Abstract PO-31: HIV-1 transactivator of transcription deregulates key Burkitt lymphoma associated oncogenes at both the transcriptional and post-transcriptional level

Abstract

Abstract Burkitt lymphoma (BL) is one of the most common HIV-associated lymphomas. This cancer represents one of the most frequent causes of mortality among HIV+ people in Southern Africa, which has the highest incidence of HIV/AIDS worldwide. As is the case for Kaposi sarcoma, recent reports associate an oncogenic function with HIV in lymphoma development. This study explores the oncogenic potential of HIV-1 protein Transactivator of transcription (Tat) in BL via its ability to manipulate the expression of c-MYC and AID, two key drivers of disease in BL. The ability of HIV-1 Tat to influence the activity of the full-length (FL) (2861 bp) c-MYC promoter was assessed using Dual Luciferase Reporter (DLR) assays. Using sequential deletions, the minimal promoter region mediating the effect was identified. Site-directed mutagenesis (SDM) was used to identify promoter elements mediating the response. Co-immunoprecipitation assays (Co-IP) were used to assess interactions between Tat and the AP-1 factor JunB. Chromatin Immunoprecipitation (ChIP) assays were done to confirm that JunB bound the promoter in vivo. To assess the ability of Tat to mediate the expression of miRNA-181b-5p, a known repressor of AICDA, Ramos cells were transfected with Tat followed by qRT-PCR. Corresponding changes in AID protein levels were determined using Western blot. To assess whether miRNA-181b-5p targets AICDA in B cells, the FL AICDA 3'UTR was cloned into the pGL3-Promoter vector and DLR assays were performed in the presence of miRNA-181b-5p mimic. Our data reveal that Tat enhances c-MYC promoter activity in a dose-dependent manner, with a maximum activity of 2.6-fold recorded when 500ng of pcDNA-Tat was used on 400ng of pGL3-c-MYC plasmid harboring the FL promoter. A loss of 35% activity was observed [1.7 (±0.05) fold] upon promoter deletion and two AP-1 sites (positions -1128 bp and -1376 bp) within the deleted region were found to mediate the Tat-dependent increase. A loss in activity of 20.6% was recorded when AP-1 site 1 was mutated, and 25% when AP-1 site 2 was mutated. A double mutant decreased the activity by 29%. Co-IP revealed a strong interaction between Tat and AP-1 factor JunB, and ChIP data showed that JunB was strongly recruited to those two AP-1 sites in the presence of Tat. Our result also reveals that the expression of the AICDA specific miRNA181b-5p is significantly reduced in the presence of Tat, which correlates with an increase in AID protein levels. We also confirm that the AICDA FL 3'UTR, which contains putative miRNA-181b-5p binding sites, is partially repressed by a miRNA-181b-5p mimic. Our study reveals that Tat potentiates oncogenesis by enhancing the expression of c-MYC and AID, two key lymphoma drivers. We show that Tat can enhance c-MYC expression by enhancing the recruitment of cellular TFs to the promoter. We also show that the same HIV protein can enhance the expression of AID by modulating the expression of miRNA-181b-5p. Citation Format: Leonardo Alves de Souza Rios, Nontlantla Mdletshe, Shaheen Mowla. HIV-1 transactivator of transcription deregulates key Burkitt lymphoma associated oncogenes at both the transcriptional and post-transcriptional level [abstract]. In: Proceedings of the AACR Virtual Meeting: Advances in Malignant Lymphoma; 2020 Aug 17-19. Philadelphia (PA): AACR; Blood Cancer Discov 2020;1(3_Suppl):Abstract nr PO-31.