Inhibition of Murine Cytomegalovirus Infection in Animals by RNase P-Associated External Guide Sequences

Wei Li; Jingxue Sheng; Mengqiong Xu; Gia-Phong Vu; Zhu Yang; Yujun Liu; Xu Sun; Phong Trang; Sangwei Lu; Fenyong Liu

doi:10.1016/j.omtn.2017.10.007

Inhibition of Murine Cytomegalovirus Infection in Animals by RNase P-Associated External Guide Sequences

Mol Ther Nucleic Acids. 2017 Dec 15:9:322-332. doi: 10.1016/j.omtn.2017.10.007. Epub 2017 Oct 16.

Authors

Wei Li¹, Jingxue Sheng², Mengqiong Xu¹, Gia-Phong Vu³, Zhu Yang⁴, Yujun Liu⁵, Xu Sun⁶, Phong Trang⁷, Sangwei Lu², Fenyong Liu⁸

Affiliations

¹ Department of Biotechnology, College of Life Science and Technology, Jinan University, Guangzhou, Guangdong 510632, China.
² Program in Comparative Biochemistry, University of California, Berkeley, Berkeley, CA 94720, USA; School of Public Health, University of California, Berkeley, Berkeley, CA 94720, USA.
³ School of Public Health, University of California, Berkeley, Berkeley, CA 94720, USA.
⁴ Jiangsu Affynigen Biotechnolgies, Inc., Taizhou, Jiangsu 225300, China; Guangzhou Qinheli Biotechnolgies, Inc., Guangzhou, Guangdong 510600, China.
⁵ Department of Biotechnology, College of Life Science and Technology, Jinan University, Guangzhou, Guangdong 510632, China; School of Medicine, St. George's University, Grenada, West Indies; School of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, Shandong 250355, China.
⁶ Department of Biotechnology, College of Life Science and Technology, Jinan University, Guangzhou, Guangdong 510632, China; Guangzhou Qinheli Biotechnolgies, Inc., Guangzhou, Guangdong 510600, China.
⁷ Program in Comparative Biochemistry, University of California, Berkeley, Berkeley, CA 94720, USA.
⁸ Department of Biotechnology, College of Life Science and Technology, Jinan University, Guangzhou, Guangdong 510632, China; Program in Comparative Biochemistry, University of California, Berkeley, Berkeley, CA 94720, USA; School of Public Health, University of California, Berkeley, Berkeley, CA 94720, USA. Electronic address: liu_fy@berkeley.edu.

Abstract

External guide sequence (EGS) RNAs are associated with ribonuclease P (RNase P), a tRNA processing enzyme, and represent promising agents for gene-targeting applications as they can direct RNase-P-mediated cleavage of a target mRNA. Using murine cytomegalovirus (MCMV) as a model system, we examined the antiviral effects of an EGS variant, which was engineered using in vitro selection procedures. EGSs were used to target the shared mRNA region of MCMV capsid scaffolding protein (mCSP) and assemblin. In vitro, the EGS variant was 60 times more active in directing RNase P cleavage of the target mRNA than the EGS originating from a natural tRNA. In MCMV-infected cells, the variant reduced mCSP expression by 92% and inhibited viral growth by 8,000-fold. In MCMV-infected mice hydrodynamically transfected with EGS-expressing constructs, the EGS variant was more effective in reducing mCSP expression, decreasing viral production, and enhancing animal survival than the EGS originating from a natural tRNA. These results provide direct evidence that engineered EGS variants with higher targeting activity in vitro are also more effective in reducing gene expression in animals. Furthermore, our findings imply the possibility of engineering potent EGS variants for therapy of viral infections.

Keywords: RNase P; antisense; gene targeting; gene therapy; herpesvirus.

Abstract

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