بررسي بيان ژن اينوزين تري فسفات پيروفسفوهيدرولاز در سلو لهاي K562

Study of Inosine triphosphate pyrophosphohydrolase gene expression in K562 cells

Objective: The ITPA gene is responsible to remove free deaminated purine nucleotides of ITP, dITP and XTP from nucleotide pool of the cells. It seems that dysfunction in its activity, not only can increas the base substitution mutations frequency but also can works as a contrived factor to creating instability in genetic materials of the cells. There are several reports about the existence of structural and numerical genetic instability in the K562 cell genome. In this research, we examined the expression of ITPA gene as a possible contrived factor in observed genetic instability of this cell line. Materials and Methods: To evaluate the expression of target gene semi-quantitative RT-PCR technique was used. Then to examine the functionality of gene products, its cDNAs were cloned and their sequences were determined. Their proteins products were predicted using available bioinformatics soft wares and the results were compared. Results: The result of structural prediction of second mRNA showed that it has ability to encode a protein which has inability in substrate binding and also in its normal enzymatic activity. With regard to the fact that enzymatic activity of protein is dependent on the dimer formation, the function of hetero-dimer enzyme is changed. Therefore the catalytic activity of ITPase is predicted to be abnormal and it can be considered as a contrived factor for creating genetic instability in K562 cell line. Conclusion: The study of gene expression showed that ITPA gene is expressed in moderate level compared to GAPDH expression as an internal control in K562 cell. Two types of transcripts were detected in this line. One of them was the normal product of splicing process of primary hnRNA, but the second one contained a 51 nucleotides deletion in the mRNA coding region. It seems, this transcript is the product of a rare splicing process in this line.