Revealing the impact of centromeric DNA on centromere formation, integrity and function – CENtegrity

The preservation of centromere function/integrity is a crucial challenge for cells in order to maintain a correct chromosome karyotype following cell division. Indeed, defects in centromere formation or function lead to numerical (aneuploidy) and structural chromosome alterations, the latter often present at centromeric regions. These alterations are common features of cancer cells and developmental disorders. Centromeres are a complex DNA/protein structure that links chromosomes to the spindle microtubules. In most species, they contain a specific chromatin enriched for the histone H3-variant CENP-A and repetitive DNA, known in human as satellite DNA and bound by CENP-B. Despite recent progress in the identification of the epigenetic processes required for centromere specification dependent on CENP-A, the role of repetitive DNA and CENP-B in centromere structure, formation and function is unclear. The objectives of this proposal are to provide novel information on how centromere position and integrity is preserved and to determine the role of repetitive centromeric DNA and its architecture. My team will combine our recently developed genome editing technique to achieve rapid protein degradation with state of the art microscopy and high-throughput analyses to assess how centromeric DNA/CENP-B counteract whole chromosome-aneuploidy. We will dissect the molecular mechanisms required for de novo centromere specification in real time by combining microscopy, proteomics and genome-wide approaches following removal of the centromere epigenetic mark CENP-A. Finally, we will ascertain how genetic or physical manipulation of the centromere architecture affects chromosome integrity using a combination of cell biology, biochemistry and single molecule technique. This proposal will convey crucial insight into the role of centromeric DNA and its DNA binding protein in centromere and chromosome biology that will be fundamental to understand the genesis of chromosome instability