Efficient conditional mutation of the vertebrate CENP-C gene

¹Cancer Research Campaign Chromosome Molecular Biology Group, Biochemistry Department, Oxford University, South Parks Road, Oxford OX1 3QU, UK. tfukagaw@bioch.ox.ac.uk

|March 21, 1998

Summary

We created a conditional CENP-C mutant cell line. This mutant arrests at the metaphase/anaphase transition and undergoes apoptosis, revealing CENP-C

Area of Science:

Background:

Centromere proteins are crucial for chromosome segregation during cell division.
CENP-C is a key component of the centromere, but its precise function in anaphase remains incompletely understood.
Understanding CENP-C's role is vital for comprehending genome stability.

Purpose of the Study:

To investigate the essential function of CENP-C during cell division.
To develop a novel system for studying conditional loss-of-function mutants of essential proteins.
To elucidate the role of CENP-C in anaphase initiation and chromosome movement.

Main Methods:

Gene targeting was employed in the DT40 cell line to create a conditional CENP-C mutant.
A fusion protein between CENP-C and a mouse steroid receptor was expressed.
Cells were cultured under restrictive conditions to induce CENP-C loss-of-function.

Main Results:

The engineered cells exhibited cell cycle arrest at the metaphase/anaphase junction under restrictive conditions.
A delay of approximately 2.5 hours preceded cell death via apoptosis.
These findings strongly implicate CENP-C in anaphase chromosome movement or signaling.

Conclusions:

CENP-C is essential for proper anaphase progression and chromosome segregation.
The developed conditional mutant system is a versatile tool for studying essential proteins.
This research provides critical insights into the molecular mechanisms of chromosome segregation and cell cycle control.

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