The Anti-apoptotic Protein HAX-1 Interacts with SERCA2 and Regulates Its Protein Levels to Promote Cell Survival

Cardiac contractility is regulated through the activity of various key Ca2+-handling proteins. The sarco(endo) plasmic reticulum (SR) Ca2+ transport ATPase (SERCA2a) and its inhibitor phospholamban (PLN) control the uptake of Ca2+ by SR membranes during relaxation. Recently, the antiapoptotic HS-1-associated protein X-1 (HAX-1) was identified as a binding partner of PLN, and this interaction was postulated to regulate cell apoptosis. In the current study, we determined that HAX-1 can also bind to SERCA2. Deletion mapping analysis demonstrated that amino acid residues 575-594 of SERCA2's nucleotide binding domain are required for its interaction with the C-terminal domain of HAX-1, containing amino acids 203-245. In transiently cotransfected human embryonic kidney 293 cells, recombinant SERCA2 was specifically targeted to the ER, whereas HAX-1 selectively concentrated at mitochondria. On triple transfections with PLN, however, HAX-1 massively translocated to the ER membranes, where it codistributed with PLN and SERCA2. Overexpression of SERCA2 abrogated the protective effects of HAX-1 on cell survival, after hypoxia/reoxygenation or thapsigargin treatment. Importantly, HAX-1 overexpression was associated with down-regulation of SERCA2 expression levels, resulting in significant reduction of apparent ER Ca2+ levels. These findings suggest that HAX-1 may promote cell survival through modulation of SERCA2 protein levels and thus ER Ca2+ stores.