Approximately 8% of the genomes of mammals, including humans and mice, are comprised of retroviral elements acquired by infection of germ line cells during the course of evolution. Retroviral insertions in our genome number about 40,000. Most endogenous retrovirus elements are defective for replication however several contain one or more viral genes that are expressed during development and certain physiological or pathological conditions. Little is known about the control of retrovirus expression or the influence of such expression on the physiology or pathology of the host. An extensively investigated group of endogenous retroviruses are those giving rise to recombinant murine leukemia viruses (MuLVs) in mice. Upon infection of mice with exogenous ecotropic MuLVs, members of this group undergo recombination to generate new MuLVs with an altered infectious host range. Recombination requires transcription of the endogenous retroviruses. Although the endogenous polytropic proviruses are transcribed;replication of the endogenous polytropic viruses in the absence of recombination has not been observed. This may, in many cases, reflect defects such as point mutations or deletions in the endogenous viral genome but may also be influenced by the activity of various restriction factors. The fact that exogenous MuLVs are capable of replicating in mice indicates that they have evolved mechanisms to circumvent the activity of at least some of the restriction factors such as the murine APOBEC3. Thus, exogenous retroviruses might facilitate through complementation, active replication of endogenous retroviruses. We have found that infection of mice by an exogenous virus results in the infectious transfer of complete endogenous proviral genetic sequences. This includes proviruses which are severely defective and possess large deletions as well as proviruses that are full-length. Furthermore, the transferred sequences are transcribed and packaged into virions released from the newly infected cells. At early times after infection with the Friend MuLV, packaging and transfer of intact endogenous retroviruses is much more prevalent than recombination. Endogenous retroviruses are transferred as early as one day after infection. Thus, the transcripts are captured within a single in vivo replication cycle and originate from among the initially infected host cells. The mobilization of intact endogenous retroviruses is unprecedented and may have important implications for the involvement of endogenous retroviruses in disease processes. We have extended our observations to further characterize the endogenous viruses mobilized after infection by exogenous retroviruses. We have found that the mobilized endogenous viruses appear to be specifically limited to endogenous polytropic proviruses at the exclusion of other endogenous retroviral elements. The endogenous polytropic proviruses are comprised of two structural subclasses termed Polytropic (PT) and Modified Polytropic (mPT). We observed a distinct shift in the subclass of proviruses from the mPT subclass to the PT subclass of polytropic proviruses detected during the course of infection. These results suggest the spread of infection to cells expressing different classes of polytropic viruses or an alteration in the expression of the endogenous proviruses in infected cells. Exogenous mouse retrovirusesencode a glycosylated gag protein (gGag) originating from an alternate translation start site upstream of the methionine start site of the gag structural polyproteins. The functions of gGag remain unclear, but mutations that eliminate its synthesis severely impede in vivo replication of the virus with little effect on replication in fibroblastic cell lines. APOBEC3 proteins have evolved as innate defenses against retroviral infections. Both mice and humans express APOBEC3 proteins that have cytidine deaminase activity leading to hypeermutation of viral transcripts and inactivation of infecting retroviruses. HIV encodes the VIF protein to evade human APOBEC3G (hA3G), however mouse retroviruses do not encode a VIF homologue and it has not been understood how they evade mouse APOBEC3 (mA3). We have found that a mouse retrovirus utilizes its glycosylated gag protein (gGag) to evade APOBEC3. gGag is critical for infection of in vitro cell lines in the presence of APOBEC3. Furthermore, a gGag-deficient virus restricted for replication in wild-type mice replicates efficiently in APOBEC3 knockout mice implicating a novel role of gGag in circumventing the action of APOBEC3 in vivo. In 2013 we have continued to focus on the elucidation of the mechanism by which the gGag protein abrogates the action of APOBEC3. Human APOBEC3G (hA3g), is counteracted by the Vif protein of HIV which depletes hA3g from infected cells by facilitating its degradation through the proteosome. We have found that a gGag-containing MuLV does not deplete mA3 from an infected cell We have observed that inhibition by mA3 packaged within gGag-deficient virions correlates with a decrease in the level of transcripts upon infection of cells and appears largely independent of deamination activity. However, we have found that mA3 packaged at very high levels in wild-type virions results in a loss of fidelity of the viral polymerase resulting a substantial increase in the overall mutation rate, but not hypermutation at G residues. Further, we have found that the ecotropic MuLV, AKV does undergo G to A hypermutation in the presence of mA3 even though AKV does encode a gGag protein. The endogenous retroviral envelope glycoprotein, gp70 is implicated in murine lupus nephritis. This protein is secreted by hepatocytes as an acute phase protein and has been believed to be a product of an endogenous xenotropic virus. However,we have found that endogenous polytropic viruses provide additional sources of serum gp70. To better understand the genetic basis of the expression of serum gp70, we analyzed the abundance of xenotropic and polytropic gp70 RNAs in livers and the genomic composition of corresponding endogenous proviruses in various strains of mice, including two different Sgp (serum gp70 production) congenic mice (Sgp3 and Sgp4). These studies revealed a significant contribution of polytropic gp70s to serum gp70 and a tight regulation of control by the Sgp3 and Sgp4 genes. Sgp3 regulates the transcription of xenotropic, polytropic and modified polytropic (mPT)viruses, and Sgp4 the transcription of only xenotropic viruses. In 2013 these studies were extended to examine the effects of a third gene, Sgp5. The Sgp5 locus enhanced the expression of xenotropic and mPT viruses, thereby upregulating the production of serum gp70. These data indicate a distinct action of the Sgp5 locus on the expression of endogenous retroviruses as compared with two other Sgp loci. Moreover, it was found that Sgp3 and Sgp4 acted synergistically to elevate the transcription of a potentially replication-competent Xmv18 provirus and the production of serum gp70. The results of these studies indicate that the combined effect of three different Sgp loci markedly enhance the expression of endogenous retroviruses and their gene product, serum gp70, thereby contributing to the formation of nephritogenic gp70-anti-gp70 immune complexes in murine lupus.

Project Start
Project End
Budget Start
Budget End
Support Year
5
Fiscal Year
2013
Total Cost
$257,427
Indirect Cost
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