Investigation into the Role of ARMT1 in Oestrogen Receptor Positive Breast Cancer
Breast cancer is the most common malignancy in women and accounts for more than 400,000 deaths per year worldwide. Oestrogen receptor alpha -positive (ER+) tumours account for approximately 80% of all breast tumours. A breast cancer susceptibility locus at 6q25.1, identified by the use of genome wide association studies (GWAS), is located directly upstream of the ESR1 gene that encodes the oestrogen receptor (Zheng et al., 2009). Three genes that are co-expressed with the ESR1 gene - ARMT1, CCDC170 and RMND1 - lie within this breast cancer susceptibility locus (Dunbier et al., 2011). Recent studies have shown a link between ARMT1 and the susceptibility and progression of ER+ breast tumours Yamamoto-Ibusuki et al., 2015). To investigate the function of ARMT1, MCF-7 cells with ARMT1 knockdown by stably transfected shRNA, and ARMT1 over expression by stable transfection with pDHA-neo-ARMT1 were obtained. After validation of these clones by qPCR and western blotting techniques, doubling time analysis showed no difference between either the ARMT1 knockdown and over expression clones and their respective controls, suggesting there is not a direct link between ARMT1 and proliferation rate. Similarly, no difference was seen between the knockdown clones and their controls when senescence-associated beta-galactosidase activity was assessed, before and after the cells were exposed to DNA damage and stress, suggesting ARMT1 levels do not affect senescence induction in these clones. Again no difference was observed when the clones were analysed for anchorage independent growth, indicating ARMT1 is not involved with the evading of growth suppressors. Cell wounding assays produced a significant increase in migration in the ARMT1 overexpression clones compared to a control, however no significant differences were seen between the ARMT1 knockdown clones and controls. DNA damage response was analysed via clonogenic growth of cells after exposure to doxorubicin and UV. Small differences in survival between the knockdown and over expression clones and their respective controls were observed, but the magnitude of these changes was inconsistent with previous findings (Perry et al., 2015). Interestingly, expression analysis with qPCR revealed reduced expression of ESR1 in ARMT1 knockdown clones. This expression pattern is mirrored by CCDC170, RMND1, and the oestrogen responsive genes TFF1 and GREB. Cumulatively these results suggest ARMT1 may have a role in regulating ESR1 expression. Further analysis is needed however to investigate the mechanism through which this occurs.
Advisor: Dunbier, Anita
Degree Name: Master of Science
Degree Discipline: Biochemistry
Publisher: University of Otago
Keywords: ARMT1; breast; cancer; oestrogen; receptor
Research Type: Thesis