Morena Pinheiro

Morena

Pinheiro

MMRES student at IRBB and CRG
Class 2019-2020

Generating CRISPR-Cas9 knockouts in DNA repair genes to create DNA damage maps upon genotoxic agents’ treatment

Summary

Cancer cells prioritize survival and proliferation, overriding signals to properly repair DNA damage or even repair it at all. This can lead to increased mutagenesis, aggressiveness, and resistance to drug treatments. Cancerous mutations are the result of the interplay between DNA damage and repair, so understanding this crosstalk is paramount to design new therapeutic strategies. To understand mutation mechanisms in cancer cells more in detail, we focus on its origin: DNA damage and the cell’s ability to mend it via DNA repair pathways. In particular, a valuable tool to understand mutagenesis is analysing mutation profiles in knockouts of DNA repair genes. We have used the CRISPR-Cas9 technique to design a DNA repair knockout library. After validation, the cells can be exposed to genotoxic agents and use a custom DNA damage library to pinpoint the lesions. Similar to what is seen for mutations, the distribution of the damage is not expected to be homogeneous, rather distributed depending on epigenetic factors. In this project, I focused on the construction of a XPF nuclease knockout. The XPF gene is involved in nucleotide excision repair, Fanconi Anemia and homologous recombination pathways. Its mutation leaves cells unable to repair ultraviolet light DNA damage, among other damaging events, causing melanomas in patients exposed to sunlight. Therefore, these findings can be key to understand the role of mutagenesis in cancer risk, progression and therapeutic response.

Major project supervisor

Núria López-Bigas
IRB Barcelona

Minor project supervisor

Maria Pia Cosma
CRG