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Published online on February 20, 2007, 10.1073/pnas.0611617104
PNAS | February 27, 2007 | vol. 104 | no. 9 | 3129-3134


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BIOLOGICAL SCIENCES / BIOCHEMISTRY
RNA-binding proteins that inhibit RNA virus infection

Jian Zhu*,{dagger}, Kodetham Gopinath{ddagger}, Ayaluru Murali{ddagger}, Guanghui Yi{ddagger}, S. Diane Hayward*, Heng Zhu*,{dagger},§, and Cheng Kao{ddagger},§

*Department of Pharmacology and Molecular Sciences and {dagger}High Throughput Biology Center, Johns Hopkins University School of Medicine, Baltimore, MD 21205; and {ddagger}Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843

Communicated by Ignacio Tinoco, Jr., University of California, Berkeley, CA, December 27, 2006 (received for review October 9, 2006)

Arrays of >5,000 Saccharomyces cerevisiae proteins were screened to identify proteins that can preferentially bind a small RNA hairpin that contains a clamped adenine motif (CAM). A CAM is required for the replication of Brome Mosaic Virus (BMV), a plant-infecting RNA virus that can replicate in S. cerevisiae. Several hits were selected for further characterization in Nicotiana benthamiana. Pseudouridine Synthase 4 (Pus4) and the Actin Patch Protein 1 (App1) modestly reduced BMV genomic plus-strand RNA accumulation, but dramatically inhibited BMV systemic spread in plants. Pus4 also prevented the encapsidation of a BMV RNA in plants and the reassembly of BMV virions in vitro. These results demonstrate the feasibility of using proteome arrays to identify specific RNA-binding proteins for antiviral activities. Furthermore, the effects of Pus4 suggest that the CAM-containing RNA motif provides a regulatory link between RNA replication and encapsidation.

Brome mosaic virus | protein–RNA interaction | pseudouridine synthase 4 | yeast proteome chip | viral RNA replication

Author contributions: J.Z. and K.G. contributed equally to this work; J.Z., H.Z., and C.K. designed research; J.Z., K.G., A.M., G.Y., and C.K. performed research; J.Z., K.G., A.M., G.Y., H.Z., and C.K. contributed new reagents/analytic tools; J.Z., K.G., A.M., G.Y., S.D.H., H.Z., and C.K. analyzed data; and J.Z., K.G., H.Z., and C.K. wrote the paper.

The authors declare no conflict of interest.

This article contains supporting information online at www.pnas.org/cgi/content/full/0611617104/DC1.

§To whom correspondence may be addressed. E-mail: ckao{at}tamu.edu or hzhu4{at}jhmi.edu

© 2007 by The National Academy of Sciences of the USA


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