J Vet Sci.  2019 Sep;20(5):e50. 10.4142/jvs.2019.20.e50.

Regulation of porcine endogenous retrovirus by dual LTR1+2 (Long Terminal Region) miRNA in primary porcine kidney cells

Affiliations
  • 1Department of Veterinary Medicine Virology Lab, College of Veterinary Medicine and Research Institute for Veterinary Science, Seoul National University, Seoul 08826, Korea. parkx026@snu.ac.kr
  • 2Department of Veterinary Microbiology and Infectious Diseases, Faculty of Veterinary Medicine, Vietnam National University of Agriculture, Hanoi 100000, Vietnam.
  • 3Research Unit, Green Cross Veterinary Products, Yongin 17066, Korea.
  • 4Department of Veterinary Microbiology, College of Veterinary Medicine and Research Institute for Veterinary Science, Seoul National University, Seoul 08826, Korea. yhp@snu.ac.kr

Abstract

Porcine endogenous retroviruses (PERVs) integrate into germline DNA as proviral genome that enables vertical transmission from parents to their offspring. The provirus usually survives as part of the host genome rather than as an infectious agent, but may become pathogenic if it crosses species barriers. Therefore, replication-competent PERV should be controlled through selective breeding or knockout technologies. Two microRNAs (miRNAs), dual LTR1 and LTR2, were selected to inhibit the expression of PERV in primary porcine kidney cells. The inhibition efficiency of the miRNAs was compared based on their inhibition of different PERV regions, specifically long terminal repeats (LTRs), gag, pol, and env. Gene expression was quantified using real-time polymerase chain reaction and the C-type reverse transcriptase (RT) activity was determined. The messenger RNA (mRNA) expression of the PERV LTR and env regions was determined in HeLa cells co-cultured with primary porcine kidney cells. The mRNA expression of the LTR, gag, pol, and env regions of PERV was dramatically inhibited by dual miRNA from 24 to 144 h after transfection, with the highest inhibition observed for the LTR and pol regions at 120 h. Additionally, the RT activity of PERV in the co-culture experiment of porcine and human cells was reduced by 84.4% at the sixth passage. The dual LTR 1+2 miRNA efficiently silences PERV in primary porcine kidney cells.

Keyword

Porcine endogenous retrovirus; inhibition; miRNA; long terminal region; primary porcine kidney cell

MeSH Terms

Coculture Techniques
DNA
Endogenous Retroviruses*
Gene Expression
Genome
HeLa Cells
Humans
Kidney*
MicroRNAs*
Parents
Proviruses
Real-Time Polymerase Chain Reaction
RNA, Messenger
RNA-Directed DNA Polymerase
Selective Breeding
Terminal Repeat Sequences
Transfection
DNA
MicroRNAs
RNA, Messenger
RNA-Directed DNA Polymerase
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