A new multigene family encoding calcium-dependent calmodulin-binding membrane proteins of Paramecium tetraurelia

Ca2+/calmodulin (CaM) regulates various physiological processes in a wide v ariety of organisms, metazoa and protists alike. To better understand Ca2+/ CaM-dependent processes, particularly those with membrane-associated compon ents, we studied Ca2+/CaM-binding membrane proteins in Paramecium tetraurel ia, a unicellular model system. A CaM-binding protein, PCM1 (paramecium CaM -binding membrane-bound protein), from a detergent-solubilized ciliary memb rane fraction was identified and purified through Ca2+-dependent CaM-affini ty chromatography. PCM1 has an apparent molecular mass of approx. 65 kDa. I t binds radiolabeled CaM in blot overlay assays and binds to CaM-affinity c olumns, both only in the presence of 10 mu M or higher Ca2+. Three peptide sequences from PCM1 were obtained, and polymerase chain reaction (PCR) and Southern hybridization experiments were designed accordingly, leading to a partial cDNA clone for PCM1 and the discovery of three homologs: PCM2, PCM3 and PCM4. Amino-acid sequences predicted by the full-length coding sequenc e for PCM3 and partial genes for PCM1, PCM2 and PCM4 are very similar (appr ox. 85% amino-acid identities). Their sequences indicate that they are hith erto novel proteins with beta/gamma-crystallin domains, cysteine-rich regio ns and potential CaM-binding domains. These protein motifs are suggested to mediate protein-protein interaction important for Ca2+/CaM signal transduc tion event(s) through the PCM family of proteins. (C) 1999 Elsevier Science B.V. All rights reserved.