Research

Accurate distribution of chromosomes to the daughter cells during cell division requires selective stabilisation of chromosome-microtubule attachments, capable of supporting chromosome bi-orientation (where sister chromatids are attached to microtubules emanating from opposite spindle poles) and maintaining sister-chromatid cohesion until all sister-chromatids achieve bipolar attachment. Two chromosomal sites work at the heart of these processes: the centromere, defined by the enrichment of CENP-A (a Histone H3 variant) nucleosomes, and the inner centromere, which lies between the two sister-chromatids. The centromere acts as an assembly site for the kinetochore, where microtubules attach. Unlike canonical chromatin, CENP-A nucleosome undergo DNA replication-mediated dilution due to the distribution of existing CENP-A to the newly made DNA strand during each round of the cell cycle. To preserve centromere identity and hence  to maintain the microtubule attachment site at the right place, CENP-A levels must be replenished during each cell cycle round. The inner centromere acts as a signalling/regulatory hub, recruiting factors that regulate kinetochore-microtubule attachments and control timely sister-chromatid separation. 

We have a good understanding of the mechanisms controlling the assembly and function of the kinetochore. However, structural and molecular bases for the mechanisms underlying the maintenance of centromere identity and the establishment of the centromere-associated regulatory interaction network are just emerging.  The overarching goal of our current work is to obtain high-resolution, mechanistic understanding of centromere/inner centromere assembly and their function in ensuring accurate segregation of chromosomes during cell division.  This is crucial as defective chromosome segregation often results in aneuploidy, a chromosomal numerical aberration implicated in miscarriages, infertility, birth defects and several human cancers. 

Exploiting our experience in integrating structure-function approaches (X-ray crystallography, cryo electron microscopy, Crosslinking/Mass Spectrometry, biochemical/biophysical methods with human cell-line based functional assays) to study chromosome segregation, we currently aim to address three important questions:

1.    How is the inner centromere signalling/regulatory platform established?

2.    How does the inner centromere recruit enzymatic activities to ensure accurate chromosome segregation?

3.    How is the centromere identity preserved through generations of cell division?

Recently, we discovered that the Chromosomal Passenger Complex (CPC), which is a major centromere associated  regulator of chromosome segregation has an intrinsic nucleosome binding activity essential for its chromosome  association and function (Abad et al., 2019, J Cell Biol). We have also characterised the molecular basis for how CPC  interacts with Sgo1, a key regulator of sister-chromatid cohesion (Abad et al., 2021, bioRXiv).

Our ongoing and future work will provide unprecedented details of centromere-mediated control of chromosome  segregation and allow us to build a comprehensive mechanistic model for error-free chromosome segregation, a process  that has been fascinating researchers for more than a century. 

Image

A.    Overview of proposed pathways responsible for the centromere localization of the Chromosomal Passenger Complex (CPC; Borealin, Survivin, INCENP and Aurora B), a master regulator of chromosome segregation. Two histone phosphorylations, Histone H3 Thr3 (H3T3p) and Histone H2A Thr120 (H2AT20p), mediated by Haspin and Bub1 kinases respectively, recruit CPC to the inner centromere. CPC binds H3T3p directly via Survivin and H2AT120p indirectly via Sgo1.

B.    Molecular basis for CPC-Sgo1 interaction: CPC-Sgo1 binding requires physical recognition of Histone H3 like N-terminal tail of Sgo1 by Survivin. Disrupting this interaction perturbs CPC centromere association and leads to chromosome missegregation.

JP lab, Gene Center Munich