PF16 ENCODES A PROTEIN WITH ARMADILLO REPEATS AND LOCALIZES TO A SINGLE MICROTUBULE OF THE CENTRAL APPARATUS IN CHLAMYDOMONAS FLAGELLA

Several studies have indicated that the central pair of microtubules a nd their associated structures play a significant role in regulating f lagellar motility. To begin a molecular analysis of these components w e have generated central apparatus-defective mutants in Chlamydomonas reinhardtii using insertional mutagenesis. One paralyzed mutant recove red in our screen, D2, is an allele of a previously identified mutant, pf16. Mutant cells have paralyzed flagella, and the C1 microtubule of the central apparatus is missing in isolated axonemes. We have cloned the wild-type PF16 gene and confirmed its identity by rescuing pf16 m utants upon transformation. The rescued pf16 cells were wild-type in m otility and in axonemal ultrastructure. A full-length cDNA clone for P F16 was obtained and sequenced. Database searches using the predicted 566 amino acid sequence of PF16 indicate that the protein contains eig ht contiguous armadillo repeats. A number of proteins with diverse cel lular functions also contain armadillo repeats including pendulin, Rch 1, importin, SRP-1, and armadillo. An antibody was raised against a fu sion protein expressed from the cloned cDNA. Immunofluorescence labeli ng of wild-type flagella indicates that the PF16 protein is localized along the length of the flagella while immunogold labeling further loc alizes the PF16 protein to a single microtubule of the central pair. B ased on the localization results and the presence of the armadillo rep eats in this protein, we suggest that the PF16 gene product is involve d in protein-protein interactions important for C1 central microtubule stability and flagellar motility.