Transport and arrangement of the outer-dynein-arm docking complex in the flagella of Chlamydomonas mutants that lack outer dynein arms.
The outer dynein arms of Chlamydomonas flagella are attached to a precise site on the outer doublet microtubules and repeat at a regular interval of 24 nm. This binding is mediated by the outer dynein arm docking complex (ODA-DC), which is composed of three protein subunits. In this study, antibodies against the 83- and 62-kD subunits (DC83 and DC62) of the ODA-DC were used to analyze its state of association with outer arm components within the cytoplasm, and its localization in the axonemes of oda mutants. Immunoprecipitation indicates that DC83 and DC62 are preassembled within the cytoplasm, but that they are not associated with outer arm dynein. Both proteins are lost or greatly diminished in oda1 and oda3, mutants in the structural genes of DC62 and DC83, respectively, demonstrating that their association is necessary for their stable presence in the cytoplasm. Immunoelectron microscopy indicates that DC83 repeats at 24-nm intervals along the length of the doublet microtubules of oda6, which lacks outer arms; thus, outer arm periodicity may be determined by the ODA-DC. Flagellar regeneration and temporary dikaryon experiments indicate that the ODA-DC can be rapidly transported into the flagellum and assembled on the doublet microtubules independently of the outer arms and independently of flagellar growth. Unexpectedly, the intensity of ODA-DC labeling decreased toward the distal ends of axonemes of oda6 but not wild-type cells, suggesting that the outer arms reciprocally contribute to the assembly/stability of the ODA-DC.
Department of Biological Sciences, Graduate School of Science, University of Tokyo, Tokyo, Japan.
K
K
Wakabayashi
S
S
Takada
G B
GB
Witman
R
R
Kamiya
D000818:Animals; D001692:Biological Transport / Q000502:physiology; D016825:Chlamydomonas reinhardtii; D003593:Cytoplasm / Q000378:metabolism; D004398:Dyneins / Q000235:genetics* / Q000378:metabolism*; D005407:Flagella / Q000378:metabolism* / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D016253:Microscopy, Immunoelectron; D009154:Mutation / Q000502:physiology; D010507:Periodicity
D004398:Dyneins