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Journal of Virology, July 1999, p. 5481-5489, Vol. 73, No. 7
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Identification of Replication Specificity Determinants in Two Strains of Tomato Leaf Curl Virus from New Delhi

Anju Chatterji, Malla Padidam, Roger N. Beachy,dagger and Claude M. Fauquet*

International Laboratory for Tropical Agricultural Biotechnology, Division of Plant Biology, The Scripps Research Institute, La Jolla, California 92037

Received 2 November 1998/Accepted 29 March 1999

We used two strains of tomato leaf curl virus from New Delhi to investigate specificity in replication of their cognate genomes. The strains share 94% sequence identity and are referred to as severe and mild on the basis of symptoms on tomato and tobacco. Replication assays in tobacco protoplasts and plants showed that a single amino acid change, Asn10 to Asp in the N terminus of Rep protein, determines specificity for replication of the two strains based upon its interaction with the origin of replication (ori) sequences. The change of Asp10 to Asn in Rep protein of the mild strain coupled with point mutations at the 3rd and 10th nucleotides of the 13-mer binding site altered its replication ability, resulting in increased levels of virus accumulation. Similarly, changing Asn10 to Asp in Rep protein of the severe strain impaired replication of the virus and altered its severe phenotype in plants. Site-directed mutations made in ori and Asn10 of Rep protein suggested that Asn10 recognizes the third base pair of the putative binding site sequence GGTGTCGGAGTC in the severe strain.

* Corresponding author. Mailing address: International Laboratory for Tropical Agricultural Biotechnology (ILTAB/ORSTOM-TSRI), Division of Plant Biology, BCC 206, The Scripps Research Institute, 10550 N. Torrey Pines Rd., La Jolla, CA 92037. Phone: (619) 784-2909. Fax: (619) 784-2994. E-mail: iltab{at}scripps.edu.

dagger Present address: Donald Danforth Plant Center, St. Louis, MO 63105.

Journal of Virology, July 1999, p. 5481-5489, Vol. 73, No. 7
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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