PKC-θ selectively controls the adhesion-stimulating molecule Rap1 (original) (raw)
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PKC- selectively controls the adhesion-stimulating molecule Rap1
Blood, 2008
selectively controls the adhesion-stimulating molecule Rap1 θ PKChttp://bloodjournal.hematologylibrary.org/content/112/12/4617.full.html Updated information and services can be found at: (5019 articles) Immunobiology Articles on similar topics can be found in the following Blood collections http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#repub\_requests Information about reproducing this article in parts or in its entirety may be found online at: http://bloodjournal.hematologylibrary.org/site/misc/rights.xhtml#reprints Information about ordering reprints may be found online at: http://bloodjournal.hematologylibrary.org/site/subscriptions/index.xhtml
Journal of Biological Chemistry, 2011
Although essential for T cell function, the identity of the T cell receptor (TCR) "inside-out" pathway for the activation of lymphocyte function-associated antigen 1 (LFA-1) is unclear. SKAP1 (SKAP-55) is the upstream regulator needed for TCRinduced RapL-Rap1 complex formation and LFA-1 activation. In this paper, we show that SKAP1 is needed for RapL binding to membranes in a manner dependent on the PH domain of SKAP1 and the PI3K pathway. A SKAP1 PH domain-inactivating mutation (i.e. R131M) markedly impaired RapL translocation to membranes for Rap1 and LFA-1 binding and the up-regulation of LFA-1-intercellular adhesion molecule 1 (ICAM-1) binding. Further, N-terminal myr-tagged SKAP1 for membrane binding facilitated constitutive RapL membrane and Rap1 binding and effectively substituted for PI3K and TCR ligation in the activation of LFA-1 in T cells. Integrins regulate T cell migration to sites of inflammation, movement in lymph nodes, and conjugate formation between T cells and antigen-presenting cells (1-3). They comprise as many as 20 ␣ heterodimers that are activated by changes in conformation and clustering (2, 3). T cell function is primarily regulated by the 2 family members, such as lymphocyte function-associated antigen (LFA-1, 2 ␣L (CD11a)/2 (CD18)) and ␣4 integrins such as ␣41 (Very Late Antigen-4 (VLA-4)). Binding to intercellular adhesion molecule 1 (ICAM-1) is mediated by the LFA-1 ␣ and  subunits that interact to create a headpiece in the  subunit, termed the I-like domain (4). LFA-1 binds to ICAM-1 and ICAM-2 on major histocompatibility complex bearing antigen-presenting cells (5). While in low-affinity conformation on resting T-cells, TCR and chemokine receptor ligation generates "inside-out" signals that activate integrins (3, 6, 7). Generic upstream regulators of
Blood, 2007
Regulated adhesion of T cells by the integrins LFA-1 (lymphocyte function-associated antigen-1) and VLA-4 (very late antigen-4) is essential for T-cell trafficking. The small GTPase Rap1 is a critical activator of both integrins in murine lymphocytes and T-cell lines. Here we examined the contribution of the Rap1 regulatory pathway in integrin activation in primary CD3+ human T cells. We demonstrate that inactivation of Rap1 GTPase in human T cells by expression of SPA1 or Rap1GAP blocked stromal cell-derived factor-1α (SDF-1α)–stimulated LFA-1–ICAM-1 (intercellular adhesion molecule-1) interactions and LFA-1 affinity modulation but unexpectedly did not significantly affect binding of VLA-4 to its ligand VCAM-1 (vascular cell adhesion molecule 1). Importantly, silencing of the Rap1 guanine exchange factor CalDAG-GEFI inhibited SDF-1α- and phorbol 12-myristate 13-acetate (PMA)–induced adhesion to ICAM-1 while having no effect on adhesion to VCAM-1. Pharmacologic inhibition of Phospho...
Immunology Letters, 2004
Dynamic regulation of integrin-mediated adhesion is central to lymphocyte trafficking and antigen recognition. The small GTPase Rap1 is a potent stimulator of leukocyte integrins through modulation of affinity and avidity. In addition, lymphocyte Rap1 has unique abilities to trigger cell polarization and enhance cell motility. These characteristics of Rap1 contribute to adhesive interactions with antigen-presenting cells (APC) and the vascular endothelium. In the process of elucidating the molecular mechanisms of Rap1-mediated integrin activation, we have identified a novel Rap1-binding molecule, regulator of adhesion and cell polarization enriched in lymphoid tissues (RAPL). RAPL is predominantly expressed in immune cells, and mediates Rap1-triggered integrin activation upon TCR engagement and chemokine stimulation. Importantly, Rap1/RAPL signaling cooperatively regulates cell polarization and integrin activation. The linkage between cell polarization and integrin activation through Rap1/RAPL signaling likely provides immune cells with their dynamic trafficking capability.
Journal of Biological Chemistry, 2002
In T-lymphocytes the Ras-like small GTPase Rap1 plays an essential role in stimulus-induced inside-out activation of integrin LFA-1 (␣ L  2) and VLA-4 (␣ 4  1). Here we show that Rap1 is also involved in the direct activation of these integrins by divalent cations or activating antibodies. Inhibition of Rap1 either by Rap GT-Pase-activating protein (RapGAP) or the Rap1 binding domain of RalGDS abolished both Mn 2؉-and KIM185 (anti-LFA-1)-induced LFA-1-mediated cell adhesion to intercellular adhesion molecule 1. Mn 2؉-and TS2/16 (anti-VLA-4)-induced VLA-4-mediated adhesion were inhibited as well. Interestingly, both Mn 2؉ , KIM185 and TS2/16 failed to induce elevated levels of Rap1GTP. These findings indicate that available levels of GTPbound Rap1 are required for the direct activation of LFA-1 and VLA-4. Pharmacological inhibition studies demonstrated that both Mn 2؉-and KIM185-induced adhesion as well as Rap1-induced adhesion require intracellular calcium but not signaling activity of the MEK-ERK pathway. Moreover, functional calmodulin signaling was shown to be a prerequisite for Rap1-induced adhesion. From these results we conclude that in addition to stimulus-induced inside-out activation of integrins, active Rap1 is required for cell adhesion induced by direct activation of integrins LFA-1 and VLA-4. We suggest that Rap1 determines the functional availability of integrins for productive binding to integrin ligands.
Immunological Reviews, 2007
Integrins are critical for the migration of T cells to lymphoid organs and to sites of inflammation and are also necessary for productive interactions between T cells and antigen-presenting cells. Integrin activation is enhanced following T-cell receptor (TCR) engagement, as signals initiated by the TCR increase affinity and avidity of integrins for their ligands. This process, known as inside-out signaling, has been shown to require several molecular components including the cytosolic adapter proteins adhesion and degranulation-promoting adapter protein and Src homology 2 domain-containing adapter protein of 55 kDa, the low molecular weight guanosine triphosphatase Rap1, and the Rap1 effector proteins Rap1 guanosine triphosphate-interacting adapter molecule, regulator of adhesion and cell polarization enriched in lymphoid tissues, and protein kinase D1. Herein, we review recent findings about how the TCR is linked to integrin activation through inside-out signaling.
Rap1 up-regulation and activation on plasma membrane regulates T cell adhesion
The Journal of Cell Biology, 2004
ap1 and Ras are closely related GTPases that share some effectors but have distinct functions. We studied the subcellular localization of Rap1 and its sites of activation in living cells. Both GFP-tagged Rap1 and endogenous Rap1 were localized to the plasma membrane (PM) and endosomes. The PM association of GFP-Rap1 was dependent on GTP binding, and GFP-Rap1 was rapidly up-regulated on this compartment in response to mitogens, a process blocked by inhibitors of endosome recycling. A novel fluorescent probe for GTP-bound Rap1 revealed that R this GTPase was transiently activated only on the PM of both fibroblasts and T cells. Activation on the PM was blocked by inhibitors of endosome recycling. Moreover, inhibition of endosome recycling blocked the ability of Rap1 to promote integrin-mediated adhesion of T cells. Thus, unlike Ras, the membrane localizations of Rap1 are dynamically regulated, and the PM is the principle platform from which Rap1 signaling emanates. These observations may explain some of the biological differences between these GTPases.
Rap1A-deficient T and B cells show impaired integrin-mediated cell adhesion
Molecular and cellular biology, 2006
Studies in tissue culture cells have demonstrated a role for the Ras-like GTPase Rap1 in the regulation of integrin-mediated cell-matrix and cadherin-mediated cell-cell contacts. To analyze the function of Rap1 in vivo, we have disrupted the Rap1A gene by homologous recombination. Mice homozygous for the deletion allele are viable and fertile. However, primary hematopoietic cells isolated from spleen or thymus have a diminished adhesive capacity on ICAM and fibronectin substrates. In addition, polarization of T cells from Rap1-/- cells after CD3 stimulation was impaired compared to that of wild-type cells. Despite this, these defects did not result in hematopoietic or cell homing abnormalities. Although it is possible that the relatively mild phenotype is a consequence of functional complementation by the Rap1B gene, our genetic studies confirm a role for Rap1A in the regulation of integrins.