Peflin and ALG-2, members of the penta-EF-hand protein family, form a heterodimer that dissociates in a Ca2+-dependent manner

Peflin, a newly identified 30-kDa Ca2+-binding protein, belongs to the pent a-EF-hand (PEF) protein family, which includes the calpain small subunit, s orcin, grancalcin, and ALG-2 (apoptosis-linked gene 2). We prepared a monoc lonal antibody against human peflin, The antibody immunoprecipitated a 22-k Da protein as well as the 30-kDa protein from the lysate of Jurkat cells. W estern blotting of the immunoprecipitates revealed that the 22-kDa protein corresponds to ALG-2, This was confirmed by Western blotting of the immunop recipitates of epitope-tagged peflin or ALG-2 whose cDNA expression constru cts were transfected to human embryonic kidney (HEK) 293 cells. Gel filtrat ion of the cytosolic fraction of Jurkat cells revealed co-elution of peflin and ALG-S in fractions eluting earlier than recombinant ALG-S, further sup porting the notion of heterodimerization of the two PEF proteins. Surprisin gly, peflin dissociated from ALG-S in the presence of Ca2+ Peflin and ALG-S co-localized in the cytoplasm, but ALG-2 was also detected in the nuclei a s revealed by immunofluorescent staining and subcellular fractionation, Pef lin was recovered in the cytosolic fraction in the absence of Ca2+ but in t he membrane/cytoskeletal fraction in the presence of Ca2+. These results su ggest that peflin has features common to those of other PEF proteins (dimer ization and translocation to membranes) and may modulate the function of AL G-2 in Ca2+ signaling.