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Dive into the research topics where Chinten James Lim is active.

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Featured researches published by Chinten James Lim.


Journal of Biological Chemistry | 2009

RIAM Activates Integrins by Linking Talin to Ras GTPase Membrane-targeting Sequences

Ho-Sup Lee; Chinten James Lim; Wilma Puzon-McLaughlin; Sanford J. Shattil; Mark H. Ginsberg

Rap1 small GTPases interact with Rap1-GTP-interacting adaptor molecule (RIAM), a member of the MRL (Mig-10/RIAM/Lamellipodin) protein family, to promote talin-dependent integrin activation. Here, we show that MRL proteins function as scaffolds that connect the membrane targeting sequences in Ras GTPases to talin, thereby recruiting talin to the plasma membrane and activating integrins. The MRL proteins bound directly to talin via short, N-terminal sequences predicted to form amphipathic helices. RIAM-induced integrin activation required both its capacity to bind to Rap1 and to talin. Moreover, we constructed a minimized 50-residue Rap-RIAM module containing the talin binding site of RIAM joined to the membrane-targeting sequence of Rap1A. This minimized Rap-RIAM module was sufficient to target talin to the plasma membrane and to mediate integrin activation, even in the absence of Rap1 activity. We identified a short talin binding sequence in Lamellipodin (Lpd), another MRL protein; talin binding Lpd sequence joined to a Rap1 membrane-targeting sequence is sufficient to recruit talin and activate integrins. These data establish the mechanism whereby MRL proteins interact with both talin and Ras GTPases to activate integrins.


Nature Cell Biology | 2009

Slit2-Robo4 signalling promotes vascular stability by blocking Arf6 activity.

Christopher A. Jones; Naoyuki Nishiya; Nyall R. London; Weiquan Zhu; Lise K. Sorensen; Aubrey C. Chan; Chinten James Lim; Haoyu Chen; Qisheng Zhang; Peter G. Schultz; Alaa M. Hayallah; Kirk R. Thomas; Michael Famulok; Kang Zhang; Mark H. Ginsberg; Dean Y. Li

Slit–Roundabout (Robo) signalling has a well-understood role in axon guidance. Unlike in the nervous system, however, Slit-dependent activation of an endothelial-specific Robo, Robo4, does not initiate a guidance program. Instead, Robo4 maintains the barrier function of the mature vascular network by inhibiting neovascular tuft formation and endothelial hyperpermeability induced by pro-angiogenic factors. In this study, we used cell biological and biochemical techniques to elucidate the molecular mechanism underlying the maintenance of vascular stability by Robo4. Here, we demonstrate that Robo4 mediates Slit2-dependent suppression of cellular protrusive activity through direct interaction with the intracellular adaptor protein paxillin and its paralogue, Hic-5. Formation of a Robo4–paxillin complex at the cell surface blocks activation of the small GTPase Arf6 and, consequently, Rac by recruitment of Arf-GAPs (ADP-ribosylation factor- directed GTPase-activating proteins) such as GIT1. Consistent with these in vitro studies, inhibition of Arf6 activity in vivo phenocopies Robo4 activation by reducing pathologic angiogenesis in choroidal and retinal vascular disease and VEGF-165 (vascular endothelial growth factor-165)-induced retinal hyperpermeability. These data reveal that a Slit2–Robo4–paxillin–GIT1 network inhibits the cellular protrusive activity underlying neovascularization and vascular leak, and identify a new therapeutic target for ameliorating diseases involving the vascular system.


Journal of Biological Chemistry | 2009

β Integrin Tyrosine Phosphorylation Is a Conserved Mechanism for Regulating Talin-induced Integrin Activation

Nicholas J. Anthis; Jacob R. Haling; Camilla L. Oxley; Massimiliano Memo; Kate L. Wegener; Chinten James Lim; Mark H. Ginsberg; Iain D. Campbell

Integrins are large membrane-spanning receptors fundamental to cell adhesion and migration. Integrin adhesiveness for the extracellular matrix is activated by the cytoskeletal protein talin via direct binding of its phosphotyrosine-binding-like F3 domain to the cytoplasmic tail of the β integrin subunit. The phosphotyrosine-binding domain of the signaling protein Dok1, on the other hand, has an inactivating effect on integrins, a phenomenon that is modulated by integrin tyrosine phosphorylation. Using full-length tyrosine-phosphorylated 15N-labeled β3, β1A, and β7 integrin tails and an NMR-based protein-protein interaction assay, we show that talin1 binds to the NPXY motif and the membrane-proximal portion of β3, β1A, and β7 tails, and that the affinity of this interaction is decreased by integrin tyrosine phosphorylation. Dok1 only interacts weakly with unphosphorylated tails, but its affinity is greatly increased by integrin tyrosine phosphorylation. The Dok1 interaction remains restricted to the integrin NPXY region, thus phosphorylation inhibits integrin activation by increasing the affinity of β integrin tails for a talin competitor that does not form activating membrane-proximal interactions with the integrin. Key residues governing these specificities were identified by detailed structural analysis, and talin1 was engineered to bind preferentially to phosphorylated integrins by introducing the mutation D372R. As predicted, this mutation affects talin1 localization in live cells in an integrin phosphorylation-specific manner. Together, these results indicate that tyrosine phosphorylation is a common mechanism for regulating integrin activation, despite subtle differences in how these integrins interact with their binding proteins.


EMBO Reports | 2004

Chemoattractant-induced Ras activation during Dictyostelium aggregation.

Helmut Kae; Chinten James Lim; George B. Spiegelman; Gerald Weeks

Ras proteins are highly conserved molecular switches that regulate cellular response to external stimuli. Dictyostelium discoideum contains an extensive family of Ras proteins that function in regulation of mitosis, cytoskeletal function and motility, and the onset of development. Little is known about the events that lead to the activation of Ras proteins in Dictyostelium, primarily owing to a lack of a biochemical assay to measure the levels of activated Ras. We have adapted an assay, used successfully to measure activated Ras in mammalian cells, to monitor activation of two Dictyostelium Ras proteins, RasC and RasG. We have found that the Ras‐binding domain (RBD) of mammalian Raf1 was capable of binding to the activated form of RasG, but not to the activated form of RasC; however, the RBD of Schizosaccharomyces pombe Byr2 was capable of binding preferentially to the activated forms of both RasC and RasG. Using this assay, we discovered that RasC and RasG showed a rapid and transient activation when aggregation‐competent cells were stimulated with the chemoattractant cAMP, and this activation did not occur in a number of cAMP signalling mutants. These data provide further evidence of a role for both RasC and RasG in the early development of Dictyostelium.


The EMBO Journal | 2001

RasC is required for optimal activation of adenylyl cyclase and Akt/PKB during aggregation

Chinten James Lim; George B. Spiegelman; Gerald Weeks

Disruption of Dictyostelium rasC, encoding a Ras subfamily protein, generated cells incapable of aggregation. While rasC expression is enriched in a cell type‐specific manner during post‐aggregative development, the defect in rasC− cells is restricted to aggregation and fully corrected by application of exogenous cAMP pulses. cAMP is not produced in rasC− cells stimulated by 2′‐deoxy‐cAMP, but is produced in response to GTPγS in cell lysates, indicating that G‐protein‐coupled cAMP receptor activation of adenylyl cyclase is regulated by RasC. However, cAMP‐induced ERK2 phosphorylation is unaffected in rasC− cells, indicating that RasC is not an upstream activator of the mitogen‐activated protein kinase required for cAMP relay. rasC− cells also exhibit reduced chemotaxis to cAMP during early development and delayed response to periodic cAMP stimuli produced by wild‐type cells in chimeric mixtures. Furthermore, cAMP‐induced Akt/PKB phosphorylation through a phosphatidylinositide 3‐kinase (PI3K)‐dependent pathway is dramatically reduced in rasC− cells, suggesting that G‐protein‐coupled serpentine receptor activation of PI3K is regulated by RasC. Cells lacking the RasGEF, AleA, exhibit similar defects as rasC− cells, suggesting that AleA may activate RasC.


Nature Cell Biology | 2007

α4 Integrins are Type I cAMP-dependent protein kinase-anchoring proteins

Chinten James Lim; Jaewon Han; Nima Yousefi; Yuliang Ma; Paul S. Amieux; G. Stanley McKnight; Susan S. Taylor; Mark H. Ginsberg

A-kinase anchoring proteins (AKAPs) control the localization and substrate specificity of cAMP-dependent protein kinase (PKA), tetramers of regulatory (PKA-R) and catalytic (PKA-C) subunits, by binding to PKA-R subunits. Most mammalian AKAPs bind Type II PKA through PKA-RII (ref. 2), whereas dual specificity AKAPs bind both PKA-RI and PKA-RII (ref. 3). Inhibition of PKA–AKAP interactions modulates PKA signalling. Localized PKA activation in pseudopodia of migrating cells phosphorylates α4 integrins to provide spatial cues governing cell motility. Here, we report that the α4 cytoplasmic domain is a Type I PKA-specific AKAP that is distinct from canonical AKAPs in two ways: the α4 interaction requires the PKA holoenzyme, and is insensitive to amphipathic peptides that disrupt most PKA–AKAP interactions. We exploited type-specific PKA anchoring peptides to create genetically encoded baits that sequester specific PKA isoforms to the mitochondria and found that mislocalization of Type I, but not Type II, PKA disrupts α4 phosphorylation and markedly inhibits the velocity and directional persistence of cell migration.


Molecular Biology of the Cell | 2013

Two Modes of Integrin Activation Form a Binary Molecular Switch in Adhesion Maturation

Ho-Sup Lee; Praju Anekal; Chinten James Lim; Chi-Chao Liu; Mark H. Ginsberg

Talin-mediated integrin activation drives integrin-based adhesions. A simple binary switch—vinculin competitively displacing RIAM from talin—is found to play a central role in the maturation and evolving functions of integrin-based adhesions.


Eukaryotic Cell | 2004

RasC plays a role in transduction of temporal gradient information in the cyclic-AMP wave of Dictyostelium discoideum.

Deborah Wessels; Rebecca Brincks; Spencer Kuhl; Vesna Stepanovic; Karla J. Daniels; Gerald Weeks; Chinten James Lim; George B. Spiegelman; Danny Fuller; Negin Iranfar; William F. Loomis; David R. Soll

ABSTRACT To define the role that RasC plays in motility and chemotaxis, the behavior of a rasC null mutant, rasC−, in buffer and in response to the individual spatial, temporal, and concentration components of a natural cyclic AMP (cAMP) wave was analyzed by using computer-assisted two-dimensional and three-dimensional motion analysis systems. These quantitative studies revealed that rasC− cells translocate at the same velocity and exhibit chemotaxis up spatial gradients of cAMP with the same efficiency as control cells. However, rasC− cells exhibit defects in maintaining anterior-posterior polarity along the substratum and a single anterior pseudopod when translocating in buffer in the absence of an attractant. rasC− cells also exhibit defects in their responses to both the increasing and decreasing temporal gradients of cAMP in the front and the back of a wave. These defects result in the inability of rasC− cells to exhibit chemotaxis in a natural wave of cAMP. The inability to respond normally to temporal gradients of cAMP results in defects in the organization of the cytoskeleton, most notably in the failure of both F actin and myosin II to exit the cortex in response to the decreasing temporal gradient of cAMP in the back of the wave. While the behavioral defect in the front of the wave is similar to that of the myoA−/myoF− myosin I double mutant, the behavioral and cytoskeletal defects in the back of the wave are similar to those of the S13A myosin II regulatory light-chain phosphorylation mutant. Expression array data support the premise that the behavioral defects exhibited by the rasC− mutant are the immediate result of the absence of RasC function.


Molecular and Cellular Biology | 2012

Protein Tyrosine Phosphatase α Phosphotyrosyl-789 Binds BCAR3 To Position Cas for Activation at Integrin-Mediated Focal Adhesions

Guobin Sun; Suzanne Y. S. Cheng; Min Chen; Chinten James Lim; Catherine J. Pallen

ABSTRACT Integrin-mediated focal adhesions connect the extracellular matrix and cytoskeleton to regulate cell responses, such as migration. Protein tyrosine phosphatase α (PTPα) regulates integrin signaling, focal adhesion formation, and migration, but its roles in these events are incompletely understood. The integrin-proximal action of PTPα activates Src family kinases, and subsequent phosphorylation of PTPα at Tyr789 acts in an unknown manner to promote migration. PTPα-null cells were used in reconstitution assays to distinguish PTPα-Tyr789-dependent signaling events. This showed that PTPα-Tyr789 regulates the localization of PTPα and the scaffolding protein Cas to adhesion sites where Cas interacts with and is phosphorylated by Src to initiate Cas signaling. Linking these events, we identify BCAR3 as a molecular connector of PTPα and Cas, with phospho-Tyr789 PTPα serving as the first defined cellular ligand for the BCAR3 SH2 domain that recruits BCAR3-Cas to adhesions. Our findings reveal a novel role of PTPα in integrin-induced adhesion assembly that enables Src-mediated activation of the pivotal function of Cas in migration.


Journal of Muscle Research and Cell Motility | 2002

Cytoskeletal regulation by Dictyostelium Ras subfamily proteins

Chinten James Lim; George B. Spiegelman; Gerald Weeks

The Ras subfamily proteins are monomeric GTPases that function as molecular switches in cellular signal transduction. The roles of six of these proteins in regulating actin cytoskeletal functions in Dictyostelium discoideum are discussed in this review.

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Chi-Chao Liu

University of British Columbia

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George B. Spiegelman

University of British Columbia

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Gerald Weeks

University of British Columbia

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Mu Chiao

University of British Columbia

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Pascal Leclair

University of British Columbia

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Gregor S. D. Reid

University of British Columbia

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Farzad Khademolhosseini

University of British Columbia

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