Regulation of the Small GTPase Rap1 and Extracellular Signal-Regulated Kinases by the Costimulatory Molecule CTLA-4

Abstract

The mitogen-activated protein kinase extracellular signal-regulated kinase (ERK) is activated following engagement of the T-cell receptor and is required for interleukin 2 (IL-2) production and T-cell proliferation. This activation is enhanced by stimulation of the coreceptor CD28 and inhibited by the coreceptor CTLA-4. We show that the small G protein Rap1 is regulated in the opposite manner; it is inhibited by CD28 and activated by CTLA-4. Together, CD3 and CTLA-4 activate Rap1 in a sustained manner. To delineate T-cell function in the absence of Rap1 activity, we generated transgenic mice expressing Rap1GAP1, a Rap1-specific GTPaseactivating protein. Transgenic mice showed lymphadenopathy, and transgenic T cells displayed increased ERK activation, proliferation, and IL-2 production. More significantly, the inhibitory effect of CTLA-4 on T-cell function in Rap1GAP1-transgenic T cells was reduced. We demonstrate that CTLA-4 activates Rap1, and we propose that intracellular signals from CTLA-4 antagonize CD28, at least in part, at the level of Rap1.The activation of T cells requires antigen recognition through the T-cell receptor (TCR) and costimulation via the CD28 coreceptor, which binds the family of B7 ligands (B7-1 and B7-2) expressed on antigen-presenting cells (APCs) (35). Additional complexity comes with another coreceptor, CTLA-4, that also binds B7 and can antagonize the ability of CD28 to stimulate T-cell activation and interleukin 2 (IL-2) accumulation (25,27,28,42,44). Recent studies have suggested that inhibition of extracellular signal-regulated kinases (ERKs) may contribute to this action (6, 9, 38). ERKs are activated following costimulation by TCR/CD28 and are required for AP-1-dependent transcription of IL-2, a cytokine that is essential for T-cell proliferation (32, 46). Cross-linking anti-CTLA-4 antibodies (that activate CTLA-4) inhibit both ERK activation and IL-2 production following costimulation by TCR/CD28 (6, 9). The mechanism by which CTLA-4 inhibits ERK activation is not known.The small G protein Ras mediates TCR activation of ERKs (7). TCR stimulation triggers the activation of Ras via the Ras guanine nucleotide exchange factors (GEF) Cal DAG-GEF II (also called Ras-GRP1) (17) and SOS (47). Ras is a positive activator of the mitogen-activated protein (MAP) kinase kinase kinase Raf-1 that can phosphorylate and activate the MAP kinase kinase MEK, which, in turn, can activate ERK. Potential negative regulatory molecules are also activated by TCR stimulation. One of these is the small G protein Rap1 that, like Ras, can be inhibited by specific GTPase-activating proteins (Rap1GAPs). Rap1 is activated following stimulation of the TCR (1, 8) and, like Ras, is rapidly recruited to the plasma membrane upon activation (3). During CD28 costimulation, Rap1 activation is inhibited (8,22,33) and ERK signaling is enhanced (8). Conversely, T lymphocytes lacking the Rap1GAP Spa-1 show constitutively elevated Rap1 activity, diminished ERK activation, and a decreased response to TCR stimulation (21). ...

Suppressing T cell motility induced by anti–CTLA-4 monotherapy improves antitumor effects

“…Instead, our data provide evidence for a context-dependent regulation of T cell motility by CTLA-4, which is modulated by co-ligation of NKG2D. Integrin activation may be one component of this effect (21,62).…”

Multiple roles of Rap1 in hematopoietic cells: complementary versus antagonistic functions

“…on June 19, 2019. by guest www.bloodjournal.org From activation by TCR with Rap1GAP1 expression and enhanced TCR-mediated ERK activation. 116 These and related studies 67 have demonstrated a role for Rap1 to uncouple Ras-mediated ERK activation in peripheral T cells. 113 TCR function is enhanced by stimulation of the coreceptor CD28, which is characterized by enhanced ERK activation and IL-2 production 67,117 and the inhibition of Rap1 52,54 without effecting Ras.…”

“…113 TCR function is enhanced by stimulation of the coreceptor CD28, which is characterized by enhanced ERK activation and IL-2 production 67,117 and the inhibition of Rap1 52,54 without effecting Ras. 113,116 It has been proposed that CD28 does this by inducing endogenous Rap1GAP activity. 67 In contrast to CD28, the coreceptor CTLA-4 (cytotoxic T lymphocyte associated protein-4) inhibits T-cell activation by limiting multiple signals downstream of TCR, including ERKs.…”

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“…kip1 that are implicated in T cell anergy and inhibition of cell cycle (49,50). In human filariasis, T cell unresponsiveness is associated with increased expression of E3 ubiquitin ligases (c-cbl, cbl-b, Itch, and Nedd4) that are often up-regulated in anergic T cells (10).…”