The Bok-IP3R interaction and the impact it has on apoptosis, Bok stability, and IP3R-mediated calcium mobilization

Abstract

Bcl-2-related ovarian killer, Bok, is often considered a pro-apoptotic Bcl-2 family member due to its ability to induce mitochondrial outer membrane permeabilization (MOMP) when over-expressed. However, there are many conflicting reports regarding endogenous Bok's apoptotic function and stability. Some studies claim that endogenous Bok is constitutively a MOMP-mediator but is very unstable and kept at very low levels via ubiquitin proteasome pathway (UPP)-mediated degradation. On the other hand, many studies, including those from the Wojcikiewicz Lab, indicate that endogenous Bok is a stable protein constitutively bound to inositol 1,4,5-trisphosphate receptors (IP3Rs), a calcium (Ca2+) release channel in the endoplasmic reticulum (ER) membrane, where it would be unable to induce MOMP. In fact, Bok KO mice have no developmental of phenotypical abnormalities, further suggesting that Bok's "killer" characterization needs to be reconsidered. Since endogenous Bok is an ER protein, studying its MOMP-like properties seems futile. Therefore, focusing on the significance of the Bok-IP3R interaction has been the priority of the Wojcikiewicz Lab since we discovered the complex. Over the years, we found that Bok protects IP3Rs from proteolytic cleavage and that Bok stability is dependent on its ability to bind to IP3Rs. However, surprisingly, there is no clear evidence to date that Bok influences the Ca2+ mobilizing activity of IP3Rs. Here, I study in detail the Bok-IP3R interaction and the impact it has on apoptosis, Bok stability, and IP3R-mediated Ca2+ mobilization. I resolve the controversy surrounding the apoptotic function of exogenous and endogenous Bok and hypothesize why there is a discrepancy between the two. Also, I show that IP3R binding in the ER membrane is essential for Bok stability, and without them Bok is rapidly degraded by the UPP. This demonstrates that the Bok-IP3R interaction is critical to Bok function, likely including its newly emerged non-apoptotic roles, such as regulating mitochondrial dynamics and mitochondria associated-ER membranes. Finally, for the first time, I reveal that Bok has suppressive effects on IP3R-mediated Ca2+ mobilization and describe how these effects are reversed when Bok is phosphorylated at serine-8. Overall, this thesis demonstrates the importance of the Bok-IP3R interaction on both Bok and IP3R function.