Radial spokes of Chlamydomonas flagella: polypeptide composition and phosphorylation of stalk components.

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Abstract
Polypeptides from flagella or axonemes of C. reinhardtii were analyzed by labeling cellular proteins by prolonged growth on 35S-containing media and using 1- and 2-dimensional electrophoresis which resolved > 170 axonemal components. A paralyzed mutant that lacked axonemal radial spokes, pf14, lacked 17 polypeptides in the MW range of 20,000-124,000 and in the isoelectric point range of 4.8-7.1. Of those polypeptides were missing in the mutant pf-1 which lacked only radial spokeheads. Identification of the 17 polypeptides missing in pf-14 as components of radial spoke structures and localization of the polypeptides lacking in pf-1 within the spokehead were determined by chemical dissection of wild-type axonemes. Extraction procedures that solubilized outer and inner dynein arms preserved the structure of the radial spokes along with the 17 polypeptides in question. Six radial spoke polypeptides were solubilized in conditions that caused disassembly of radial spokeheads from the stalks and those components included the 5 polypeptides missing in pf-1. No Ca2+- or Mg2+-activated ATPase activities were associated with solubilized preparations of wild-type radial spokeheads. In vivo pulse 32P incorporation experiments showed that > 80 axonemal components were labeled by 32P and that 5 of the radial spoke stalk polypeptides were modified to different extents.