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Quantitative Immunofluorescence Assay to Measure the Variation in Protein Levels at Centrosomes
Published on: December 20, 2014
Plk4-induced centriole biogenesis in human cells
Julia Kleylein-Sohn1, Jens Westendorf, Mikael Le Clech
1Department of Cell Biology, Max-Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany.
Developmental Cell
|August 8, 2007
Summary
Overexpression of Polo-like kinase 4 (Plk4) causes human cells to generate extra centrioles. This study maps the essential components and stages involved in centriole assembly and elongation.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Centrioles are crucial for cell division and organization.
- Understanding centriole biogenesis is key to deciphering cellular function and dysfunction.
Purpose of the Study:
- To dissect the molecular mechanisms and intermediates of human cell centriole assembly.
- To identify and order the critical protein components involved in procentriole formation.
Main Methods:
- Centrosome amplification induced by Polo-like kinase 4 (Plk4) overexpression.
- siRNA screening to identify essential assembly factors.
- Immunoelectron microscopy for protein localization.
Main Results:
- Identified Plk4, hSas-6, CPAP, Cep135, gamma-tubulin, and CP110 as key players in a staged assembly pathway.
- hSas-6 transiently associates with nascent procentrioles.
- Cep135 and CPAP form a stable core structure.
- CP110 acts as a cap during centriole elongation via tubulin insertion.
Conclusions:
- A comprehensive model for human cell centriole biogenesis is presented.
- The study elucidates the sequential recruitment and function of proteins in building new centrioles.
- Elongation mechanism involves tubulin insertion beneath a CP110 cap.
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