Carles Serra-Pagès

The LAR transmembrane protein tyrosine phosphatase and a coiled-coil LAR-interacting protein co-localize at focal adhesions.

Focal adhesions are sites of cell‐extracellular matrix interactions that function in anchoring stress fibers to the plasma membrane and in adhesion‐mediated signal transduction. Both focal adhesion structure and signaling ability involve protein tyrosine phosphorylation. LAR is a broadly expressed transmembrane protein tyrosine phosphatase comprised of a cell adhesion‐like ectodomain and two intracellular protein tyrosine phosphatase domains. We have identified a novel cytoplasmic 160 kDa phosphoserine protein termed LAR‐interacting protein 1 (LIP.1), which binds to the LAR membrane‐distal D2 protein tyrosine phosphatase domain and appears to localize LAR to focal adhesions. Both LAR and LIP.1 decorate the ends of focal adhesions most proximal to the cell nucleus and are excluded from the distal ends of focal adhesions, thus localizing to regions of focal adhesions presumably undergoing disassembly. We propose that LAR and LIP.1 may regulate the disassembly of focal adhesions and thus help orchestrate cell‐matrix interactions.

Interaction between GRIP and Liprin-α/SYD2 Is Required for AMPA Receptor Targeting

Abstract Interaction with the multi-PDZ protein GRIP is required for the synaptic targeting of AMPA receptors, but the underlying mechanism is unknown. We show that GRIP binds to the liprin-α/SYD2 family of proteins that interact with LAR receptor protein tyrosine phosphatases (LAR-RPTPs) and that are implicated in presynaptic development. In neurons, liprin-α and LAR-RPTP are enriched at synapses and coimmunoprecipitate with GRIP and AMPA receptors. Dominant-negative constructs that interfere with the GRIP-liprin interaction disrupt the surface expression and dendritic clustering of AMPA receptors in cultured neurons. Thus, by mediating the targeting of liprin/GRIP-associated proteins, liprin-α is important for postsynaptic as well as presynaptic maturation.

The Trio Guanine Nucleotide Exchange Factor Is a RhoA Target BINDING OF RhoA TO THE TRIO IMMUNOGLOBULIN-LIKE DOMAIN

Trio is a complex protein containing two guanine nucleotide exchange factor domains each with associated pleckstrin homology domains, a serine/threonine kinase domain, two SH3 domains, an immunoglobulin-like domain, and spectrin-like repeats. Trio was originally identified as a LAR tyrosine phosphatase-binding protein and is involved in actin remodeling, cell migration, and cell growth. Herein we provide evidence that Trio not only activates RhoA but is also a RhoA target. The RhoA-binding site was mapped to the Trio immunoglobulin-like domain. RhoA isoprenylation is necessary for the RhoA-Trio interaction, because mutation of the RhoA carboxyl-terminal cysteine residue blocked binding. The existence of an intramolecular functional link between RhoA activation and RhoA binding is suggested by the finding that Trio exchange activity enhanced RhoA binding to Trio. Furthermore, immunofluorescence studies of HeLa cells showed that although ectopically expressed Trio was evenly distributed within the cell, co-expression of Trio with RhoA resulted in relocalization of Trio into punctate structures. Relocalization was not observed with Trio constructs lacking the immunoglobulin-like domain, indicating that RhoA acts to regulate Trio localization via binding to the immunoglobulin-like domain. We propose that Trio-mediated RhoA activation and subsequent RhoA-mediated relocalization of Trio functions to modulate and coordinate Trio signaling.

Association of the Cell Cycle Regulatory Proteins p45SKP2 and CksHs1 FUNCTIONAL EFFECT ON CDK2 COMPLEX FORMATION AND KINASE ACTIVITY

In mammalian cells, CDK2 is part of a multiprotein complex that includes Cyclin A or E and cell cycle regulatory proteins such as p21Cip1, PCNA, p27Kip1, p45SKP2, p19SKP1, and CksHs1/CksHs2. While the role of some of these proteins has been well studied, the function of other proteins in the complex remains unclear. In this study, we showed that the carboxyl-terminal region of p45SKP2 associates directly with CksHs1 and that CksHs1 negatively regulated the interaction between p45SKP2 and CDK2. Moreover, we showed that overexpression of CksHs1 inhibits CDK2 kinase activity and that additional expression of p45SKP2overcame this inhibition and restored CDK2 kinase activity. We proposed that the association of CksHs1 and p45SKP2 prevented CksHs1 from binding CDK2 and negatively regulating the CDK2 kinase activity.