Role of the breast cancer susceptibility gene BRCA1 in radio-resistance
© Current Science Ltd 2000
Published: 12 March 2000
Evidence that BRCA1 is involved in DNA repair stems from its interaction with proteins like RAD51, RAD50, etc. We hypothesised that overexpression or antisense inhibition of the mouse homologue Brca1 in an ovarian cancer cell line could result in changes of sensitivity to γ-radiation.
Mouse ovarian cancer cell line ID-8 was transfected or infected with Neo-resistant vector or retrovirus expressing full-length Brca1 in the sense or antisense orientation. G418-resistant colonies were isolated (two sense, S2 and S13, and three antisense, AS1, AS2 and AS3) and levels of Brca1 were quantified by real time quantitative RT-PCR. Cells were incubated in 96-well culture plates at 37°C for 24 hours. They were irradiated (0, 1, 3, 5 and 10 Gy), incubated at 37°C for 24 hours, washed, and postincubated at 37°C in fresh medium for 48 hours, after which cell density was measured using a colorimetric assay with sulforhodamine B.
Colonies S2 and S13 respectively expressed very low and high levels of Brca1 transgene, and expression of transgene in colony AS2 was 12- and 60-times higher than in colonies AS1 and AS3 respectively. Overexpression of Brca1 resulted in enhanced resistance to irradiation (1.2 to 1.5-fold), whereas inhibition of expression of Brca1 resulted in a very slight increase in sensitivity. Increased radio-resistance induced by overexpression of Brca1 is consistent with other studies, and supports a role for BRCA1 in DNA damage repair. On the other hand, the discrepancy between our results obtained with inhibition expression of Brca1 and data in the literature (ionising radiation hypersensitivity in human cancer cells containing mutated BRCA1 and mouse embryonic stem cells carrying a Brca1 null mutation) is unexpected. We need to investigate further the inhibition of expression of protein Brca1 and above all its functional properties, analysing expression profiles of BRCA1 downstream target genes like p21, GADD45, c-myc etc