Comprehensive Analysis of Human Cytomegalovirus MicroRNA Expression during Lytic and Quiescent InfectionReportar como inadecuado




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Background

Human cytomegalovirus HCMV encodes microRNAs miRNAs that function as post-transcriptional regulators of gene expression during lytic infection in permissive cells. Some miRNAs have been shown to suppress virus replication, which could help HCMV to establish or maintain latent infection. However, HCMV miRNA expression has not been comprehensively examined and compared using cell culture systems representing permissive lytic and semi-permissive vs. non-permissive latent-like infection.

Methods

Viral miRNAs levels and expression kinetics during HCMV infection were determined by miRNA-specific stem-loop RT-PCR. HCMV infected THP-1 non-permissive, differentiated THP-1 d-THP-1, semi-permissive and human embryo lung fibroblasts HELs, fully-permissive were examined. The impact of selected miRNAs on HCMV infection gene expression, genome replication and virus release was determined by Western blotting, RT-PCR, qPCR, and plaque assay.

Results

Abundant expression of 15 HCMV miRNAs was observed during lytic infection in HELs; highest peak inductions 11- to 1502-fold occurred at 48 hpi. In d-THP-1s, fourteen mRNAs were detected with moderate induction 3- to 288-fold, but kinetics of expression was generally delayed for 24 h relative to HELs. In contrast, only three miRNAs were induced to low levels 3- to 4-fold during quiescent infection in THP-1s. Interestingly, miR-UL70-3p was poorly induced in HEL 1.5-fold, moderately in THP-1s 4-fold, and strongly 58-fold in d-THP-1s, suggesting a potentially specific role for miR-UL70-3p in THP-1s and d-THP-1s. MiR-US33 -UL22A and -UL70 were further evaluated for their impact on HCMV replication in HELs. Ectopic expression of miR-UL22A and miR-UL70 did not affect HCMV replication in HELs, whereas miR-US33 inhibited HCMV replication and reduced levels of HCMV US29 mRNA, confirming that US29 is a target of miR-US33.

Conclusions

Viral miRNA expression kinetics differs between permissive, semi-permissive and quiescent infections, and miR-US33 down-regulates HCMV replication. These results suggest that miR-US33 may function to impair entry into lytic replication and hence promote establishment of latency.



Autor: Zhang-Zhou Shen, Xing Pan, Ling-Feng Miao, Han-Qing Ye, Stéphane Chavanas, Christian Davrinche, Michael McVoy, Min-Hua Luo

Fuente: http://plos.srce.hr/



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