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Replication of the complex retrovirus mouse mammary tumor virus (MMTV) is antagonized by murine Apobec3 (mA3), a member of the Apobec family of cytidine deaminases. We have shown that MMTV-encoded Rem protein inhibits proviral mutagenesis by the Apobec enzyme, activation-induced cytidine deaminase (AID) during viral replication in BALB/c mice. To further study the role of Rem in vivo, we have infected C57BL/6 (B6) mice with a superantigen-independent lymphomagenic strain of MMTV (TBLV-WT) or a mutant strain that is defective in Rem and its cleavage product Rem-CT (TBLV-SD). Compared to BALB/c, B6 mice were more susceptible to TBLV infection and tumorigenesis. Furthermore, unlike MMTV, TBLV induced T-cell tumors in B6 μMT mice, which lack membrane-bound IgM and conventional B-2 cells. At limiting viral doses, loss of Rem expression in TBLV-SD-infected B6 mice accelerated tumorigenesis compared to TBLV-WT in either wild-type B6 or AID-knockout mice. Unlike BALB/c results, high-throughput sequencing indicated that proviral G-to-A or C-to-T mutations were unchanged regardless of Rem expression in B6 tumors. However, knockout of both AID and mA3 reduced G-to-A mutations. Ex vivo stimulation showed higher levels of mA3 relative to AID in B6 compared to BALB/c splenocytes, and effects of agonists differed in the two strains. RNA-Seq revealed increased transcripts related to growth factor and cytokine signaling in TBLV-SD-induced tumors relative to TBLV-WT-induced tumors, consistent with another Rem function. Thus, Rem-mediated effects on tumorigenesis in B6 mice are independent of Apobec-mediated proviral hypermutation. Author summary: Retroviruses cause lifelong infections resulting from their ability to thwart innate immunity. The Apobec family of cytidine deaminases are part of the innate immune response that recognizes and mutates foreign nucleic acids, including those from multiple viruses. Retroviral antagonists of Apobecs have been identified, including mouse mammary tumor virus (MMTV)-encoded Rem protein. Previous experiments have shown that Rem-null MMTV or closely related TBLV proviruses from BALB/c tumors accumulate G-to-A and C-to-T mutations typical of Apobecs compared to wild-type proviruses expressing Rem. The difference in mutations between Rem-expressing and non-expressing MMTV strains largely disappeared in mice lacking the Apobec family member, activation-induced cytidine deaminase (AID). These results suggested that Rem is an AID antagonist. In this study, we attempted to study AID-mediated mutations of TBLV proviruses lacking Rem expression obtained from tumors in C57BL/6 (B6) wild-type and AID-knockout backgrounds. Surprisingly, no differences in G-to-A mutations were observed in TBLV proviruses regardless of Rem expression, yet such mutations were significantly reduced in proviruses obtained from mA3/AID-double knockout mice relative to those from wild-type B6 or AID-knockout mice. Many cellular mRNAs involving the innate immune response, but not Apobecs, were elevated in the absence relative to the presence of Rem expression on the B6 AID-knockout background. These results revealed that Apobec-mediated mutagenesis is dependent on mouse strain and suggested a second means of Rem-dependent immune evasion. [ABSTRACT FROM AUTHOR]

Abstract Background Mouse mammary tumor virus (MMTV) encodes the Rem protein, an HIV Rev-like protein that enhances nuclear export of unspliced viral RNA in rodent cells. We have shown that Rem is expressed from a doubly spliced RNA, typical of complex retroviruses. Several recent reports indicate that MMTV can infect human cells, suggesting that MMTV might interact with human retroviruses, such as human immunodeficiency virus (HIV), human T-cell leukemia virus (HTLV), and human endogenous retrovirus type K (HERV-K). In this report, we test whether the export/regulatory proteins of human complex retroviruses will increase expression from vectors containing the Rem-responsive element (RmRE). Results MMTV Rem, HIV Rev, and HTLV Rex proteins, but not HERV-K Rec, enhanced expression from an MMTV-based reporter plasmid in human T cells, and this activity was dependent on the RmRE. No RmRE-dependent reporter gene expression was detectable using Rev, Rex, or Rec in HC11 mouse mammary cells. Cell fractionation and RNA quantitation experiments suggested that the regulatory proteins did not affect RNA stability or nuclear export in the MMTV reporter system. Rem had no demonstrable activity on export elements from HIV, HTLV, or HERV-K. Similar to the Rem-specific activity in rodent cells, the RmRE-dependent functions of Rem, Rev, or Rex in human cells were inhibited by a dominant-negative truncated nucleoporin that acts in the Crm1 pathway of RNA and protein export. Conclusion These data argue that many retroviral regulatory proteins recognize similar complex RNA structures, which may depend on the presence of cell-type specific proteins. Retroviral protein activity on the RmRE appears to affect a post-export function of the reporter RNA. Our results provide additional evidence that MMTV is a complex retrovirus with the potential for viral interactions in human cells.