CBAP interacts with the un-liganded common β-subunit of the GM-CSF/IL-3/IL-5 receptor and induces apoptosis via mitochondrial dysfunction

Abstract

The cytoplasmic domain of the common b-chain (bc) of the granulocyte-macrophage-colony-stimulating factor (GM-CSF)/interleukin-3 (IL-3)/IL-5 receptor contains a membrane proximal region that is sufficient to mediate ligand-dependent mitogenic activity. Within this region two motifs, designated as box 1 and box 2, are highly conserved among members of the cytokine receptor superfamily. Whereas box 1 is required for the recruitment and phosphorylation of Janus kinase-2, the function of box 2 remains largely unknown. Here, we report the identification of a novel transmembrane protein (common b-chain associated protein (CBAP)) which directly associated with bc via the box 2 motif. Interestingly, such an association only occurred in the absence of GM-CSF in vivo. Ectopic overexpression of CBAP triggered apoptosis of factor-dependent cells via mitochondrial dysfunction, which could be inhibited by Bcl-2 overexpression. Reduced expression of endogenous CBAP by small interfering RNA did not interfere GM-CSF-activated signaling molecules, but such treatment significantly inhibited apoptosis induced by GM-CSF deprivation, but not other death stimuli. Domain mapping studies indicated that one apoptogenic domain of CBAP correlated with its ability to interact with bc. Taken together, these results suggest that CBAP modulates GM-CSF-deprivation-induced apoptosis possibly via a novel mechanism involving interaction with an un-liganded bc molecule.

CBAP promotes thymocyte negative selection by facilitating T-cell receptor proximal signaling

“… 18 There was no significant difference between WT and CBAP-deficient thymocytes in this in vitro assay system ( Figure 1b ). Because CBAP has a pro-apoptotic function in a variety of cell lines, 16 we next examined whether it is involved in apoptosis of DN thymocytes that fail to rearrange the TCR β gene, a process that is at least partly FADD-mediated. 19 We generated RAG1 and CBAP double-deficient mice and found that CBAP deficiency could not rescue cells from death caused by a failure of TCR β rearrangement.…”

“…We previously reported that GM-CSF/IL-3/IL-5 receptor common β -chain-associated protein (CBAP) plays a dual role in in vitro cell apoptosis 16 and in vivo T-cell migration and adhesion. 17 In T cells, CBAP is a component of the β 1-integrin complex and is involved in ZAP70-mediated VAV1 phosphorylation.…”

CBAP Functions as a Novel Component in Chemokine-Induced ZAP70-Mediated T-Cell Adhesion and Migration

“…CBAP was first identified as a binding protein of the βc subunit in hematopoietic cells [27], whereas integrin β1 was found to associate with βc in endothelial cells and plays an important role during vasculogenesis and tumor angiogenesis [25], [26], leading us to hypothesize that CBAP may also be involved in integrin-related cellular function, such as cell adhesion and migration. To this end, we generated several stable human Jurkat T-cell clones in which CBAP expression was decreased using a lentiviral vector encoding a CBAP-specific shRNA (see Materials and Methods) and tested the effect of reducing CBAP levels on T cell adhesion and migration induced by chemokine CXCL12, which is the ligand for CXCR4, the major chemokine receptor in Jurkat T cells.…”

“…The mouse monoclonal antibody against human CBAP was generated as described [27]. Polyclonal anti-mouse CBAP was generated using bacterially produced recombinant full-length GST-tagged CBAP (Geness Biotech, Taipei, Taiwan).…”

“…Our earlier study identified a novel transmembrane protein, the GM-CSF/IL-3/IL-5 receptor common beta-chain-associated protein (CBAP), from a human lymphocyte cDNA library using the yeast two-hybrid screen with the cytoplasmic domain of βc as the bait [27]. Upon cytokine deprivation, CBAP could interact directly with the unliganded βc subunit and accelerate apoptosis through a mitochondria-dependent pathway [27]. Therefore, we investigated whether there is a functional link among βc, CBAP, and the integrin β1 complex.…”