Assessing the role of Syntaxin18 in controlling c-myc mRNA expression and growth in human breast cancer cells.

Messenger RNA stability is critical in the control of gene expression. Enzymes capable of cleaving RNA, termed ribonucleases, are gaining appreciation for their role in regulating gene expression. Our lab has discovered a novel protein which has been shown to cleave c-myc mRNA within the coding region determinant (CRD). This protein was tentatively identified as Syntaxin18, a known t-SNARE protein. Recombinant Syntaxin18 has been shown to possess endoribonucleolytic activity against c-myc mRNA in vitro. In order to determine whether Syntaxin18 functions as an endoribonucleae in cells, the effect of altering its expression on the proliferation, levels and stability of c-myc mRNA was examined. The expression of Syntaxin18 in various cell lines and tissues were also compared. Through Western analysis, it was discovered that the expression of Syntaxin18 was similar in all immortalized cancer cell lines examined. However, higher Syntaxin18 expression was observed in cancerous breast tissue when compared to the normal breast tissue. MTT assays revealed that modulation of Syntaxin18 was able to affect the proliferation of MCF-7 cells, and this effect was observed only upon incubation with 17-β-estradiol and insulin. Finally, ribonuclease protection assay analysis showed that modulation of Syntaxin18 may have an effect on the steady-state levels of c-myc and β-actin mRNAs; however, down-regulation of Syntaxin18 did not impact the stability of the c-myc transcript. Based on results obtained from this study, it remains unclear whether Syntaxin18 can function as an endoribonuclease in MCF-7 cells.