When a cell divides in a process known as mitosis, its duplicated chromosomes need to be separated and evenly distributed into the newly created daughter cells. Despite the importance of mitosis, many of its details still remain unclear, which has hampered efforts to develop treatments for mitotic abnormalities.
A new study reported today by Osaka University researchers in Nature Cell Biology has revealed details about the protein complexes that congregate at the centromeres—sites near the centers of chromosomes. These protein complexes, called kinetochores, act as anchors that help other cellular organizers redistribute chromosomes within the cell.
For their study, the team focused on the different components that form that bind to kinetochores. One such group is the CCAN proteins, which are present at the centromere throughout the cell cycle and act as a binding site for other microtubule-associating proteins only when cell division occurs. The researchers show that a subset of the CCAN proteins that form the CENP-T pathway is pivotal for successful cell division via its binding to a protein complex called the Ndc80 complex, together enabling microtubules to attach to chromosomes.
"We chiefly investigated whether the CENP-T pathway or the roughly similar CENP-C pathway is essential for mitotic progression by selectively deleting parts of these proteins that bind to the Ndc80 complex," says corresponding author Tatsuo Fukagawa. "CENP-T mutants with an absence of domains for binding to Ndc80, but not similar CENP-C mutants, revealed a failure of chromosomes to segregate, preventing cells from dividing and ultimately leading to cell death."
The team next constructed chimeric constructs consisting partly of CENP-T and partly of CENP-C. This validated the finding that CENP-T and the molecules that bind to it are vital for mitosis. They also revealed that phosphorylation plays a vital role in regulating binding between the Ndc80 complex and CENP-T, and they obtained additional direct evidence for their findings by measuring the pulling force exerted by microtubules in the mitotic spindle.
"Our work overturns an earlier consensus by showing that it is CENP-T, not CENP-C, that acts via the Ndc80 complex for successful cell division by ensuring accurate and timely chromosome segregation," says lead author Masatoshi Hara. "The findings could lead to therapeutic options for treating diseases involving dysfunction in the kinetochores and mitotic progression, including cancer."
Image: During mitosis, spindle microtubules capture the kinetochore formed in the centromere. After mitosis, chromosomes are divided into daughter cells. Image courtesy of Osaka University.