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Quantitative Immunofluorescence Assay to Measure the Variation in Protein Levels at Centrosomes
Published on: December 20, 2014
Abnormal centrosomal structure and duplication in Cep135-deficient vertebrate cells
Burcu Inanç1, Monika Pütz, Pierce Lalor
1Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland.
Molecular Biology of the Cell
|July 19, 2013
Summary
The protein Cep135 is crucial for maintaining centriole structural integrity in vertebrate cells. Its absence leads to atypical centrioles and increased centrosome amplification during S-phase delay.
Area of Science:
- Cell Biology
- Molecular Biology
- Structural Biology
Background:
- Centrosomes organize microtubules and contain centrioles, essential for cell division.
- The protein Cep135 localizes to the cartwheel structure during centriole assembly.
- The precise function of Cep135 in vertebrate centriole biogenesis remains unclear.
Purpose of the Study:
- To investigate the role of Cep135 in vertebrate centriole function and cell cycle regulation.
- To analyze the structural and functional consequences of Cep135 disruption in DT40 cells.
Main Methods:
- Gene disruption of Cep135 in DT40 chicken B-cell line.
- Analysis of centrosome composition and function.
- Electron microscopy to examine centriole ultrastructure.
- Hydroxyurea treatment to induce S-phase delay and assess centrosome amplification.
Main Results:
- Cep135-deficient cells exhibit normal viability but reduced centriole numbers and increased monopolar spindles.
- Electron microscopy revealed atypical lumen structures in Cep135-deficient centrioles.
- Centrosome amplification significantly increased in Cep135-deficient cells upon hydroxyurea treatment.
Conclusions:
- Cep135 is essential for maintaining centriole structural integrity in proliferating vertebrate cells.
- Cep135 acts to limit centrosome reduplication during S-phase arrest.
- This study elucidates a novel role for Cep135 in regulating centrosome numbers and centriole structure.
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