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Journal of Virology, April 2007, p. 4348-4356, Vol. 81, No. 8
0022-538X/07/$08.00+0     doi:10.1128/JVI.01289-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Determination of Kaposi's Sarcoma-Associated Herpesvirus C-Terminal Latency-Associated Nuclear Antigen Residues Mediating Chromosome Association and DNA Binding

Brenna Kelley-Clarke, Mary E. Ballestas, Viswanathan Srinivasan, Andrew J. Barbera, Takashi Komatsu, Te-Ana Harris, Mia Kazanjian, and Kenneth M. Kaye^*

Channing Laboratory, Departments of Medicine, Brigham and Women's Hospital, Harvard Medical School, 181 Longwood Ave., Boston, Massachusetts 02115

Received 19 June 2006/ Accepted 29 December 2006

Kaposi's sarcoma-associated herpesvirus latency-associated nuclear antigen (LANA) tethers viral terminal repeat (TR) DNA to mitotic chromosomes to mediate episome persistence. The 1,162-amino-acid LANA protein contains both N- and C-terminal chromosome attachment regions. The LANA C-terminal domain self-associates to specifically bind TR DNA and mitotic chromosomes. Here, we used alanine scanning substitutions spanning residues 1023 to 1145 to investigate LANA self-association, DNA binding, and C-terminal chromosome association. No residues were essential for LANA oligomerization, as assayed by coimmunoprecipitation experiments, consistent with redundant roles for amino acids in self-association. Different subsets of amino acids were important for DNA binding, as assayed by electrophoretic mobility shift assay, and mitotic chromosome association, indicating that distinct C-terminal LANA subdomains effect DNA and chromosome binding. The DNA binding domains of LANA and EBNA1 are predicted to be structurally homologous; certain LANA residues important for DNA binding correspond to those with roles in EBNA1 DNA binding, providing genetic support for at least partial structural homology. In contrast to the essential role of N-terminal LANA chromosome targeting residues in DNA replication, deficient C-terminal chromosome association did not reduce LANA-mediated DNA replication.

Published ahead of print on 7 February 2007.

Present address: Department of Pediatrics, University of Alabama School of Medicine, Birmingham, AL 35233.

Present address: PTC Therapeutics, South Plainfield, NJ 07080.

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