Central helix mutations in the centrosome-associated EF-hand protein centrin

Summary Centrin is a unique member of the EF-hand superfamily that is found in calcium-modulated contractile fibers associated with the centrosomes in a wide variety of higher and lower eukaryotes. InChlamydomonas reinhardtii centrin is encoded by the genevfl 2. We previously showed that thevfl 2–220 mutation is a glutamic acid to lysine change at position 101 that results in the loss of the centrin-containing fibrous structures. Here we describe several phenotypic revertants of thevfl 2–220 mutation that carry second site suppressors at amino acid position 96 or 104. Even though these new mutations are able to suppress the variable flagellar number phenotype, they are not wild type at the ultrastractural level but instead show specific defects in their distal striated fibers and transition-region stellate fibers. The nucleus-basal body connectors are, however, functional, suggesting that the reversion of the variable flagellar number phenotype occurs through restoration of proper connections of the basal body apparatus to the nucleus, as has been previously proposed. Positions 96, 101, and 104 lie within centrin's central helix, indicating that, as is the case for calmodulin, this part of the protein is of particular importance to its function.Abbreviations NBBC nucleus-basal body connector - NFA nucleoflagellar apparatus