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Developing of isogenic cell lineages with different aleles XPG through transduction with lentiviral vectors

Grant number: 16/05569-0
Support type:Scholarships in Brazil - Scientific Initiation
Effective date (Start): July 01, 2016
Effective date (End): November 30, 2016
Field of knowledge:Biological Sciences - Biochemistry
Principal Investigator:Carlos Frederico Martins Menck
Grantee:Davi Mendes
Home Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil
Associated research grant:14/15982-6 - Consequences of repair deficiencies in damaged genome, AP.TEM

Abstract

DNA is the center of all cells' genetic information and it is constantly exposed toendogenous or exogenous genotoxic agents that can cause damage which eventually lead tomutagenesis or cell death. The most common endogenous agents are reactive oxygen species(ROS), originated mainly in processes such as cellular respiration and inflammation. Theseagents oxidize bases, sugars, phosphates and causing single strand breaks (SSBs) or double(DSBs) in the DNA molecule. During the evolution, cells have developed DNA repairsystems that correct the damage. Dysfunctions in these systems lead to the accumulation ofDNA damage and human syndromes are important for understanding processes such as agingand neurodegeneration, linked mainly to the accumulation of damage in both the nuclear andmitochondrial DNA. Several knowledge gaps exist, lacking models to better investigate thecauses for these processes. The main goal for this project is to develop isogenic lineages fromXPCS1LV cell line, deleted for the gene encoding for the XPG protein, which has a role asendonuclease in the nucleotide excision repair (NER). Xeroderma pigmentosum patientscarrying homozygous mutations in the XPG present high skin cancer incidence, as well asclinical phenotypes may also include neurodegeneration and premature aging. This cell lineswill be transduced with lentiviral vectors carrying the wild-type XPG gene or alleles of XPGmutated genes identified in patients from Cabo Frio, Rio de Janeiro, Brazil and characterizedby Dr. Daniela Soltys. Thus, it is expected to obtain four new cell lines, one of these beingwild-type, two sensitive only to ultraviolet light while the third one will be sensitive tooxidative stress and UV (transduced with a control vector). Thus we expect to obtain morestable and reliable cell models to identify the XPG relationship to repair the damage causedby oxidative stress and if such damage is related with mitochondrial dysfunction.