The Inducible Expression Of MicroRNAs In HeLa Cells Infected By CVB3 And The Regulation Of Novel MicroRNAs On Viral Infection

Objective:It has been known that RNA interference(RNAi) is a nucleic acid-based immunity against viral infection.Over the past years,it has become clear that a variety of viruses,first in plants but recently in insect and mammalian viruses as well, encode suppressors of the RNAi pathway.More recent findings have revealed that certain viruses encode their own microRNAs(miRNAs) or miRNA-like RNA molecules,which are processed by the mammalian RNAi machinery.Furthermore, host-encoded miRNAs have been shown to both silence and enhance intracellular levels of viral RNAs.These findings suggest that interactions between the RNAi pathway including miRNA pathway and viral genomes can profoundly affect the outcomes of the viral life cycles and contribute to the pathogenic signatures of the infectious agents.So,we examined the miRNAs which may regulate both the viral life cycle of Coxsackievirus type B3(CVB3) and the interaction between viruses and their hosts.Methods:The miRNA cloning was used to clone the novel miRNAs from HeLa cells infectecd by CVB3.The genomewide expression profile of miRNAs in HeLa cells infected by CVB3 was detected by miRNA microarray.And on the base of these results,antisense oligonucleotides and a kind of over expression system have been employed to find the real roles of some candidate miRNAs.Results: 1 A series of small RNAs was cloned from HeLa cells infected by CVB3 through miRNA cloning.2 Through Northern blotting and bioinformatic methods which were used to predict the precursor of miRNA,four novel miRNAs related with CVB3 infection were identified.3 We reported the genomewide expression profiling of miRNAs in HeLa cell infected by CVB3 by using a microarray containing 245 human miRNA oligonucleotide probes.This approach allowed us to find some miRNAs which were significantly different in miRNome expression between HeLa cells infected by CVB3 and control HeLa cells.Among these miRNAs,five are up-regulated and three are down-regulated.4 The study about the function of miRv-2 and miRv-4 demonstrated that the two novel miRNAs can slow down the cytopathic effect(CPE) induced by CVB3.Conclusion:The miRNA cloning can be used to find novel miRNAs,and the miRNA microarray can be used to study the genomewide expression profiling of miRNAs. After CVB3 had infected HeLa cells,not only the expression of identified miRNAs was changed,but also novel miRNAs were induced to appear.And the following investigation showed two novel miRNAs——miRv-2 and miRv-4 can inhibit the CPE of CVB3.These results provided the proofs that these miRNAs were associated with CVB3 infection for the first time.