The CNOT complex contributes to the maintenance of genome stability

Chalabi Hagkarim, Nafiseh ORCID: 0000-0002-7985-3326 (2020). The CNOT complex contributes to the maintenance of genome stability. University of Birmingham. Ph.D.

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Abstract

The yeast CCR4-NOT (CNOT in mammals) complex is a large (1.0-MDa) and highly conserved multifunctional set of proteins. It is involved in many different aspects of mRNA metabolism, including repression and activation of mRNA initiation, control of mRNA elongation, and deadenylation-dependent mRNA turnover; it also has a role in ubiquitin-protein transferase activity and histone methylation. Some studies have suggested that the yeast complex may be involved in the recognition and repair of DNA damage. To investigate whether similar properties are attributable to the mammalian complex we have examined the effects of inactivation of the complex on various aspects of the DNA damage response. Inactivation was achieved by depletion of CNOT1, the largest of the CNOT proteins, which forms a scaffold to the complex. Ablation of CNOT1 expression disrupts cell cycle progression through S and G2/M phases, which subsequently arrests the cell cycle in G1, with markedly elevated levels of cyclin E, p27 and p21. At later times, the cells appear to senesce and /or undergo autophagy. As expected, depletion of CNOT1 affects global transcription and can lead to transcription-dependent replication stress and R-loop formation. CNOT1 depletion can also affect DNA replication by reducing dNTP synthesis. Activity of the RNase H2 complex decreases following loss of CNOT1, which increases the sensitivity of genomic DNA to alkaline lysis due to an increase in embedded ribonucleotides. In addition, depletion of CNOT1 results in DNA damage as seen by comet assay, and formation of micronuclei. This is accompanied by activation of Chk2 in the absence of extraneous DNA damage. In this study, we have demonstrated that the CNOT complex contributes to the maintenance of genome stability and to the response to DNA damage.

Type of Work:

Thesis (Doctorates > Ph.D.)

Award Type:

Doctorates > Ph.D.

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