The overall goal of this proposal is to determine the role that HIV plays directly in the pathogenesis of AIDS-associated lymphoma. In preliminary studies of 18 non-B-cell malignancies, a clonally integrated form of HIV was identified in 14 using the inverse polymerase chain reaction (IPCR) method. IPCR sensitivity controls suggest that at least 5% of the cells within a tumor cell population would have to have a clonal HIV in order to give a positive IPCR result. The spectrum of malignancies containing clonal forms of HIV included polyclonal mixed immunophenotype large cell lymphoma, cutaneous and systemic T-cell lymphomas, Hodgkin's disease, and angioimmunoblastic lymphadenopathy (AILD). In the majority of cases tested HIV expression was localized to tumor-associated macrophages. A common integration site in the 3' exon of the fur gene just 5' to the c-fes oncogene was identified in six of the tumors and intense fes immunostaining was observed in several cases analyzed. These observations suggest that HIV may be associated with cell transformation through an insertional mutagenesis process. In the current proposal we will identify the clinical spectrum of lymphomas containing clonal HIV and map integration sites through sequencing of IPCR products. The exact identification of cells containing the clonal HIV will be determined through cell sorting and IPCR. Cytokines expressed by separated cells will be determined by reverse transcriptase PCR (RT-PCR). Tumors with HIV near fes will be evaluated for fes and activated fes expression by flow cytometry and immunostaining. The identity of fes expressing cells will also be determined by flow cytometry. Expression vector studies using the 5' fes promoter region with a 5' full-length HIV and a 3' luciferase reporter gene will allow mutational studies on HIV genes that may influence fes expression. HIV's will be cloned from two tumors showing increased fes expression for sequence analysis and expression vector construction. Because the six cases of fes associated HIV integrations were integrated in the fur gene, studies on fur expression will be performed. As new genes are identified adjacent to tumor associated clonal forms of HIV, the influence of HIV on those genes and transformation will be studied. These studies are designed to define mechanisms of tumorigenesis in the AIDS non-B-cell lymphoma as well as evaluate the role of clonal forms of HIV in early stages of B lymphoma development.