CD154 Costimulated Ovine Primary B Cells, a Cell Culture System That Supports Productive Infection by Bovine Leukemia Virus

Abstract

Bovine leukemia virus (BLV) is closely associated with the development of B-cell leukemia and lymphoma in cattle. BLV infection has also been studied extensively in an in vivo ovine model that provides a unique system for studying B-cell leukemogenesis. There is no evidence that BLV can directly infect ovine B cells in vitro, and there are no direct data regarding the oncogenic potential of the viral Tax transactivator in B cells. Therefore, we developed ovine B-cell culture systems to study the interaction between BLV and its natural target, the B cell. In this study, we used murine CD154 (

Transcriptional Regulation of the Bovine Leukemia Virus Promoter by the Cyclic AMP-response Element Modulator τ Isoform

“…Human HeLa cells were maintained in Dulbecco's modified Eagle's medium (Invitrogen) supplemented with 5% fetal bovine serum, 50 units of penicillin/ml, and 50 g of streptomycin/ml. The YR2 B-cell line is derived from leukemic cells of a BLV-infected sheep and contains a single, monoclonally integrated silent provirus with two mutations in Tax BLV that impair the infectious potential of the integrated provirus (20). The YR2 cell line was maintained in OptiMEM medium (Invitrogen) supplemented with 10% fetal bovine serum, 50 units of penicillin/ml, and 50 g of streptomycin/ml.…”

“…To this end, we performed supershift EMSA experiments using the three 21-bp TxREs as probes. These probes were incubated with nuclear extracts prepared from YR2 cells derived from leukemic cells of a BLV-infected sheep (20), either mock-treated or treated for 1 h with phorbol ester phorbol 12-myristate 13-acetate plus Ca 2ϩ ionophore ionomycin (P ϩ I), a combination known to activate BLV expression (29,30). Incubations were carried out in the presence of either a polyclonal anti-CREM antibody (Fig.…”

Deacetylase Inhibitors and the Viral Transactivator TaxBLV Synergistically Activate Bovine Leukemia Virus Gene Expression via a cAMP-responsive Element- and cAMP-responsive Element-binding Protein-dependent Mechanism

“…The YR2 B-cell line is derived from leukemic cells of a BLV-infected sheep and contains a single, monoclonally integrated silent provirus with two mutations in Tax BLV that impair the infectious potential of the integrated provirus. The YR2 LTaxSN cell line is derived from YR2 cells and contains a retroviral vector gene encoding for a transactivation-competent Tax BLV (42). YR2 and YR2 LTaxSN cell lines were maintained in Opti-MEM medium supplemented with 10% fetal bovine serum, 50 units of penicillin/ml, and 50 g of streptomycin/ml.…”

Disruption of B-cell homeostatic control mediated by the BLV-Tax oncoprotein: association with the upregulation of Bcl-2 and signaling through NF-κB

“…Our main objective was thus to evaluate the oncogenic potential of the transcriptional transactivator in ovine B-cells, a relevant cell type for studying leukemogenic processes. We have previously shown that B-cells isolated from a variety of lymphoid tissues from sheep including ileal and jejunal PP were permissive to BLV infection (Van den Broeke et al, 2001). To examine the impact of Tax in a pure ovine B-cell population, we used ovine B-cell clones derived from the jejunal PP of a lamb (Griebel et al, 2000).…”

“…Our main objective was thus to evaluate the oncogenic potential of the viral Tax transactivator in a relevant cell type. We previously developed primary ovine B-cell cultures and showed that B-cells isolated from a variety of sheep lymphoid tissues including ileal and jejunal Peyer's patches (PP) were permissive to BLV infection Van den Broeke et al, 2001). This culture system, dependent on costimulation with CD154 and g chain (g c )-common cytokines, provides an excellent in vitro model for investigating Tax BLV -associated leukemogenic processes.…”