | MEDLINE | ID: mdl-23305660

RESUMO

Chemical gradients are used by cells to provide positional information. Two new studies reveal that polarity proteins are highly dynamic in yeast cells responding to a pheromone gradient and suggest that this behavior is important for robust directional growth.


Assuntos
Polaridade Celular , Saccharomyces cerevisiae/citologia , Schizosaccharomyces/citologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
17.
Nat Cell Biol ; 13(12): 1424-30, 2011 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-21964439

RESUMO

Polarity is key to the function of eukaryotic cells. On the establishment of a polarity axis, cells can vectorially target secretion, generating an asymmetric distribution of plasma membrane proteins. From Saccharomyces cerevisiae to mammals, the small GTPase Cdc42 is a pivotal regulator of polarity. We used a fluorescent probe to visualize the distribution of phosphatidylserine in live S. cerevisiae. Remarkably, phosphatidylserine was polarized in the plasma membrane, accumulating in bud necks, the bud cortex and the tips of mating projections. Polarization required vectorial delivery of phosphatidylserine-containing secretory vesicles, and phosphatidylserine was largely excluded from endocytic vesicles, contributing to its polarized retention. Mutants lacking phosphatidylserine synthase had impaired polarization of the Cdc42 complex, leading to a delay in bud emergence, and defective mating. The addition of lysophosphatidylserine resulted in resynthesis and polarization of phosphatidylserine, as well as repolarization of Cdc42. The results indicate that phosphatidylserine--and presumably its polarization--are required for optimal Cdc42 targeting and activation during cell division and mating.


Assuntos
Polaridade Celular/fisiologia , Fosfatidilserinas/metabolismo , Saccharomyces cerevisiae/citologia , Saccharomyces cerevisiae/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo , CDPdiacilglicerol-Serina O-Fosfatidiltransferase/genética , CDPdiacilglicerol-Serina O-Fosfatidiltransferase/metabolismo , Divisão Celular/fisiologia , Mutagênese/fisiologia , Saccharomyces cerevisiae/enzimologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia
18.
Curr Biol ; 21(3): 184-94, 2011 Feb 08.
Artigo em Inglês | MEDLINE | ID: mdl-21277209

RESUMO

BACKGROUND: Polarization in yeast has been proposed to involve a positive feedback loop whereby the polarity regulator Cdc42p orients actin cables, which deliver vesicles carrying Cdc42p to the polarization site. Previous mathematical models treating Cdc42p traffic as a membrane-free flux suggested that directed traffic would polarize Cdc42p, but it remained unclear whether Cdc42p would become polarized without the membrane-free simplifying assumption. RESULTS: We present mathematical models that explicitly consider stochastic vesicle traffic via exocytosis and endocytosis, providing several new insights. Our findings suggest that endocytic cargo influences the timing of vesicle internalization in yeast. Moreover, our models provide quantitative support for the view that integral membrane cargo proteins would become polarized by directed vesicle traffic given the experimentally determined rates of vesicle traffic and diffusion. However, such traffic cannot effectively polarize the more rapidly diffusing Cdc42p in the model without making additional assumptions that seem implausible and lack experimental support. CONCLUSIONS: Our findings suggest that actin-directed vesicle traffic would perturb, rather than reinforce, polarization in yeast.


Assuntos
Polaridade Celular , Modelos Biológicos , Saccharomyces cerevisiae/citologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Citoesqueleto de Actina/química , Citoesqueleto de Actina/ultraestrutura , Endocitose , Exocitose , Retroalimentação Fisiológica , Transporte Proteico , Proteínas SNARE/metabolismo , Septinas/análise , Septinas/metabolismo , Septinas/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/genética , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/metabolismo
19.
FEBS J ; 276(24): 7253-64, 2009 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-20050180

RESUMO

The conserved Rho-type GTPase Cdc42p is a key regulator of signal transduction and polarity in eukaryotic cells. In the yeast Saccharomyces cerevisiae, Cdc42p promotes polarized growth through the p21-activated kinases Ste20p and Cla4p. Previously, we demonstrated that Ste20p forms a complex with Erg4p, Cbr1p and Ncp1p, which all catalyze important steps in sterol biosynthesis. CLA4 interacts genetically with ERG4 and NCP1. Furthermore, Erg4p, Ncp1p and Cbr1p play important roles in cell polarization during vegetative growth, mating and filamentation. As Ste20p and Cla4p are involved in these processes it seems likely that sterol biosynthetic enzymes and p21-activated kinases act in related pathways. Here, we demonstrate that the deletion of either STE20 or CLA4 results in increased levels of sterols. In addition, higher concentrations of steryl esters, the storage form of sterols, were observed in cla4Delta cells. CLA4 expression from a multicopy plasmid reduces enzyme activity of Are2p, the major steryl ester synthase, under aerobic conditions. Altogether, our data suggest that Ste20p and Cla4p may function as negative modulators of sterol biosynthesis. Moreover, Cla4p has a negative effect on steryl ester formation. As sterol homeostasis is crucial for cell polarization, Ste20p and Cla4p may regulate cell polarity in part through the modulation of sterol homeostasis.


Assuntos
Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Proteínas Serina-Treonina Quinases/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/crescimento & desenvolvimento , Esteróis/metabolismo , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Polaridade Celular/efeitos dos fármacos , Homeostase , Peptídeos e Proteínas de Sinalização Intracelular/genética , MAP Quinase Quinase Quinases , Proteínas Serina-Treonina Quinases/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética , Esterol O-Aciltransferase/fisiologia
20.
Traffic ; 7(9): 1224-42, 2006 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-17004323

RESUMO

Polarized cell growth requires the establishment of an axis of growth along which secretion can be targeted to a specific site on the cell cortex. How polarity establishment and secretion are choreographed is not fully understood, though Rho GTPase- and Rab GTPase-mediated signaling is required. Superimposed on this regulation are the functions of specific lipids and their cognate binding proteins. In a screen for Saccharomyces cerevisiae genes that interact with Rho family CDC42 to promote polarity establishment, we identified KES1/OSH4, which encodes a homologue of mammalian oxysterol-binding protein (OSBP). Other yeast OSH genes (OSBP homologues) had comparable genetic interactions with CDC42, implicating OSH genes in the regulation of CDC42-dependent polarity establishment. We found that the OSH gene family (OSH1-OSH7) promotes cell polarization by maintaining the proper localization of septins, the Rho GTPases Cdc42p and Rho1p, and the Rab GTPase Sec4p. Disruption of all OSH gene function caused specific defects in polarized exocytosis, indicating that the Osh proteins are collectively required for a secretory pathway implicated in the maintenance of polarized growth.


Assuntos
Polaridade Celular/fisiologia , Proteínas de Membrana/fisiologia , Proteínas de Saccharomyces cerevisiae/fisiologia , Saccharomyces cerevisiae/fisiologia , Proteína cdc42 de Saccharomyces cerevisiae de Ligação ao GTP/fisiologia , Proteínas rho de Ligação ao GTP/fisiologia , Polaridade Celular/genética , Humanos , Proteínas de Membrana/genética , Mutação , Receptores de Esteroides/genética , Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/genética
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