Exposure of phosphatidylserine by Xk-related protein family members during apoptosis

Apoptotic cells expose phosphatidylserine (PtdSer) on their surface as an "eat me" signal. Mammalian Xk-related (Xkr) protein 8, which is predicted to contain six transmembrane regions, and its Caenorhabditis elegans homolog CED-8 promote apoptotic PtdSer exposure. The mouse and human Xkr families consist of eight and nine members, respectively. Here, we found that mouse Xkr family members, with the exception of Xkr2, are localized to the plasma membrane. When Xkr8-deficient cells, which do not expose PtdSer during apoptosis, were transformed by Xkr family members, the transformants expressing Xkr4, Xkr8, or Xkr9 responded to apoptotic stimuli by exposing cell surface PtdSer and were efficiently engulfed by macrophages. Like Xkr8, Xkr4 and Xkr9 were found to possess a caspase recognition site in the C-terminal region and to require its direct cleavage by caspases for their function. Site-directed mutagenesis of the amino acid residues conserved among CED-8, Xkr4, Xkr8, and Xkr9 identified several essential residues in the second transmembrane and second cytoplasmic regions. Real time PCR analysis indicated that unlike Xkr8, which is ubiquitously expressed, Xkr4 and Xkr9 expression is tissue-specific.