Target identification of microRNA repressed by enterovirus 71 infection

• Main Authors: Li Ting Kao, 高儷庭
• Other Authors: J. T. Horng
• Format: Others
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/91796307505030868005

碩士 === 長庚大學 === 生物醫學研究所 === 99 === MicroRNAs are a group of small RNAs (18–24 nucleotides) that inhibit gene expression by binding to specific 3’untranslated region (3’UTR) of mRNA. microRNAs plays an important role in virus replication, such as helping virus replication or inhibition virus replication. Previously, we have observed that four specific microRNAs expression levels were decreased after EV71 infection ten hours in RD cells. I want to identify the target genes of microRNAs and investigate their functions in EV71 replication. First, I used online prediction tools, such as miRanda and Targetscan combined with literature research to obtain four candidate target genes, FKBP1A, PEG10, KPNB1 and SLC2A3. Our result showed that only FKBP1A and PEG10 mRNA expression level were increased after EV71 infection by Q-RT-PCR. I then used Q-RT-PCR, western blot and 3’UTR reporter assay to verify whether candidates are bona fide target genes. Our result showed that FKBP1A is target gene of miR-181-d and PEG10 is target gene of miR-197 and miR-574-5p. In order to understand the biological role of these genes in viral replication, we applied RNA interference to knock down FKBP1A and PEG10. Knockdown of FKBP1A resulted in increased viral protein expression whereas knockdown of PEG10 resulted in decreased viral protein expression. FKBP1A and PEG10 may thus regulate EV71 replication but the exact mechanisms need further investigation.