马立克病病毒编码的miR-M11-5p宿主靶基因的筛选与鉴定

doi:10.11843/j.issn.0366-6964.2019.05.013

Abstract

Abstract:

As one of the most important Alphaherpesvirus, Marek's disease virus (MDV) causes immunosuppression and rapid-onset of T-cell lymphoma in its natural host chicken, namely Marek's disease (MD). A large number of viral microRNAs (miRNAs) have been identified in the MDV genomes, which may play critical roles in virus replication, latency, and tumourigenesis. It has been previously confirmed that deletion of miR-M11-5p from the viral genome significantly enhances MDV pathogenicity and oncogenicity, suggesting that it may be a tumor suppressor during MDV tumourigenesis. To further reveal the regulatory mechanism mediated by miR-M11-5p in MDV oncogenesis, we have presently performed a hybrid-PCR to amplify the candidate host target genes using the cDNA derived from chicken embryo fibroblasts (CEF) cellular RNA as template. The PCR products were ligated into the pMD19-T vector and transformed into E. coli JM109 to construct a cDNA library for screening the candidate targets for miR-M4-5p. Based on PCR identification, DNA sequencing and BLAST analysis, a total of 77 candidate genes were obtained, of which 37 binding sites recognized by miR-M11-5p were located in the 3'-UTRs of the mRNA genes. The further analysis of dual luciferase reporter assay (DLRA), miRNA over-expression and RT-qPCR confirmed that three host genes (MAFB, LOC776816 and RFX7) were the final target genes for miR-M11-5p. Our data provide an important foundation for further elucidation of the regulatory mechanism mediated by miR-M11-5p in MDV tumourigenesis.

CLC Number:

S852.659.1

LIU Haoli, TENG Man, LI Huizhen, YU Zuhua, LIU Jingling, DING Ke, ZHANG Gaiping, LUO Jun. Screening and Identification of Host mRNA Targets for the Viral microRNA miR-M11-5p Encoded by Marek's Disease Virus[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2019, 50(5): 1026-1038.

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Screening and Identification of Host mRNA Targets for the Viral microRNA miR-M11-5p Encoded by Marek's Disease Virus

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