SYNTHESIS AND TURNOVER OF EMBRYONIC SEA-URCHIN CILIARY PROTEINS DURING SELECTIVE-INHIBITION OF TUBULIN SYNTHESIS AND ASSEMBLY

When ciliogenesis first occurs in sea urchin embryos, the major buildi ng block proteins, tubulin and dynein, exist in substantial pools, but most 9+2 architectural proteins must be synthesized de novo. Pulse-ch ase labeling with [H-3]leucine demonstrates that these proteins are co ordinately up-regulated in response to deciliation so that regeneratio n ensues and the tubulin and dynein pools are replenished. Protein lab eling and incorporation into already-assembled cilia is high, indicati ng constitutive ciliary gene expression and steady-state turnover. To determine whether either the synthesis of tubulin or the size of its a vailable pool is coupled to the synthesis or turnover of the other 9+2 proteins in some feedback manner, fully-ciliated mid-or late-gastrula stage Strongylocentrotus droebachiensis embryos were pulse labeled in the presence of colchicine or taxol at concentrations that block cili ary growth. As a consequence of tubulin autoregulation mediated by inc reased free tubulin, no labeling of ciliary tubulin occurred in colchi cine-treated embryos. However, most other proteins were labeled and in corporated into steady-state cilia at near-control levels in the prese nce of colchicine or taxol. With taxol, tubulin was labeled as well. A n axoneme-associated 78 kDa cognate of the molecular chaperone HSP70 c orrelated with length during regeneration; neither colchicine nor taxo l influenced the association of this protein in steady-state cilia. Th ese data indicate that 1) ciliary protein synthesis and turnover is in dependent of tubulin synthesis or tubulin pool size; 2) steady-state i ncorporation of labeled proteins cannot be due to formation or elongat ion of cilia; 3) substantial tubulin exchange takes place in fully-mot ile cilia; and 4) chaperone presence and association in steady-state c ilia is independent of background ciliogenesis, tubulin synthesis, and tubulin assembly state.