A novel ATM-Chk2-INCENP pathway delays abscission to prevent chromosome breakage in cytokinesis with chromatin bridges
Session type: E-poster/poster
Theme: Cancer research and COVID-19
Chromatin bridges are strands of missegregated chromatin connecting the anaphase poles or daughter nuclei and have been linked to tumourigenesis. In response to chromatin bridges in cytokinesis, cells delay abscission, the severing of the narrow cytoplasmic canal that connects the two daughter cells, to prevent chromatin breakage or tetraploidization by regression of the cleavage furrow that are associated with genomic instability and cancer predisposition. In mammalian cells, this abscission-delay is called “the abscission checkpoint” and is dependent on the localization of the Chromosomal Passenger Complex (CPC) at the midbody. The CPC comprises the catalytic subunit Aurora B kinase, the scaffolding protein INCENP and the non-enzymatic subunits Survivin and Borealin; however, the molecular mechanisms that signal chromatin bridges to the CPC have not been previously identified.
To investigate this, we used siRNA depletion-replacement and in vitro reconstitution experiments, in combination with confocal microscopy, live cell imaging and biochemistry techniques.
We show that inhibition of the DNA damage kinases ATM or Chk2 impairs CPC-localization to the midbody and correlates with premature abscission and chromatin breakage in cytokinesis with trapped chromatin in human carcinoma cell lines. ATM phosphorylates Chk2-threonine 68 (T68) to activate Chk2 at the midbody. In turn, active Chk2 phosphorylates INCENP at a newly identified site in cytokinesis, to promote CPC-localization to the midbody and delay abscission. Expression of a phosphomimetic mutant INCENP at the Chk2 site, but not the wild-type protein, rescues CPC-midbody-localization and prevents chromatin breakage in Chk2-deficient or ATM-deficient cells. In contrast, a non-phosphorylatable mutant INCENP does not localize to the midbody; furthermore, expression of this non-phosphorylatable INCENP accelerates abscission, promotes chromatin breakage and reduces cell proliferation compared with the wild-type protein.
These results identify a novel signaling pathway that prevents chromosome breakage in cytokinesis.
These results describe a novel mechanism that could protect against tumourigenesis and support targeting abscission-control proteins as a potential therapeutic strategy to kill proliferating tumour cells.