Control of flagellar assembly and disassembly by a microtubule depolymerase CrKinesin13
Junmin Pan
Protein Science Laboratory of the Ministry of Education, School of Life Sciences, Tsinghua University, Beijing 100084, China
In steady growing cultures, Chlamydomonas reinhardtii maintains a pair of flagella of 10-12 um long. The flagella are assembled upon completion of cell division. The flagella, however, are removed prior to next cell cycle, during zygote development and upon cellular stress. Upon flagellar loss, full length flagella may be regenerated rapidly within 2 h. Though the flagellar structure, flagellar proteome and the intraflagllar transport (IFT) system are well studied, the regulation of flagellar assembly and disassembly are poorly understood. We report here that a Chlamydomonas homologue of kinesin 13 family members - CrKinesin13 (CrKin13) controls both flagellar assembly and disassembly by regulating microtubule dynamics. During flagellar shortening, CrKin13, which is localized predominantly in the cell body, is timely transported to the flagella. Flagellar transport of CrKinesin13 is defective at restrictive temperature in fla10, a temperature sensitive mutant defective in IFT motor protein FLA10. Furthermore, CrKin13 is co-immunoprecipitated with IFT139, indicating that IFT is required for CrKin13 transport, which is in contrast with diffusion model of mammalian kinesin13 transport. Suppressing expression of CrKinesin13 by RNAi impairs flagellar disassembly. During flagellar assembly, CrKin13 is phosphorylated and RNAi knockdown strains of CrKin13 exhibits shorter flagella. The cell body pool of tubulin is increased before flagellar assembly shortly after flagellar loss. Taxol treatment impairs cell body pool of tubulin and completely blocks flagellar assembly of CrKin13 RNAi strains. These data demonstrate that CrKin13 controls flagellar assembly and disassembly by temporal and spatial regulation of microtubule dynamics. This work was supported by NSFC (Grants 30830057, 30988004 and 30771084), National Basic Research Program of China [also called 973 program] (Grant 2007CB914401) and SRFDP.
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