Schoppmeier, Jutta and Mages, Wolfgang and Lechtreck, Karl‐Ferdinand (2005) GFP as a tool for the analysis of proteins in the flagellar basal apparatus of Chlamydomonas. CELL MOTILITY AND THE CYTOSKELETON, 61 (4). pp. 189-200. ISSN 0886-1544,
Full text not available from this repository. (Request a copy)Abstract
Green fluorescent protein (GFP) was used to analyse three proteins in the flagellar basal apparatus of C. reinhardtii: (1) Striated fiber assemblin (SFA), the major component of the striated microtubule-associated fibers; (2) Centrin, present in the nucleus basal body connectors (NBBCs) and the distal connecting fiber (dCF) between the two basal bodies; and (3) DLP13, the Chlamdomonas homologue of human autoantigen NA14 The fusions co-localized with the wild-type proteins when expressed moderately. Overexpression of centrin-GH and DIP13-GFP resulted in the formation of large aggregates and disturbed the distribution of the respective wild-type proteins. The amount of wild-type DIP13 was significantly reduced in cells overexpressing DIP13-GFP. Moreover, the cells frequently failed to assemble full-length flagella and flagellar regeneration was delayed, indicating a role of DEP13 during flagellar assembly. In contrast, overexpression of GFP-SFA, which retained more wild-type properties than SFA-GFP, increased the size of the striated fibers without altering the cross-shaped pattern. Abnormal patterns were observed in centrin-deficient cells, suggesting that centrin is required for proper localization of SFA. Photobleaching of GFP-SFA fibers indicated that GFP-SFA in the fibers is turned over slowly. Conditionally expressed centrin-GFP was incorporated into NBBCs in regions close to the basal bodies, but underrepresented in the dCF, indicative of a different dynamic of these two centrin fibers. Bending of the NBBCs was observed in vivo during flagellar motion, indicating that the filaments are flexible. In conclusion, in Chlamydomonas GFP-tagging is a useful tool for yielding new insights into the function and properties of the analyzed proteins.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | GREEN FLUORESCENT PROTEIN; NUCLEAR TRANSFORMATION; SF-ASSEMBLIN; REINHARDTII; CENTRIN; GENE; CELLS; CENTROSOME; EXPRESSION; MOVEMENT; centrin; DIP13; striated fiber assemblin; flagella; basal body |
| Subjects: | 500 Science > 570 Life sciences |
| Divisions: | Biology, Preclinical Medicine > Institut für Biochemie, Genetik und Mikrobiologie > Cell Cycle Control |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 03 May 2021 05:00 |
| Last Modified: | 03 May 2021 05:00 |
| URI: | https://pred.uni-regensburg.de/id/eprint/35792 |
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