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PMID: 15616187 Published · ppublish English Journal Article Research Support, N.I.H., Extramural Research Support, U.S. Gov't, P.H.S.

The FLA3 KAP subunit is required for localization of kinesin-2 to the site of flagellar assembly and processive anterograde intraflagellar transport.

Molecular biology of the cell ·Vol. 16 ·No. 3 ·2005-03-00 ·Pages 1341-54

Mueller J, Perrone CA, Bower R, Cole DG, Porter ME

Abstract

Intraflagellar transport (IFT) is a bidirectional process required for assembly and maintenance of cilia and flagella. Kinesin-2 is the anterograde IFT motor, and Dhc1b/Dhc2 drives retrograde IFT. To understand how either motor interacts with the IFT particle or how their activities might be coordinated, we characterized a ts mutation in the Chlamydomonas gene encoding KAP, the nonmotor subunit of Kinesin-2. The fla3-1 mutation is an amino acid substitution in a conserved C-terminal domain. fla3-1 strains assemble flagella at 21 degrees C, but cannot maintain them at 33 degrees C. Although the Kinesin-2 complex is present at both 21 and 33 degrees C, the fla3-1 Kinesin-2 complex is not efficiently targeted to or retained in the basal body region or flagella. Video-enhanced DIC microscopy of fla3-1 cells shows that the frequency of anterograde IFT particles is significantly reduced. Anterograde particles move at near wild-type velocities, but appear larger and pause more frequently in fla3-1. Transformation with an epitope-tagged KAP gene rescues all of the fla3-1 defects and results in preferential incorporation of tagged KAP complexes into flagella. KAP is therefore required for the localization of Kinesin-2 at the site of flagellar assembly and the efficient transport of anterograde IFT particles within flagella.

MeSH Terms
Amino Acid Sequence Animals Binding Sites Biological Transport Blotting, Southern Blotting, Western Centromere/ultrastructure Chlamydomonas/metabolism Cilia/metabolism Cloning, Molecular DNA, Complementary/metabolism Electrophoresis, Polyacrylamide Gel Epitopes/chemistry Flagella/physiology Genetic Linkage Kinesins/biosynthesis,chemistry,metabolism,physiology Microscopy, Fluorescence Microscopy, Video Models, Genetic Molecular Sequence Data Mutation Phenotype Protein Structure, Tertiary RNA/chemistry Sequence Homology, Amino Acid Temperature Time Factors
Chemicals
DNA, Complementary Epitopes RNA Kinesins
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Mueller Joshua
Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, MN 55455, USA.
Perrone Catherine A
Bower Raqual
Cole Douglas G
Porter Mary E
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Article Info
Journal
Molecular biology of the cell
Abbr.
Mol Biol Cell
ISSN
1059-1524
DOI
10.1091/mbc.e04-10-0931
Published
2005-03-00
Epub
2004-00-22
Pages
1341-54
Language
English
Region
United States
NLM ID
9201390
PMCID
PMC551497
Subset
IM
Grants
NIGMS NIH HHS · R01 GM055667 · United States
NIGMS NIH HHS · R01 GM061920 · United States
NIGMS NIH HHS · R01 GM-55667 · United States
NIGMS NIH HHS · R01 GM-61920 · United States
External Links
PubMed source DOI: 10.1091/mbc.e04-10-0931 PMC Full Text View journal details
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