Scaffolding function of theChlamydomonasprocentriole protein CRC70, a member of the conserved Cep70 family

Abstract

Centriole duplication occurs once per cell cycle through the assembly of daughter centrioles on the side wall of pre-existing centrioles. Little is known about the molecules involved in the assembly of new centrioles. Here, we identify CRC70 as a Chlamydomonas protein with an important role in the accumulation of centriole proteins at the site of assembly. CRC70 contains a highly conserved ~50-amino-acid sequence shared by mammalian Cep70 and preferentially localizes to immature centrioles (the procentrioles). This localization is maintained in the mutant bld10, in which centriole formation is blocked before the assembly of centriolar microtubules. RNA interference (RNAi)-mediated knockdown of CRC70 produces flagella-less cells and inhibits the recruitment of other centriole components, such as SAS-6 and Bld10p to the centriole. Overexpression of CRC70 induces an accumulation of these proteins in discrete spots in the cytoplasm. Overexpression of EGFP-tagged CRC70 in mouse NIH3T3 cells causes the formation of structures apparently related to centrioles. These findings suggest that CRC70 is a member of a conserved protein family and functions as a scaffold for the assembly of the centriole precursor.