The molecular basis of IP3R recognition by the ubiquitin-proteasome pathway

Abstract

Inositol 1,4,5-trisphosphate (IP3) receptors (IP3Rs) form ~1.2MDa tetrameric Ca2+ channels in the endoplasmic reticulum (ER) membrane of mammalian cells. IP3R1 is the most ubiquitously expressed among all three IP3R isoforms. Upon activation by the second messenger, IP3, IP3Rs undergo a conformational change that leads to channel opening and allows Ca2+ ions to flow from the ER stores into the cytosol. IP3R-dependent Ca2+ signaling is crucial to many cellular events. The Wojcikiewicz laboratory has found that active IP3Rs are quickly processed by the ubiquitin-proteasome pathway (UPP), which is initiated by their association with the erlin1/2 complex. The association also recruits the E3 ligaseRNF170 to active IP3Rs. However, how activated IP3Rs are recognized by the erlin1/2 complex remains unclear. Using IP3R mutants, we discovered that the erlin1/2 complex binding site is on the third intraluminal loop (IL3) of IP3R and also found that a region at the N-terminus of IL3 is critical to IP3R channel activity. We also used UPP inhibitor TAK-243 to confirm the sequence of events that leads to IP3R processing by the UPP: the erlin1/2 complex association is prior to IP3R ubiquitination and degradation. Surprisingly, we found that long-term treatment with UPP inhibitors can inhibit IP3R-mediated Ca2+ signaling and affect other aspects of Ca2+ handling in cells. Overall, these results help us understand how the large ion channels are deconstructed and further our knowledge of substrate processing by the UPP.