Screening and computational analysis of colorectal associated non-synonymous polymorphism in CTNNB1 gene in Pakistani population

Abstract

AbstractBackgroundColorectal cancer (CRC) is categorized by alteration of vital pathways such as β-catenin(CTNNB1) mutations,WNTsignaling activation,tumor protein 53 (TP53) inactivation,BRAF, Adenomatous polyposis coli (APC) inactivation,KRAS, dysregulation of epithelial to mesenchymal transition (EMT) genes,MYCamplification, etc. In the present study an attempt was made to screenCTNNB1gene in colorectal cancer samples from Pakistani population and investigated the association ofCTNNB1gene mutations in the development of colorectal cancer.Methods200 colorectal tumors approximately of male and female patients with sporadic or familial colorectal tumors and normal tissues were included. DNA was extracted and amplified through polymerase chain reaction (PCR) and subjected to exome sequence analysis. Immunohistochemistry was done to study protein expression. Molecular dynamic (MD) simulations of CTNNB1WTand mutant S33F and T41A were performed to evaluate the stability, folding, conformational changes and dynamic behaviors of CTNNB1 protein.ResultsSequence analysis revealed two activating mutations (S33F and T41A) in exon 3 ofCTNNB1gene involving the transition of C.T and A.G at amino acid position 33 and 41 respectively (p.C33T and p.A41G). Immuno-histochemical staining showed the accumulation of β-catenin protein both in cytoplasm as well as in the nuclei of cancer cells when compared with normal tissue. Further molecular modeling, docking and simulation approaches revealed significant conformational changes in the N-terminus region of normal to mutantCTNNB1gene critical for binding with Glycogen synthase kinase 3-B (GSK3) and transducin containing protein1 (TrCp1).ConclusionPresent study on Pakistani population revealed an association of two non-synonymous polymorphisms in theCTNNB1gene with colorectal cancer. These genetic variants led to the accumulation of theCTNNB1, a hallmark of tumor development. Also, analysis of structure to function alterations inCTNNB1gene is crucial in understanding downstream biological events.