scholarly journals Activation of Cdc42 GTPase upon CRY2-Induced Cortical Recruitment Is Antagonized by GAPs in Fission Yeast

Cells ◽  
2020 ◽  
Vol 9 (9) ◽  
pp. 2089 ◽  
Author(s):  
Iker Lamas ◽  
Nathalie Weber ◽  
Sophie G. Martin
Keyword(s):  
Fission Yeast ◽  
Positive Feedback ◽  
Antagonistic Activity ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Cell Architecture

The small GTPase Cdc42 is critical for cell polarization in eukaryotic cells. In rod-shaped fission yeast Schizosaccharomyces pombe cells, active GTP-bound Cdc42 promotes polarized growth at cell poles, while inactive Cdc42-GDP localizes ubiquitously also along cell sides. Zones of Cdc42 activity are maintained by positive feedback amplification involving the formation of a complex between Cdc42-GTP, the scaffold Scd2, and the guanine nucleotide exchange factor (GEF) Scd1, which promotes the activation of more Cdc42. Here, we use the CRY2-CIB1 optogenetic system to recruit and cluster a cytosolic Cdc42 variant at the plasma membrane and show that this leads to its moderate activation also on cell sides. Surprisingly, Scd2, which binds Cdc42-GTP, is still recruited to CRY2-Cdc42 clusters at cell sides in individual deletion of the GEFs Scd1 or Gef1. We show that activated Cdc42 clusters at cell sides are able to recruit Scd1, dependent on the scaffold Scd2. However, Cdc42 activity is not amplified by positive feedback and does not lead to morphogenetic changes, due to antagonistic activity of the GTPase activating protein Rga4. Thus, the cell architecture is robust to moderate activation of Cdc42 at cell sides.

scholarly journals Clustering-mediated activation of Cdc42 GTPase antagonized by GAPs in fission yeast

2020 ◽  
Author(s):  
Iker Lamas ◽  
Nathalie Weber ◽  
Sophie G Martin
Keyword(s):  
Fission Yeast ◽  
Positive Feedback ◽  
Antagonistic Activity ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Cell Architecture

AbstractThe small GTPase Cdc42 is critical for cell polarization in eukaryotic cells. In rod-shaped fission yeast Schizosaccharomyces pombe cells, active GTP-bound Cdc42 promotes polarized growth at cell poles, while inactive Cdc42-GDP localizes ubiquitously also along cell sides. Zones of Cdc42 activity are maintained by positive feedback amplification involving the formation of a complex between Cdc42-GTP, the scaffold Scd2 and the guanine nucleotide exchange factor (GEF) Scd1, which promotes the activation of more Cdc42. Here, we use the CRY2-CIB1 optogenetic system to recruit and cluster a cytosolic Cdc42 allele at the plasma membrane and show that this leads to its moderate activation also on cell sides. Surprisingly, activation of CRY2-Cdc42 does not individually depend on Scd1 or the GEF Gef1. We show that activated Cdc42 clusters at cell sides are able to recruit Scd1, dependent on the scaffold Scd2. However, Cdc42 activity is not amplified by positive feedback and does not lead to morphogenetic changes, due to antagonistic activity of the GTPase activating protein Rga4 on cell sides. Thus, the cell architecture is robust to moderate activation of Cdc42 at cell sides.

scholarly journals PLEKHG3 enhances polarized cell migration by activating actin filaments at the cell front

2016 ◽  
Vol 113 (36) ◽  
pp. 10091-10096 ◽  
Author(s):  
Trang Thi Thu Nguyen ◽  
Wei Sun Park ◽  
Byung Ouk Park ◽  
Cha Yeon Kim ◽  
Yohan Oh ◽  
...  
Keyword(s):  
Positive Feedback ◽  
Actin Polymerization ◽  
Leading Edge ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Cell Front ◽  
Control Protein

Cells migrate by directing Ras-related C3 botulinum toxin substrate 1 (Rac1) and cell division control protein 42 (Cdc42) activities and by polymerizing actin toward the leading edge of the cell. Previous studies have proposed that this polarization process requires a local positive feedback in the leading edge involving Rac small GTPase and actin polymerization with PI3K likely playing a coordinating role. Here, we show that the pleckstrin homology and RhoGEF domain containing G3 (PLEKHG3) is a PI3K-regulated Rho guanine nucleotide exchange factor (RhoGEF) for Rac1 and Cdc42 that selectively binds to newly polymerized actin at the leading edge of migrating fibroblasts. Optogenetic inactivation of PLEKHG3 showed that PLEKHG3 is indispensable both for inducing and for maintaining cell polarity. By selectively binding to newly polymerized actin, PLEKHG3 promotes local Rac1/Cdc42 activation to induce more local actin polymerization, which in turn promotes the recruitment of more PLEKHG3 to induce and maintain cell front. Thus, autocatalytic reinforcement of PLEKHG3 localization to the leading edge of the cell provides a molecular basis for the proposed positive feedback loop that is required for cell polarization and directed migration.

scholarly journals cTAGE5 acts as a Sar1 GTPase regulator for collagen export

2018 ◽  
Author(s):  
Norito Sasaki ◽  
Masano Shiraiwa ◽  
Miharu Maeda ◽  
Tomohiro Yorimitsu ◽  
Ken Sato ◽  
...  
Keyword(s):  
Small Gtpase ◽  
Coated Vesicles ◽  
Secretory Proteins ◽  
Efficient Production ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
Collagen Secretion ◽  
Nucleotide Exchange Factor ◽  
Er Exit Sites

AbstractSecretory proteins synthesized within the endoplasmic reticulum (ER) are exported via coat protein complex II (COPII)-coated vesicles. The formation of the COPII-coated vesicles is initiated by activation of the small GTPase, Sar1. cTAGE5 directly interacts with a guanine-nucleotide exchange factor (GEF), Sec12, and a GTPase-activating protein (GAP) of Sar1, Sec23. We have previously shown that cTAGE5 recruits Sec12 to the ER exit sites for efficient production of activated Sar1 for collagen secretion. However, the functional significance of the interaction between cTAGE5 and Sec23 has not been fully elucidated. In this study, we showed that cTAGE5 enhances the GAP activity of Sec23 toward Sar1. In addition, the interaction of cTAGE5 with Sec23 is necessary for collagen exit from the ER. Our data suggests that cTAGE5 acts as a Sar1 GTPase regulator for collagen secretion.

scholarly journals Eng2, a new player involved in feedback loop regulation of Cdc42 activity in fission yeast

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Patricia García ◽  
Pedro M. Coll ◽  
Francisco del Rey ◽  
M. Isabel Geli ◽  
Pilar Pérez ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Feedback Loop ◽  
Oscillatory Behavior ◽  
Small Gtpase ◽  
Dual Function ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Endocytic Machinery ◽  
Mutant Cells

AbstractCell polarity and morphogenesis are regulated by the small GTPase Cdc42. Even though major advances have been done in the field during the last years, the molecular details leading to its activation in particular cellular contexts are not completely understood. In fission yeast, the β(1,3)-glucanase Eng2 is a “moonlighting protein” with a dual function, acting as a hydrolase during spore dehiscence, and as component of the endocytic machinery in vegetative cells. Here, we report that Eng2 plays a role in Cdc42 activation during polarized growth through its interaction with the scaffold protein Scd2, which brings Cdc42 together with its guanine nucleotide exchange factor (GEF) Scd1. eng2Δ mutant cells have defects in activation of the bipolar growth (NETO), remaining monopolar during all the cell cycle. In the absence of Eng2 the accumulation of Scd1 and Scd2 at the poles is reduced, the levels of Cdc42 activation decrease, and the Cdc42 oscillatory behavior, associated with bipolar growth in wild type cells, is altered. Furthermore, overexpression of Eng2 partially rescues the growth and polarity defects of a cdc42-L160S mutant. Altogether, our work unveils a new factor regulating the activity of Cdc42, which could potentially link the polarity and endocytic machineries.

scholarly journals Spatial control of Cdc42 activation determines cell width in fission yeast

2011 ◽  
Vol 22 (20) ◽  
pp. 3801-3811 ◽  
Author(s):  
Felice D. Kelly ◽  
Paul Nurse
Keyword(s):  
Fission Yeast ◽  
Constant Width ◽  
Growth Zone ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Cell Width ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Genome Wide ◽  
Ideal System

The fission yeast Schizosaccharomyces pombe is a rod-shaped cell that grows by linear extension at the cell tips, with a nearly constant width throughout the cell cycle. This simple geometry makes it an ideal system for studying the control of cellular dimensions. In this study, we carried out a near-genome-wide screen for mutants wider than wild-type cells. We found 11 deletion mutants that were wider; seven of the deleted genes are implicated in the control of the small GTPase Cdc42, including the Cdc42 guanine nucleotide exchange factor (GEF) Scd1 and the Cdc42 GTPase-activating protein (GAP) Rga4. Deletions of rga4 and scd1 had additive effects on cell width, and the proteins localized independently of one another, with Rga4 located at the cell sides and Scd1 at the cell tips. Activated Cdc42 localization is altered in rga4Δ, scd1Δ, and scd2Δ mutants. Delocalization and ectopic retargeting experiments showed that the localizations of Rga4 and Scd1 are crucial for their roles in determining cell width. We propose that the GAP Rga4 and the GEF Scd1 establish a gradient of activated Cdc42 within the cellular tip plasma membrane, and it is this gradient that determines cell growth-zone size and normal cell width.

scholarly journals The Identification of cDNAs That Affect the Mitosis-to-Interphase Transition in Schizosaccharomyces pombe, Including sbp1, Which Encodes a spi1p-GTP–Binding Protein

Genetics ◽  
1998 ◽  
Vol 148 (2) ◽  
pp. 645-656
Author(s):  
Xiangwei He ◽  
Naoyuki Hayashi ◽  
Nathan G Walcott ◽  
Yoshiaki Azuma ◽  
Thomas E Patterson ◽  
...  
Keyword(s):  
Fission Yeast ◽  
Genetic Interaction ◽  
Binding Protein ◽  
Medial Septum ◽  
Regulatory Proteins ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
System A ◽  
Nucleotide Exchange Factor

Abstract Perturbations of the spi1p GTPase system in fission yeast, caused by mutation or overexpression of several regulatory proteins, result in a unique terminal phenotype that includes condensed chromosomes, a wide medial septum, and a fragmented nuclear envelope. To identify potential regulators or targets of the spi1p GTPase system, a screen for cDNAs whose overexpression results in this terminal phenotype was conducted, and seven clones that represent three genes, named med1, med2, and med3 (mitotic exit defect), were identified. Their genetic interaction with the spi1p GTPase system was established by showing that the spi1p guanine nucleotide exchange factor mutant pim1-d1ts was hypersensitive to their overexpression. med1 encodes a homologue of the human Ran-binding protein, RanBP1, and has been renamed sbp1 (spi1-binding protein). sbp1p binds to spi1p-GTP and costimulates the GTPase-activating protein (GAP)-catalyzed GTPase activity. Cells in which sbp1p is depleted or overproduced phenocopy cells in which the balance between spi1p-GTP and spi1p-GDP is perturbed by other means. Therefore, sbp1p mediates and/or regulates the essential functions of the spi1p GTPase system. med2 and med3 encode novel fission yeast proteins that, based on our phenotypic analyses, are likely to identify additional regulators or effectors of the spi1p GTPase system.

scholarly journals A Two-Tiered Mechanism Enables Localized Cdc42 Signaling during Enterocyte Polarization

2017 ◽  
Vol 37 (7) ◽  
Author(s):  
Lucas J. M. Bruurs ◽  
Susan Zwakenberg ◽  
Mirjam C. van der Net ◽  
Fried J. Zwartkruis ◽  
Johannes L. Bos
Keyword(s):  
Molecular Mechanisms ◽  
Apical Membrane ◽  
Cell Polarization ◽  
Small Gtpase ◽  
Nucleotide Exchange ◽  
Differential Diffusion ◽  
Cellular Processes ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Dual Mechanism

ABSTRACT Signaling by the small GTPase Cdc42 governs a diverse set of cellular processes that contribute to tissue morphogenesis. Since these processes often require highly localized signaling, Cdc42 activity must be clustered in order to prevent ectopic signaling. During cell polarization, apical Cdc42 signaling directs the positioning of the nascent apical membrane. However, the molecular mechanisms that drive Cdc42 clustering during polarity establishment are largely unknown. Here, we demonstrate that during cell polarization localized Cdc42 signaling is enabled via activity-dependent control of Cdc42 mobility. By performing photoconversion experiments, we show that inactive Cdc42-GDP is 30-fold more mobile than active Cdc42-GTP. This switch in apical mobility originates from a dual mechanism involving RhoGDI-mediated membrane dissociation of Cdc42-GDP and Tuba-mediated immobilization of Cdc42-GTP. Interference with either mechanism affects Cdc42 clustering and as a consequence impairs Cdc42-mediated apical membrane clustering. We therefore identify a molecular network, comprised of Cdc42, the guanine nucleotide exchange factor (GEF) Tuba, and RhoGDI, that enables differential diffusion of inactive and active Cdc42 and is required to establish localized Cdc42 signaling during enterocyte polarization.

scholarly journals Myoblast Migration and Directional Persistence Affected by Syndecan-4-Mediated Tiam-1 Expression and Distribution

2020 ◽  
Vol 21 (3) ◽  
pp. 823 ◽  
Author(s):  
Daniel Becsky ◽  
Szuzina Gyulai-Nagy ◽  
Arpad Balind ◽  
Peter Horvath ◽  
Laszlo Dux ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Underlying Mechanism ◽  
Small Gtpase ◽  
Asymmetric Distribution ◽  
Nucleotide Exchange ◽  
Muscle Stem Cells ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
T Lymphoma ◽  
Migration Capacity

Skeletal muscle is constantly renewed in response to injury, exercise, or muscle diseases. Muscle stem cells, also known as satellite cells, are stimulated by local damage to proliferate extensively and form myoblasts that then migrate, differentiate, and fuse to form muscle fibers. The transmembrane heparan sulfate proteoglycan syndecan-4 plays multiple roles in signal transduction processes, such as regulating the activity of the small GTPase Rac1 (Ras-related C3 botulinum toxin substrate 1) by binding and inhibiting the activity of Tiam1 (T-lymphoma invasion and metastasis-1), a guanine nucleotide exchange factor for Rac1. The Rac1-mediated actin remodeling is required for cell migration. Syndecan-4 knockout mice cannot regenerate injured muscle; however, the detailed underlying mechanism is unknown. Here, we demonstrate that shRNA-mediated knockdown of syndecan-4 decreases the random migration of mouse myoblasts during live-cell microscopy. Treatment with the Rac1 inhibitor NSC23766 did not restore the migration capacity of syndecan-4 silenced cells; in fact, it was further reduced. Syndecan-4 knockdown decreased the directional persistence of migration, abrogated the polarized, asymmetric distribution of Tiam1, and reduced the total Tiam1 level of the cells. Syndecan-4 affects myoblast migration via its role in expression and localization of Tiam1; this finding may facilitate greater understanding of the essential role of syndecan-4 in the development and regeneration of skeletal muscle.

scholarly journals Identification of a Rab GTPase-activating protein cascade that controls recycling of the Rab5 GTPase Vps21 from the vacuole

2015 ◽  
Vol 26 (13) ◽  
pp. 2535-2549 ◽  
Author(s):  
Meenakshi Rana ◽  
Jens Lachmann ◽  
Christian Ungermann
Keyword(s):  
Endoplasmic Reticulum ◽  
Endocytic Pathway ◽  
Rab Gtpase ◽  
Guanine Nucleotide ◽  
Membrane Recycling ◽  
Nucleotide Exchange ◽  
Gtpase Activating Protein ◽  
Guanine Nucleotide Exchange ◽  
Vacuole Fusion ◽  
Nucleotide Exchange Factor

Transport within the endocytic pathway depends on a consecutive function of the endosomal Rab5 and the late endosomal/lysosomal Rab7 GTPases to promote membrane recycling and fusion in the context of endosomal maturation. We previously identified the hexameric BLOC-1 complex as an effector of the yeast Rab5 Vps21, which also recruits the GTPase-activating protein (GAP) Msb3. This raises the question of when Vps21 is inactivated on endosomes. We provide evidence for a Rab cascade in which activation of the Rab7 homologue Ypt7 triggers inactivation of Vps21. We find that the guanine nucleotide exchange factor (GEF) of Ypt7 (the Mon1-Ccz1 complex) and BLOC-1 both localize to the same endosomes. Overexpression of Mon1-Ccz1, which generates additional Ypt7-GTP, or overexpression of activated Ypt7 promotes relocalization of Vps21 from endosomes to the endoplasmic reticulum (ER), which is indicative of Vps21 inactivation. This ER relocalization is prevented by loss of either BLOC-1 or Msb3, but it also occurs in mutants lacking endosome–vacuole fusion machinery such as the HOPS tethering complex, an effector of Ypt7. Importantly, BLOC-1 interacts with the HOPS on vacuoles, suggesting a direct Ypt7-dependent cross-talk. These data indicate that efficient Vps21 recycling requires both Ypt7 and endosome–vacuole fusion, thus suggesting extended control of a GAP cascade beyond Rab interactions.

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