A novel and frequent del T alteration in a microsatellite sequence of the 3’-UTRCDK2-AP1 gene regulation CDK2-AP1 decreased expression in MSI CRC
Background: We have demonstrated Cyclin Dependent Kinase2 –Associated Protein 1 (CDK2-AP1) expression is significantly decreased in microsatellite unstable (MSI) CRC cell lines. We observed siRNA inhibition of CDK2-AP1 associated with increased colon cell proliferation and decreased apoptosis. Recently, we also observed a novel del T in a microsatellite sequence of CDK2-AP1 associated with its decreased expression in MSI CRC. In this study, we further characterize the frequency and role of this del T associated with MSI CRC in human and knockout mouse models.
Methods: The del T was determine in 98 human CRC samples from 35 CRC cell lines (MSI, 14 and MSS, 21) and 63 CRC tissues (MSI, 31 and MSS, 32). This del T was also determined in intestine tumor tissue from MLH1 knockout mice. The function of del T in the MSI CRC was determined by employing a Lentiviral Vector mediated gene transfer system (LV-pRRL.sin.PPT.hPGK.EGFP). This del T mediating CDK2-AP1 mRNA instability and degradation was also analyzed by RNA folding structure determining.
Result/Discussion: A novel and frequent somatic alteration, del T in the 3’-UTR CDK2-AP1 gene was determined in 21% of the human MSI CRC cell lines and 13% of the human MSI CRC tissues, associated with decreased CDK2-AP1 expression. Interestingly, a similar del T alteration in the 3’-UTR CDK2-AP1 was determined in MSI intestine tumor from MLH1 knockout mice. We identified this novel and frequent del T in the 3’-UTR CDK2-AP1 gene associated function alteration decreased CDK2-AP1 expression in the MSI CRC by employing a Lentiviral Vector mediated gene transfer system. This del T presented mediating CDK2-AP1mRNA instability and degradation. These data shows that the molecular mechanism of CDK2-AP1 decreased expression in MSI deficiency is associated with del T in a poly (T)8 of the 3’-UTR of the CDK2-AP1 gene. In these results we demonstrated a strong association between MSI CRC and decreased or absent CDK2-AP1 expression.