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Journal of Virology, July 2004, p. 7619-7633, Vol. 78, No. 14
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.14.7619-7633.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Repression and Derepression of Minus-Strand Synthesis in a Plus-Strand RNA Virus Replicon

Guohua Zhang, Jiuchun Zhang, and Anne E. Simon*

Department of Cell Biology and Molecular Genetics, University of Maryland—College Park, College Park, Maryland 20742

Received 13 January 2004/ Accepted 6 March 2004

Plus-strand viral RNAs contain sequences and structural elements that allow cognate RNA-dependent RNA polymerases (RdRp) to correctly initiate and transcribe asymmetric levels of plus and minus strands during RNA replication. cis-acting sequences involved in minus-strand synthesis, including promoters, enhancers, and, recently, transcriptional repressors (J. Pogany, M. R. Fabian, K. A. White, and P. D. Nagy, EMBO J. 22:5602-5611, 2003), have been identified for many viruses. A second example of a transcriptional repressor has been discovered in satC, a replicon associated with turnip crinkle virus. satC hairpin 5 (H5), located proximal to the core hairpin promoter, contains a large symmetrical internal loop (LSL) with sequence complementary to 3'-terminal bases. Deletion of satC 3'-terminal bases or alteration of the putative interacting bases enhanced transcription in vitro, while compensatory exchanges between the LSL and 3' end restored near-normal transcription. Solution structure analysis indicated that substantial alteration of the satC H5 region occurs when the three 3'-terminal cytidylates are deleted. These results indicate that H5 functions to suppress synthesis of minus strands by sequestering the 3' terminus from the RdRp. Alteration of a second sequence strongly repressed transcription in vitro and accumulation in vivo, suggesting that this sequence may function as a derepressor to free the 3' end from interaction with H5. Hairpins with similar sequence and/or structural features that contain sequence complementary to 3'-terminal bases, as well as sequences that could function as derepressors, are located in similar regions in other carmoviruses, suggesting a general mechanism for controlling minus-strand synthesis in the genus.

* Corresponding author. Mailing address: Department of Cell Biology and Molecular Genetics, Microbiology Building, University of Maryland—College Park, College Park, MD 20742. Phone: (301) 405-8975. Fax: (301) 805-1318. E-mail: Anne_Simon{at}umail.umd.edu.

Journal of Virology, July 2004, p. 7619-7633, Vol. 78, No. 14
0022-538X/04/$08.00+0     DOI: 10.1128/JVI.78.14.7619-7633.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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