Kaposi's sarcoma (KS) associated herpesvirus (KSHV or HHV8) is a recently discovered virus which is associated with malignancies in patients, especially those with AIDS. KSHV is found in KS, primary effusion lymphocytes (PELs) and multicentric Castleman's disease, a lymphoproliferative disorder. Similar to a closely related herpesvirus, Epstein-Barr virus, KSHV infection in malignancy and in PEL cell lines is primarily latent, with viral DNA persisting as an extra-chromosomal, circular episome. The latency associated nuclear antigen (LANA or LNA) is an 1162 amino acid KSHV protein expressed in a punctate distribution in the nuclei of latently infected cells. The serum of KSHV infected patients usually reacts with LANA and epidemiologic studies have often used this reactivity to determine KSHV seropositivity. The function of LANA is unknown. We now show that LANA colocalizes with KSHV episomes on cell chromosomes and LANA permits maintenance of KSHV episomes in cells. These results are consistent with a model in which LANA mediates segregation of episomes to daughter cells by tether episomes to cell chromosomes during mitosis. This project will test this model by determine if LANA colocalization with episomes on chromosomes is essential for episome maintenance. First, extensive mutational analysis of LANA will be performed to determine the LANA domains that mediate colocalization with chromosomes and with KSHV episomes. LANA mutants which cannot colocalize with chromosomes or episomes will be tested for loss of the ability to permit maintenance of KSHV episomes in cells. Next, the KSHV DNA element essential for LANA mediated episome maintenance will be defined. Experiments will then investigate the nature of the interaction between LANA and the essential KSHV DNA element.
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