Skip to main content
NCBI home page
Springer Nature - PMC COVID-19 Collection logoLink to Springer Nature - PMC COVID-19 Collection
. 2008 Jan 13;22(3):199–206. doi: 10.1007/s12250-007-0022-3

P19 of tomato bushy stunt virus suppresses RNA silencing induced by short hairpin RNA in mammal cells

Chen Wei-xian 1, Chen Juan 1, Zhang Zhen-zhen 1, Huang Ai-long 1,✉
PMCID: PMC7090650

Abstract

To counteract the immune system in parasitic hosts, some viruses encode proteins to suppress the RNA interference (RNAi) effect. In this report, we established two RNAi systems to be easily observed with strong and obvious effect. The function of the P19 of tomato bushy stunt virus, which suppresses RNAi in mammal cells, was then studied using these two systems. Short hairpin RNAs targeting green fluorescence protein (pshRNA-GFP) and firefly luciferase (pshRNA-luc) were designed and inserted into a eukaryotic transcriptional vector pTZU6+1, respectively. The shRNA expressing vectors were co-transfected with plasmids containing the target gene with or without P19. The GFP expression level was assayed by fluorescence microscopy, Western blotting and RT-PCR. The luciferase expression level was analyzed by the dual-luciferase assay system. pshRNA designed in this study down-regulated the target gene specifically and efficiently, with a decrease of expression of both genes of about 70%, respectively. When P19 was introduced into the RNAi systems, the expression of both GFP and the luciferase were mostly recovered compared with the control groups. The RNAi systems of GFP and luciferase were constructed successfully, demonstrating that P19 of tomato bushy stunt virus has the ability to counteract the RNAi effect induced by shRNA in mammal cells.

Key words: RNA interference (RNAi), Suppressor, P19, Tomato bushy stunt virus

Footnotes

Foundation items: National Natural Science Foundation of China (No.30500428); Chongqing Science and Technology Commission (2006BA5021)

References

  • 1.Bennasser Y., Le S. Y., Benkirane M., et al. Evidence that HIV-1 encodes an siRNA and a suppressor of RNA silencing. Immunity. 2005;22(5):607–619. doi: 10.1016/j.immuni.2005.03.010. [DOI] [PubMed] [Google Scholar]
  • 2.Carrington J. C., Ambros V. Role of microRNAs in plant and animal development. Science. 2003;301:336–338. doi: 10.1126/science.1085242. [DOI] [PubMed] [Google Scholar]
  • 3.Charles H. L., Patrice D., Khalil A., et al. A cellular microRNA mediates antiviral defense in human cells. Science. 2005;308(22):557–560. doi: 10.1126/science.1108784. [DOI] [PubMed] [Google Scholar]
  • 4.Daniel S., Attila M., Alessandra L., et al. A viral protein suppresses RNA silencing and binds silencing-generated, 21-to 25-nucleotide double-stranded RNAs. EMBO J. 2002;21(12):3070–3080. doi: 10.1093/emboj/cdf312. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 5.Donald K. Breakthrough of the Year. Science. 2002;298:2283. doi: 10.1126/science.298.5602.2283. [DOI] [PubMed] [Google Scholar]
  • 6.Elbashir S. M., Harborth J., Lendeckel W., et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in culture mammalian cells. Nature. 2001;411:494–498. doi: 10.1038/35078107. [DOI] [PubMed] [Google Scholar]
  • 7.Hannon G. J. RNA interference. Nature. 2002;418:244–251. doi: 10.1038/418244a. [DOI] [PubMed] [Google Scholar]
  • 8.Kasschau K. D., Xie Z., Allen E., et al. P1/HC-Pro, a viral suppressor of RNA silencing, interferes with Arabidopsis development and miRNA function. Dev Cell. 2003;4(2):205–217. doi: 10.1016/S1534-5807(03)00025-X. [DOI] [PubMed] [Google Scholar]
  • 9.Li H. W., Li W. X., Ding S. W. Induction and suppression of RNA silencing by an animal virus. Science. 2002;296:1319–1321. doi: 10.1126/science.1070948. [DOI] [PubMed] [Google Scholar]
  • 10.Li W. X., Li H. W., Lu R., et al. Interferon antagonist proteins of influenza and vaccinia viruses are suppressors of RNA silencing. Proc Natl Acad Sci USA. 2004;101(5):1350–1355. doi: 10.1073/pnas.0308308100. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Matthew N.P., Lena E., Jan K., et al. A pancreatic islet-specific microRNA regulates insulin secretion. Nature. 2004;432:226–230. doi: 10.1038/nature03076. [DOI] [PubMed] [Google Scholar]
  • 12.Mallory A.C., Reinhart B. J., Bartel D., et al. A viral suppressor of RNA silencing differentially regulates the accumulation of short interfering RNAs and micro-RNAs in tobacco. Proc Natl Acad Sci USA. 2002;99(23):15225–15233. doi: 10.1073/pnas.232434999. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Patrice D., Charles H. L., Eneida A. P., et al. Probing the microRNA and small interfering RNA pathways with virus-encoded suppressors of RNA silencing. Plant cell. 2004;16:1235–1250. doi: 10.1105/tpc.020719. [DOI] [PMC free article] [PubMed] [Google Scholar] [Retracted]
  • 14.Rajendra M., Radhamani A., Trent H. S., et al. RNA viruses as inducers, suppressors and targets of post-transcriptional gene silencing. Plant Mol Biol. 2000;43:295–306. doi: 10.1023/A:1006456000564. [DOI] [PubMed] [Google Scholar]
  • 15.Roth B. M., Pruss G. J., Vance V. B. Plant viral suppressors of RNA silencing. Virus Res. 2004;102(1):97–108. doi: 10.1016/j.virusres.2004.01.020. [DOI] [PubMed] [Google Scholar]
  • 16.Stevenson M. Therapeutic Potential of RNA Interference. N Engl J Med. 2004;351:1772–1777. doi: 10.1056/NEJMra045004. [DOI] [PubMed] [Google Scholar]
  • 17.Vicki V., Herve V. RNA silencing in plants—defense and counterdefense. Science. 2001;292(22):2277–2280. doi: 10.1126/science.1061334. [DOI] [PubMed] [Google Scholar]
  • 18.Westhof E. How to silence silencing. Chem. Biol. 2004;11(2):158–160. doi: 10.1016/S1074-5521(04)00038-9. [DOI] [PubMed] [Google Scholar]
  • 19.Zsuzsanna L., Daniel S., Jozsef B. Double-stranded RNA-binding proteins could suppress RNA interference-mediated antiviral defences. J Gen Virol. 2003;84:975–980. doi: 10.1099/vir.0.18987-0. [DOI] [PubMed] [Google Scholar]

Articles from Virologica Sinica are provided here courtesy of Nature Publishing Group

RESOURCES