scholarly journals The Rac activator Tiam1 controls tight junction biogenesis in keratinocytes through binding to and activation of the Par polarity complex

2005 ◽  
Vol 170 (7) ◽  
pp. 1029-1037 ◽  
Author(s):  
Alexander E.E. Mertens ◽  
Tomasz P. Rygiel ◽  
Cristina Olivo ◽  
Rob van der Kammen ◽  
John G. Collard
Keyword(s):  
Cell Polarity ◽  
Epidermal Keratinocytes ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Invasion And Metastasis ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
T Lymphoma ◽  
Maturation Defect ◽  
Membrane Sealing

The GTPases Rac and Cdc42 play a pivotal role in the establishment of cell polarity by stimulating biogenesis of tight junctions (TJs). In this study, we show that the Rac-specific guanine nucleotide exchange factor Tiam1 (T-lymphoma invasion and metastasis) controls the cell polarity of epidermal keratinocytes. Similar to wild-type (WT) keratinocytes, Tiam1-deficient cells establish primordial E-cadherin–based adhesions, but subsequent junction maturation and membrane sealing are severely impaired. Tiam1 and V12Rac1 can rescue the TJ maturation defect in Tiam1-deficient cells, indicating that this defect is the result of impaired Tiam1–Rac signaling. Tiam1 interacts with Par3 and aPKCζ, which are two components of the conserved Par3–Par6–aPKC polarity complex, and triggers biogenesis of the TJ through the activation of Rac and aPKCζ, which is independent of Cdc42. Rac is activated upon the formation of primordial adhesions (PAs) in WT but not in Tiam1-deficient cells. Our data indicate that Tiam1-mediated activation of Rac in PAs controls TJ biogenesis and polarity in epithelial cells by association with and activation of the Par3–Par6–aPKC polarity complex.

scholarly journals The Rac-GAP Bcr is a novel regulator of the Par complex that controls cell polarity

2013 ◽  
Vol 24 (24) ◽  
pp. 3857-3868 ◽  
Author(s):  
Anjana S. Narayanan ◽  
Steve B. Reyes ◽  
Kyongmi Um ◽  
Joseph H. McCarty ◽  
Kimberley F. Tolias
Keyword(s):  
Cell Migration ◽  
Cell Polarity ◽  
Leading Edge ◽  
Cell Polarization ◽  
Nucleotide Exchange ◽  
Complex Activity ◽  
Guanine Nucleotide Exchange ◽  
Directed Cell Migration ◽  
Nucleotide Exchange Factor ◽  
T Lymphoma

Cell polarization is essential for many biological processes, including directed cell migration, and loss of polarity contributes to pathological conditions such as cancer. The Par complex (Par3, Par6, and PKCζ) controls cell polarity in part by recruiting the Rac-specific guanine nucleotide exchange factor T-lymphoma invasion and metastasis 1 (Tiam1) to specialized cellular sites, where Tiam1 promotes local Rac1 activation and cytoskeletal remodeling. However, the mechanisms that restrict Par-Tiam1 complex activity to the leading edge to maintain cell polarity during migration remain unclear. We identify the Rac-specific GTPase-activating protein (GAP) breakpoint cluster region protein (Bcr) as a novel regulator of the Par-Tiam1 complex. We show that Bcr interacts with members of the Par complex and inhibits both Rac1 and PKCζ signaling. Loss of Bcr results in faster, more random migration and striking polarity defects in astrocytes. These polarity defects are rescued by reducing PKCζ activity or by expressing full-length Bcr, but not an N-terminal deletion mutant or the homologous Rac-GAP, Abr, both of which fail to associate with the Par complex. These results demonstrate that Bcr is an integral member of the Par-Tiam1 complex that controls polarized cell migration by locally restricting both Rac1 and PKCζ function.

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 Linking Ras to myosin function: RasGEF Q, a Dictyostelium exchange factor for RasB, affects myosin II functions

2008 ◽  
Vol 181 (5) ◽  
pp. 747-760 ◽  
Author(s):  
Subhanjan Mondal ◽  
Deenadayalan Bakthavatsalam ◽  
Paul Steimle ◽  
Berthold Gassen ◽  
Francisco Rivero ◽  
...  
Keyword(s):  
Cell Polarity ◽  
Cell Sorting ◽  
Myosin Ii ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Myosin Heavy Chain Kinase ◽  
Null Mutants

Ras guanine nucleotide exchange factor (GEF) Q, a nucleotide exchange factor from Dictyostelium discoideum, is a 143-kD protein containing RasGEF domains and a DEP domain. We show that RasGEF Q can bind to F-actin, has the potential to form complexes with myosin heavy chain kinase (MHCK) A that contain active RasB, and is the predominant exchange factor for RasB. Overexpression of the RasGEF Q GEF domain activates RasB, causes enhanced recruitment of MHCK A to the cortex, and leads to cytokinesis defects in suspension, phenocopying cells expressing constitutively active RasB, and myosin-null mutants. RasGEF Q− mutants have defects in cell sorting and slug migration during later stages of development, in addition to cell polarity defects. Furthermore, RasGEF Q− mutants have increased levels of unphosphorylated myosin II, resulting in myosin II overassembly. Collectively, our results suggest that starvation signals through RasGEF Q to activate RasB, which then regulates processes requiring myosin II.

scholarly journals The RAC1 activator Tiam1 regulates centriole duplication through controlling PLK4 levels

2020 ◽  
Author(s):  
Andrew P. Porter ◽  
Gavin R. M. White ◽  
Erinn-Lee Ogg ◽  
Helen J. Whalley ◽  
Angeliki Malliri
Keyword(s):  
Cell Cycle ◽  
Guanine Nucleotide Exchange Factor ◽  
S Phase ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Centriole Duplication ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
F Box Protein

SummaryCentriole duplication is tightly controlled to maintain correct centriole number through the cell cycle. A key component of this control is the regulated degradation of PLK4, the master regulator of centriole duplication. Here we show that the Rac1 guanine nucleotide exchange factor (GEF) Tiam1 localises to centrosomes during S-phase, where it is required for maintenance of normal centriole number. Depletion of Tiam1 leads to an increase in centrosomal PLK4, centriole overduplication and ultimately to lagging chromosomes at anaphase and aneuploidy. The effects of Tiam1 depletion can be rescued by re-expression of wild-type Tiam1 and catalytically inactive (GEF*) Tiam1, but not by Tiam1 mutants unable to bind to the F-box protein βTRCP, implying that Tiam1 regulates PLK4 levels through promoting βTRCP-mediated degradation.

scholarly journals Dynamic association of the PI3P-interacting Mon1-Ccz1 GEF with vacuoles is controlled through its phosphorylation by the type 1 casein kinase Yck3

2014 ◽  
Vol 25 (10) ◽  
pp. 1608-1619 ◽  
Author(s):  
Gus Lawrence ◽  
Christopher C. Brown ◽  
Blake A. Flood ◽  
Surya Karunakaran ◽  
Margarita Cabrera ◽  
...  
Keyword(s):  
Fusion Reaction ◽  
Casein Kinase ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Phosphatidylinositol 3 Kinase ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Phosphatidic Acid Phosphatase ◽  
Phosphatidylinositol 3

Maturation of organelles in the endolysosomal pathway requires exchange of the early endosomal GTPase Rab5/Vps21 for the late endosomal Rab7/Ypt7. The Rab exchange depends on the guanine nucleotide exchange factor activity of the Mon1-Ccz1 heterodimer for Ypt7. Here we investigate vacuole binding and recycling of Mon1-Ccz1. We find that Mon1-Ccz1 is absent on vacuoles lacking the phosphatidic acid phosphatase Pah1, which also lack Ypt7, the phosphatidylinositol 3-kinase Vps34, and the lipid phosphatidylinositol 3-phosphate (PI3P). Interaction of Mon1-Ccz1 with wild-type vacuoles requires PI3P, as shown in competition experiments. We also find that Mon1 is released from vacuoles during the fusion reaction and its release requires its phosphorylation by the type 1 casein kinase Yck3. In contrast, Mon1 is retained on vacuoles lacking Yck3 or when Mon1 phosphorylation sites are mutated. Phosphorylation and release of Mon1 is restored with addition of recombinant Yck3. Together the results show that Mon1 is recruited to endosomes and vacuoles by PI3P and, likely after activating Ypt7, is phosphorylated and released from vacuoles for recycling.

scholarly journals The Rac activator Tiam1 is required for α3β1-mediated laminin-5 deposition, cell spreading, and cell migration

2005 ◽  
Vol 171 (5) ◽  
pp. 871-881 ◽  
Author(s):  
Irene H.L. Hamelers ◽  
Cristina Olivo ◽  
Alexander E.E. Mertens ◽  
D. Michiel Pegtel ◽  
Rob A. van der Kammen ◽  
...  
Keyword(s):  
Mouse Skin ◽  
Nucleotide Exchange ◽  
Laminin 5 ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
T Lymphoma ◽  
Guanosine Triphosphatase ◽  
And Migration ◽  
Migration Of Cells

The Rho-like guanosine triphosphatase Rac1 regulates various signaling pathways, including integrin-mediated adhesion and migration of cells. However, the mechanisms by which integrins signal toward Rac are poorly understood. We show that the Rac-specific guanine nucleotide exchange factor Tiam1 (T-lymphoma invasion and metastasis 1) is required for the integrin-mediated laminin (LN)-5 deposition, spreading, and migration of keratinocytes. In contrast to wild-type keratinocytes, Tiam1-deficient (Tiam1−/−) keratinocytes are unable to adhere to and spread on a glass substrate because they are unable to deposit their own LN5 substrate. Both Tiam1 and V12Rac1 can rescue the defects of Tiam1−/− keratinocytes, indicating that these deficiencies are caused by impaired Tiam1-mediated Rac activation. Tiam1−/− cells are unable to activate Rac upon α3β1-mediated adhesion to an exogenous LN5 substrate. Moreover, Tiam1 deficiency impairs keratinocyte migration in vitro and reepithelialization of excision wounds in mouse skin. Our studies indicate that Tiam1 is a key molecule in α3β1-mediated activation of Rac, which is essential for proper production and secretion of LN5, a requirement for the spreading and migration of keratinocytes.

scholarly journals Interaction between Tiam1 and the Arp2/3 complex links activation of Rac to actin polymerization

2006 ◽  
Vol 397 (1) ◽  
pp. 39-45 ◽  
Author(s):  
Jean Paul ten Klooster ◽  
Eva E. Evers ◽  
Lennert Janssen ◽  
Laura M. Machesky ◽  
Frits Michiels ◽  
...  
Keyword(s):  
Actin Polymerization ◽  
Morphological Changes ◽  
Coiled Coil ◽  
Pleckstrin Homology Domain ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor ◽  
Coiled Coil Region ◽  
T Lymphoma ◽  
Cell Cell

The Rac-specific GEF (guanine-nucleotide exchange factor) Tiam1 (T-lymphoma invasion and metastasis 1) regulates migration, cell–matrix and cell–cell adhesion by modulating the actin cytoskeleton through the GTPase, Rac1. Using yeast two-hybrid screening and biochemical assays, we found that Tiam1 interacts with the p21-Arc [Arp (actin-related protein) complex] subunit of the Arp2/3 complex. Association occurred through the N-terminal pleckstrin homology domain and the adjacent coiled-coil region of Tiam1. As a result, Tiam1 co-localizes with the Arp2/3 complex at sites of actin polymerization, such as epithelial cell–cell contacts and membrane ruffles. Deletion of the p21-Arc-binding domain in Tiam1 impairs its subcellular localization and capacity to activate Rac1, suggesting that binding to the Arp2/3 complex is important for the function of Tiam1. Indeed, blocking Arp2/3 activation with a WASP (Wiskott–Aldrich syndrome protein) inhibitor leads to subcellular relocalization of Tiam1 and decreased Rac activation. Conversely, functionally active Tiam1, but not a GEF-deficient mutant, promotes activation of the Arp2/3 complex and its association with cytoskeletal components, indicating that Tiam1 and Arp2/3 are mutually dependent for their correct localization and signalling. Our data suggests a model in which the Arp2/3 complex acts as a scaffold to localize Tiam1, and thereby Rac activity, which are both required for activation of the Arp2/3 complex and further Arp2/3 recruitment. This ‘self-amplifying’ signalling module involving Tiam1, Rac and the Arp2/3 complex could thus drive actin polymerization at specific sites in cells that are required for dynamic morphological changes.

scholarly journals Depletion of Ric-8B leads to reduced mTORC2 activity

2019 ◽  
Author(s):  
Maíra H. Nagai ◽  
Luciana M. Gutiyama ◽  
Victor P. S. Xavier ◽  
Cleiton F. Machado ◽  
Alice H. Reis ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Neural Tube ◽  
Adult Brain ◽  
Nucleotide Exchange ◽  
Wild Type ◽  
Hypomorphic Mutation ◽  
Downstream Target ◽  
Cellular Processes ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

AbstractmTOR, a serine/threonine protein kinase that is involved in a series of critical cellular processes, can be found in two functionally distinct complexes, mTORC1 and mTORC2. In contrast to mTORC1, little is known about the mechanisms that regulate mTORC2. Here we show that mTORC2 activity is reduced in mice with a hypomorphic mutation of the Ric-8B gene. Ric-8B is a highly conserved protein that acts as a non-canonical guanine nucleotide exchange factor (GEF) for heterotrimeric Gαs/olf type subunits. We found that Ric-8B hypomorph embryos are smaller than their wild type littermates, fail to close the neural tube in the cephalic region and die during mid-embryogenesis. Comparative transcriptome analysis revealed that signaling pathways involving GPCRs and G proteins are dysregulated in the Ric-8B mutant embryos. Interestingly, this analysis also revealed an unexpected impairment of the mTOR signaling pathway.Phosphorylation of Akt at Ser 473 is downregulated in the Ric-8B mutant embryos, indicating a decreased activity of mTORC2. In contrast, phosphorylation of S6, a downstream target of mTORC1, is unaltered. Knockdown of the endogenous Ric-8B gene in HEK293T cells leads to reduced phosphorylation levels of Akt at Ser 473, but not of S6, further supporting the selective involvement of Ric-8B in mTORC2 activity. Our results reveal a crucial role for Ric-8B in development and provide novel insights into the signals that regulate mTORC2 activity.Author SummaryGene inactivation in mice can be used to identify genes that are involved in important biological processes and that may contribute to disease. By using this approach, we found that the Ric-8B gene is essential for embryogenesis and for the normal development of the nervous system. Ric-8B mutant mouse embryos are smaller than their wild type littermates and show neural tube defects at the cranial region. This approach also allowed us to identify the biological pathways that are involved in the observed phenotypes, the G protein and mTORC2 signaling pathways. mTORC2 plays particular important roles also in the adult brain, and has been implicated in neurological disorders. Ric-8B is highly conserved in mammals, including humans. Our mutant mice provide a model to study the complex molecular and cellular processes underlying the interplay between Ric-8B and mTORC2 in neuronal function.

scholarly journals Characterization of Synthetic-Lethal Mutants Reveals a Role for the Saccharomyces cerevisiae Guanine-Nucleotide Exchange Factor Cdc24p in Vacuole Function and Na+ Tolerance

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 43-55 ◽  
Author(s):  
William H White ◽  
Douglas I Johnson
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Polarity ◽  
Synthetic Lethality ◽  
Guanine Nucleotide Exchange Factor ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Synthetic Lethal ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Abstract Cdc24p is the guanine-nucleotide exchange factor for the Cdc42p GTPase, which controls cell polarity in Saccharomyces cerevisiae. To identify new genes that may affect cell polarity, we characterized six UV-induced csl (CDC24  synthetic-lethal) mutants that exhibited synthetic-lethality with cdc24-4ts at 23°. Five mutants were not complemented by plasmid-borne CDC42, RSRI, BUD5, BEM1, BEM2, BEM3 or CLA4 genes, which are known to play a role in cell polarity. The csl3 mutant displayed phenotypes similar to those observed with calcium-sensitive, Pet−  vma mutants defective in vacuole function. CSL5 was allelic to VMA5, the vacuolar H+-ATPase subunit C, and one third of cs15 cdc24-4ts cells were elongated or had misshapen buds. A cdc24-4ts Δvma5::LEU2 double mutant did not exhibit synthetic lethality, suggesting that the csl5/vma5 cdc24-4ts synthetic-lethality was not simply due to altered vacuole function. The cdc24-4ts mutant, like Δvma5::LEU2 and cs13 mutants, was sensitive to high levels of Ca2+ as well as Na+ in the growth media, which did not appear to be a result of a fragile cell wall because the phenotypes were not remedied by 1 m sorbitol. Our results indicated that Cdc24p was required in one V-ATPase mutant and another mutant affecting vacuole morphology, and also implicated Cdc24p in Na+ tolerance.

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