Identification of MicroRNA Targeting MYC

Abstract

c‐Myc is a transcription factor that regulates numerous processes including cell cycle, apoptosis, cellular differentiation, cellular metabolism and genomic instability. In normal cells, expression of c‐Myc is tightly regulated by external signals such as growth factors and extracellular matrix. However in stressed and cancerous cells, c‐Myc is generally over‐expressed by chromosomal translocation, gene amplification, or stabilization of its mRNA. We hypothesize that MYC is a target of microRNA (miRNA). miRNAs are 20‐22bp short RNAs that regulate gene expression by binding target mRNA 3′‐UTR, leading to mRNA degradation or translational repression. We used a reporter construct carrying c‐Myc binding sites to screen miRNAs targeting MYC. The results demonstrated that miR‐212 (30%), miR‐203 (40%), miR‐33b (50%) and miR‐33a (50%) down‐regulated reporter activity. Next, we cloned the 3′‐UTR of MYC into a reporter vector and performed assays in cell transfected with miRNAs. We found that miR‐33a (60%), miR‐33b (50%), miR‐203 (40%) down‐regulated the expression of the reporter gene located upstream of the MYC 3′‐UTR. Finally, we performed colony formation assay with rat RK3E‐cmyc cells, which constitutively express c‐Myc. Our results showed that miR‐212 (85%), miR‐33a (50%), miR‐33b (60%) and miR‐203 (17%) reduced colony formation. We will continue to investigate whether MYC is an authentic target of these miRNAs.