Flagellar protein dynamics in Chlamydomonas

Cilia and flagella appear to be stable, terminal, microtubule-containing or ganelles, but they also elongate and shorten in response to a variety of si gnals. To understand mechanisms that regulate flagellar dynamics, Chlamydom onas cells with nongrowing flagella were labeled with S-35, and flagella an d basal body components were examined for labeled polypeptides. Maximal inc orporation of label into the flagella occurred within 3 h. Twenty percent o f the flagellar polypeptides were exchanged. These included tubulins, dynei ns, and 80 other axonemal and membrane plus matrix polypeptides. The most s table flagellar structure is the PF-ribbon, which comprises part of the wal l of each doublet microtubule and is composed of tubulin and three other po lypeptides. Most 35S was incorporated into the high molecular weight ribbon polypeptide, rib240, and little, if any, S-35 is incorporated into PF-ribb on-associated tubulin. Both wild-type (9 + 2) and 9 + 0 flagella, which lac k central microtubules, exhibited nearly identical exchange patterns, so la beling is not due to turnover of relatively labile central microtubules. To determine if flagellar length is balanced by protein exchange, 35S incorpo ration into disassembling flagella was examined, as was exchange in flagell a in which microtubule assembly was blocked by colchicine. Incorporation of S-35-Iabeled polypeptides was found to occur into flagellar axonemes durin g wavelength-dependent shortening in pf78 and in fla-10 cells induced to sh orten flagella by incubation at 33 degreesC. Colchicine blocked tubulin add ition but did not affect the exchange of the other exchangeable polypeptide s; nor did it induce any change in flagellar length. Basal bodies also inco rporated newly synthesized proteins. These data reveal that Chlamydomonas f lagella are dynamic structures that incorporate new protein both during ste ady state and as flagella shorten and that protein exchange does not, alone , explain length regulation.