A function for ATR kinase in the regulation of cytokinetic abscission

Abscission, the final stage of cytokinesis, separates the two daughter cells through cutting of the intercellular bridge. This occurs on either side of a proteinaceous, microtubule-dense structure known as the midbody. Abscission is mediated by the ESCRT proteins that are recruited sequentially to the midbody. These include the ESCRT-III complex which is composed of eight families of ‘CHMP’ proteins that are responsible for a series of membrane remodelling events that ultimately results in abscission. These ESCRT-III filaments in turn interact with a hexameric AAA ATPase VPS4. VPS4 is a ‘remodelling’ complex that continuously remodels ESCRT-III filaments by exchanging CHMP subunits for those with geometries that favour membrane deformation and ultimately constrict the plasma membrane. The ESCRT- III subunit CHMP4B is the main positive regulator of the ESCRT-III filament architecture, required for abscission. An isoform of CHMP4B, termed CHMP4C, negatively regulates abscission by preventing the formation of ESCRT-III filaments of the correct architecture. Abscission timing is regulated by Aurora B kinase which localises to the cytokinetic midbody to prevent premature abscission in the presence of chromatin between the separating daughter cells, which can result in binucleate cells and/or DNA damage. This so-called ‘abscission checkpoint’ controls abscission timing via the ANCHR protein which ‘anchors’ the VPS4 protein at the midbody thereby preventing VPS4 from catalysing the ESCRT-III remodelling required for abscission at the final ‘cut’ site until chromatin has been removed from the intercellular bridge I have shown that in each cell cycle, irrespective of any chromatin bridges, ATR localises to the cytokinetic midbody specifically during late cytokinesis. Furthermore, ATR at the midbody corresponds to its activated phosphorylated form (ATR-T1989p). Consistent with previous genetic studies using a chicken Atr conditional null DT40 cell line, chemical inhibition of ATR activity results in premature abscission and cytokinetic defects. Interestingly, ATR-T1989p midbody localisation is dependent upon the negative regulator CHMP4C, and ATR kinase activity is required to impede the recruitment of CHMP4B, as well as promoting the recruitment of the abscission checkpoint regulator ANCHR to the midbody. Together, this data supports a role for ATR in regulating the correct timing of abscission which is separate from its canonical role in the replication stress response.

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Research Ireland

Publisher

University of Galway

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CC BY-NC-ND

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University of Galway Theses (PhD Theses)