Master Regulation of Centromere Function by the Highly Conserved Mis18 Complex

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The cells in our body duplicate their genomes every time they divide. In order to make sure that the process of cell division proceeds to completion without any glitches, the genomic DNA packaged in chromosomes following duplication in the parental cell must be equally and accurately segregated between two daughter cells. This means that every time a cell divides, the newly formed daughter cells will inherit the same amount of genetic material that the parental cell initially harboured. Specialised structures known as 'kinetochores' assemble on specific sites on chromosomes known as 'centromeres' to ensure that chromosome segregation into daughter cells is always accurate and error-free. Centromeres are crucial for genome stability, as kinetochores that assemble on them must attach themselves to thread-like structures known as microtubules that will then pull each duplicated chromosome towards one and only one daughter cell during cell division. Centromere dysfunction and consequent kinetochore defects can lead to chromosome mis-segregation, which has been frequently observed in a variety of cancers, and in genetic diseases such as Down's syndrome.In most organisms, kinetochore-microtubule attachment happens at only one site on each individual chromosome. Each chromosome therefore must have only one centromere on it. While much of the chromosomal DNA in cells is tightly wrapped around conventional 'histone' proteins, centromeric DNA is unique in that it is wrapped around a specialised centromere-specific histone protein known as CENP-A. It is CENP-A, rather than the underlying DNA sequence itself, that specifies the site of centromere assembly on each chromosome. CENP-A is known to be essential for centromere function and kinetochore formation: mutations in this protein have been shown to adversely affect chromosome segregation. How CENP-A recognizes centromeric DNA and marks functional centromeres, has remained a mystery for long.The assembly of CENP-A at centromeres is dependent on the highly conserved centromere protein Mis18, and the CENP-A specific chaperone protein HJURP. The sequence of events that regulates the timing of CENP-A loading is first initiated by Mis18, which recruits HJURP, that in turn then deposits CENP-A at centromeres. Mis18 thus master-regulates CENP-A assembly at centromeres, and is the most fundamental factor required for centromere function. Mis18 was first identified in fission yeast in 2004, as a gene which when mutated led to chromosome mis-segregation. Subsequent studies in human cells have shown that Mis18 function is highly conserved. Not surprisingly, Mis18 has been found to be mis-regulated in a variety of human cancers. How Mis18 initially recognizes centromeres to subsequently effect CENP-A assembly, however, remains unclear. The research proposed in this application is broadly aimed at elucidating how Mis18 and its associated proteins, together regulate CENP-A assembly and centromere function. The insights gained from our research will be key to building a more complete understanding of how cells faithfully segregate their DNA during cell division, through the course of evolution.