The Protein Kinase C-Responsive Inhibitory Domain of CARD11 Functions in NF-κB Activation To Regulate the Association of Multiple Signaling Cofactors That Differentially Depend on Bcl10 and MALT1 for Association

Abstract

The activation of NF-B by T-cell receptor (TCR) signaling is critical for T-cell activation during the adaptive immune response. CARD11 is a multidomain adapter that is required for TCR signaling to the IB kinase (IKK) complex. During TCR signaling, the region in CARD11 between the coiled-coil and PDZ domains is phosphorylated by protein kinase C (PKC) in a required step in NF-B activation. In this report, we demonstrate that this region functions as an inhibitory domain (ID) that controls the association of CARD11 with multiple signaling cofactors, including Bcl10, TRAF6, TAK1, IKK␥, and caspase-8, through an interaction that requires both the caspase recruitment domain (CARD) and the coiled-coil domain. Consistent with the ID-mediated control of their association, we demonstrate that TRAF6 and caspase-8 associate with CARD11 in T cells in a signal-inducible manner. Using an RNA interference rescue assay, we demonstrate that the CARD, linker 1, coiled-coil, linker 3, SH3, linker 4, and GUK domains are each required for TCR signaling to NF-B downstream of ID neutralization. Requirements for the CARD, linker 1, and coiled-coil domains in signaling are consistent with their roles in the association of CARD11 with Bcl10, TRAF6, TAK1, caspase-8, and IKK␥. Using Bcl10-and MALT1-deficient cells, we show that CARD11 can recruit signaling cofactors independently of one another in a signal-inducible manner.The NF-B family of transcription factors plays important pleiotropic roles in the regulation of cellular activation, proliferation, and survival. The precise regulation of NF-B activity is critical for several biological processes including innate and adaptive immunity (22, 54), learning and memory (34), epidermal development and homeostasis (2), bone formation and metabolism (65), and embryonic development (19). In most normal cells, NF-B is inactive but is poised for rapid posttranslational activation by a diverse array of stimuli such as bacterial and viral products, proinflammatory cytokines, antigens recognized by lymphocytes, DNA-damaging agents, and UV irradiation.A remarkable aspect of NF-B regulation is that each of these stimuli can activate the IB kinase complex (IKK complex) (50). An emerging theme is that a ligand-receptor pair that may be specialized to a particular biological context will signal to the IKK complex through adapter molecules that can receive stimulusspecific molecular signals and transmit that information to widely expressed signaling cofactors that are of general use. The IKK complex activates NF-B by canonical and noncanonical pathways (50), both of which link IKK kinase activity to the phosphorylation and degradative processing of inhibitory proteins that keep NF-B inactive in the unstimulated state.NF-B is one of the key transcription factors that are induced when a T lymphocyte is activated by antigenic stimulation in the adaptive immune response (51). Following the engagement of antigen by the T-cell receptor (TCR) complex and concomitant triggering of costimulatory receptors like CD...

NF-κB Activation in T Cells Requires Discrete Control of IκB Kinase α/β (IKKα/β) Phosphorylation and IKKγ Ubiquitination by the ADAP Adapter Protein

“…ADAP also appears to play a dual role in regulating recruitment of NF-B signaling proteins to the PKC signalosome as distinct sites in ADAP are critical for the recruitment of TAK1 and CARMA1 to PKC. Although TAK1 and CARMA1 have been proposed to interact with each other, TAK1 recruitment to PKC and IKK␣/␤ phosphorylation is not altered in CARMA1-deficient T cells (11,16,17). We suggest that ADAP plays a central role in this CARMA1-independent recruitment of TAK1 to PKC.…”

“…The mechanism by which TAK1 is recruited to the PKC signalosome is unclear. Although CARMA1 and TAK1 have been reported to associate with each other (11,16,17), TAK1-mediated IKK␣/␤ phosphorylation is intact in CARMA1-deficient Jurkat T cells (11). This suggests that IKK␥ ubiquitination and IKK␣/␤ phosphorylation are independently controlled.…”

NF-κB signaling pathways regulated by CARMA family of scaffold proteins

“…The linker region is phosphorylated by PKCθ (or PKCβ in B cells) in a signal-dependent manner ( Figure 4A) [19,64] and two putative PKC phosphorylation sites, Ser552 and Ser645 (in mouse: Ser564 and Ser657), were identified and confirmed in the linker region of human CARMA1 [19,64,65]. The inducible phosphorylation of CARMA1 by PKC results in conformational changes that enable CARMA1 to associate with its downstream signaling components [19,64,66].…”

A Quantitative Signaling Screen Identifies CARD11 Mutations in the CARD and LATCH Domains That Induce Bcl10 Ubiquitination and Human Lymphoma Cell Survival

“…As a consequence of BCR or TCR engagement, CARD11 undergoes a conformational transition from a closed, inactive state to an open, active scaffold. This transition is controlled by an inhibitory domain (ID), located between the coiled-coil and PDZ domains, that keeps CARD11 in the closed, latent state through interactions that require the CARD and coiled-coil domains (13)(14)(15). Antigen receptor signaling leads to the neutralization of the ID through its phosphorylation at specific serine residues by protein kinase C (PKC) in T cells, PKC␤ in B cells, IKK␤, and at least one additional unidentified kinase (13,(15)(16)(17)(18).…”