title: Synthesis and turnover of embryonic sea urchin ciliary proteins during selective inhibition of tubulin synthesis and assembly.
abstract: When ciliogenesis first occurs in sea urchin embryos, the major building block proteins, tubulin and dynein, exist in substantial pools, but most 9 + 2 architectural proteins must be synthesized de novo. Pulse-chase labeling with [3H]leucine demonstrates that these proteins are coordinately up-regulated in response to deciliation so that regeneration ensues and the tubulin and dynein pools are replenished. Protein labeling and incorporation into already-assembled cilia is high, indicating constitutive ciliary gene expression and steady-state turnover. To determine whether either the synthesis of tubulin or the size of its available 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 ciliary growth. As a consequence of tubulin autoregulation mediated by increased free tubulin, no labeling of ciliary tubulin occurred in colchicine-treated embryos. However, most other proteins were labeled and incorporated into steady-state cilia at near-control levels in the presence of colchicine or taxol. With taxol, tubulin was labeled as well. An axoneme-associated 78 kDa cognate of the molecular chaperone HSP70 correlated with length during regeneration; neither colchicine nor taxol influenced the association of this protein in steady-state cilia. These data indicate that 1) ciliary protein synthesis and turnover is independent of tubulin synthesis or tubulin pool size; 2) steady-state incorporation of labeled proteins cannot be due to formation or elongation of cilia; 3) substantial tubulin exchange takes place in fully-motile cilia; and 4) chaperone presence and association in steady-state cilia is independent of background ciliogenesis, tubulin synthesis, and tubulin assembly state.
authors.affiliation: Department of Physiology, Boston University School of Medicine, Massachusetts 02118, USA.
authors.forename: R E
authors.initials: RE
authors.lastname: Stephens
mesh_terms: D000818:Animals; D002923:Cilia / Q000187:drug effects / Q000378:metabolism*; D003078:Colchicine / Q000494:pharmacology; D004398:Dyneins / Q000096:biosynthesis / Q000378:metabolism*; D005775:Gastrula / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D017239:Paclitaxel / Q000494:pharmacology; D012617:Sea Urchins / Q000196:embryology; D014404:Tubulin / Q000096:biosynthesis* / Q000378:metabolism
chemical_list: D018840:HSP70 Heat-Shock Proteins; D014404:Tubulin; D004398:Dyneins; D017239:Paclitaxel; D003078:Colchicine