Effective inhibition in animals of viral pathogenesis by a ribozyme derived from RNase P catalytic RNA.

Abstract

A functional RNase P ribozyme (M1GS RNA) was constructed to target the overlapping mRNA region of two murine cytomegalovirus (MCMV) capsid proteins essential for viral replication: the assembly protein (mAP) and M80. The customized ribozyme efficiently cleaved the target mRNA sequence in vitro. Moreover, 80% reduction in the expression of mAP and M80 and a 2,000-fold reduction in viral growth were observed in cells expressing the ribozyme. In contrast, there was no significant reduction in viral gene expression and growth in cells that either did not express the ribozyme or produced a "disabled" ribozyme carrying mutations that abolished its catalytic activity. When the ribozyme-expressing constructs were delivered into MCMV-infected SCID mice via a modified "hydrodynamic transfection" procedure, expression of ribozymes was observed in the livers and spleens. Compared with the control animals that did not receive any M1GS constructs or received the disabled ribozyme construct, animals receiving the functional ribozyme construct exhibited a significant reduction of viral gene expression and infection. Viral titers in the spleens, livers, lungs, and salivary glands of the functional ribozyme-treated SCID mice at 21 days after infection were 200- to 2,000-fold lower than those in the control animals. Moreover, survival of the infected animals significantly improved upon receiving the functional ribozyme construct. Our study examines the use of M1GS ribozymes for inhibition of gene expression in animals and demonstrates the utility of RNase P ribozymes for gene targeting applications in vivo.

• Authors: Yong Bai, Phong Trang, Hongjian Li, Kihoon Kim, Tianhong Zhou, Fenyong Liu
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 105 Issue 31 Pg. 10919-24 (Aug 05 2008) ISSN: 1091-6490 [Electronic] United States
• PMID: 18663226 (Publication Type: Comparative Study, Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• Capsid Proteins
• DNA Primers
• RNA, Catalytic
• Ribonuclease P

Topics

• Animals
• Blotting, Northern
• Blotting, Western
• Capsid Proteins (metabolism)
• DNA Primers (genetics)
• Gene Expression Regulation, Viral (genetics)
• Gene Targeting (methods)
• Mice
• Mice, SCID
• Muromegalovirus (chemistry)
• NIH 3T3 Cells
• RNA, Catalytic (metabolism)
• Ribonuclease P (metabolism)
• Virus Replication (genetics)