Identification and functional characterization of HSH2, a dynamically expressed adaptor protein implicated in B cell survival signaling

HSH2 was identified in a bioinformatics-based search for novel adaptor proteins that are preferentially expressed in B lymphocytes. Interaction of hematopoietic Src homology 2 (HSH2) with the anti-apoptotic BCL-2 family protein Hax-1 implicated HSH2 in regulation of apoptosis. Expression of HSH2 in the WEHI-231 immature B cell line by retroviral transduction attenuates antigen receptor-induced apoptosis by preventing mitochondrial membrane depolarization. Furthermore, endogenous HSH2 expression is diminished by pro-apoptotic antigen receptor stimulation. In contrast, pro-survival signaling through CD40 up-regulates HSH2 expression and prevents antigen receptor-mediated down-regulation of HSH2. HSH2 expression is also up-regulated by anti-CD40, lipopoly-saccharide (LPS), CpG DNA, and BlyS stimulation of murine splenic B cells. Anti-CD40, LPS, and CpG DNA activate numerous B cell processes; including differentiation, proliferation, and survival; BlyS primarily regulates B cell homeostasis and survival and does not induce proliferation or plasma cell differentiation. All of these stimuli promote B cell survival by up-regulation of anti-apoptotic molecules through NF-kappaB activation. Treatment of splenic B cells with inhibitors of NF-kappaB activation prevented up-regulation of HSH2 by anti-CD40, LPS, CpG DNA, and BlyS, suggesting HSH2 expression is NF-kappaB dependent. HSH2 expression is also diminished by apoptosis-inducing stimulation by Interleukin (IL)-21. IL-2l stimulation induces apoptosis of anti-CD40-, LPS-, and CpG DNA-treated splenic B cells by down-regulating anti-apoptotic molecules such as Bcl-xL and up-regulating pro-apoptotic proteins like Bim that potentiate mitochondrial membrane depolarization. Therefore, HSH2 expression is coordinately regulated with known anti-apoptotic molecules under conditions that promote B cell survival. These data suggest that HSH2 is a component of a pro-survival signaling module that functions to prevent B cell apoptosis by maintaining mitochondrial membrane stability.