Proteomic analysis of flagella from Chlamydomonas mutants lacking the central pair apparatus reveals loss of radial spoke head proteins

Abstract

The flagellar axoneme is a complex protein-based machine capable of generating motile forces by coordinating the action of thousands of dynein motors. A key element of the axoneme is the central pair apparatus, consisting of a pair of microtubules surrounded by additional structures. In an effort to better understand the organization of the central pair, we used 2D DIGE to identify proteins that are depleted from flagella isolated from two different Chlamydomonas reinhardtii mutants, pf15 and pf18, that lack the central pair. The set of proteins contained almost no components of the central apparatus. We find that three proteins of the radial spoke head RSP1, RSP9, and RSP10, as well as a number of other protein components associated with the outer doublets, are depleted from flagella of mutants lacking the central apparatus. Two of the other proteins depleted from pf15 and pf18 flagella, the microtubule inner proteins (MIPs) FAP21 and FAP161, are missing from the genome of Thalassiosira, an organism that lacks a central pair and radial spokes, and RNAi of FAP21 in planaria shows that it has a role in ciliary motility. Based on the depletion of radial spoke head proteins, as well as MIPs and other axonemal components, from flagella lacking the central pair apparatus, we hypothesize that the central apparatus may play a role in scaffolding the assembly or retention of radial spokes and other axonemal structures.