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Cloning, localization, and axonemal function of Tetrahymena centrin
Charles Guerra1, Yuuko Wada, Vagn Leick
1Department of Anatomy and Structural Biology, Albert Einstein College of Medicine, Bronx, New York 10461, USA.
Molecular Biology of the Cell
|January 17, 2003
Summary
Centrin, a calcium-binding protein, regulates inner arm dynein activity in Tetrahymena cilia. This protein controls microtubule sliding, impacting ciliary functions like reversal and chemotaxis.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Centrin is an EF-hand calcium-binding protein.
- Tetrahymena thermophila possesses multiple EF-hand proteins with distinct localizations.
- Centrin's specific localization within Tetrahymena cilia suggests a role in ciliary function.
Purpose of the Study:
- To clone and characterize centrin from Tetrahymena thermophila.
- To investigate the function of centrin in regulating axonemal dynein activity.
- To explore centrin's role in calcium-mediated ciliary responses.
Main Methods:
- Gene cloning and protein characterization of Tetrahymena centrin.
- Immunofluorescence microscopy to determine centrin localization.
- In vitro microtubule motility assays to assess dynein activity.
- Functional assays using calcium and a calcium-mimicking peptide (CALP1).
Main Results:
- Tetrahymena centrin is a 19.4 kDa protein with high homology to other centrins.
- Centrin localizes to basal bodies, oral apparatus, ciliary rootlets, and inner arm dynein (IAD) in cilia.
- Calcium or CALP1 increased microtubule sliding velocity, an effect abrogated by anti-centrin antibodies.
- This study demonstrates centrin's direct role in regulating IAD activity.
Conclusions:
- Centrin is a key regulatory protein for calcium responses in Tetrahymena cilia.
- Centrin's function is linked to the control of inner arm dynein activity.
- This regulation is crucial for ciliary functions such as ciliary reversal and chemotaxis.