Mitochondrial and Molecular Defects in Progeroid Cockayne Syndrome Cells

Ageing is dramatically accelerated in rare genetic disorders like the Cockayne syndrome (CS). Dissecting the molecular defects in these diseases is critical to develop treatments and elucidate processes possibly involved in normal ageing.

CS defects (UV-hypersensitivity, premature ageing, neurodegeneration) have been classically attributed to UV damage as proteins mutated in this disease, CSA or CSB, are implicated in the repair UV-induced DNA damage. Nevertheless, CSA or CSB impairment also results in UV-sensitive syndrome (UVSS), a disease characterized only by photosensitivity, thereby uncoupling the DNA repair defect from precocious ageing/neurodegeneration. Furthermore, not all CS patients are clinically hypersensitive to UV light. In this context, our lab identified alteration of a novel pathway specifically in CS (and not UVSS) fibroblasts, which is possibly determinant for the progeroid phenotype (Chatre et al, 2015 PNAS). Namely, oxidative and nitrosative stress promote overexpression of the HTRA3 protease, which results in degradation of the mitochondrial DNA polymerase POLG1, in turn leading to mitochondrial dysfunction.

We aim to dissect the consequences of POLG1 depletion on mitochondrial DNA replication and organelle function, and assess their rescue with MnTBAP, a scavenger of oxidative/nitrosative stress that we showed is able to revert HTRA3 and mitochondrial defects in CS patient cells (Orphan-Drug-Designation for CS). To overcome the difficulty of assessing molecular alterations in patient-derived cells with different genetic backgrounds, we generate isogenic CS cellular models using CRISPR/Cas9 technology to correct the CSB mutation in CS fibroblasts and introduce this mutation in normal cells. Multiple isogenic cellular models should help understanding the dynamics of CS cellular defects, compatibly with a continuous spectrum of severity in CS patients, the causes of which are still unclear. Preliminary results on CRISPR-edited cells will be presented.