Dissection of genetic mechanisms governing the expression of serum retroviral gp70 implicated in murine lupus nephritis

Abstract

The endogenous retroviral envelope glycoprotein, gp70, implicated in murine lupus nephritis is secreted by hepatocytes as an acute phase protein, and it has been thought to be a product of an endogenous xenotropic virus, NZB-X1. However, since endogenous polytropic (PT) and modified polytropic (mPT) viruses encode gp70s that are closely related to xenotropic gp70, these viruses can be additional sources of serum gp70. To better understand the genetic basis of the expression of serum gp70, we analyzed the abundance of xenotropic, PT, or mPT gp70 RNAs in livers and the genomic composition of corresponding proviruses in various strains of mice, including two different Sgp (serum gp70 production) congenic mice. Our results demonstrated that the expression of different viral gp70 RNAs was remarkably heterogeneous among various mouse strains and that the level of serum gp70 production was regulated by multiple structural and regulatory genes. Additionally, a significant contribution of PT and mPT gp70s to serum gp70 was revealed by the detection of PT and mPT, but not xenotropic transcripts in 129 mice, and by a closer correlation of serum levels of gp70 with the abundance of PT and mPT gp70 RNAs than with that of xenotropic gp70 RNA in Sgp3 congenic mice. Furthermore, the injection of lipopolysaccharides selectively up-regulated the expression of xenotropic and mPT gp70 RNAs, but not PT gp70 RNA. Our data indicate that the genetic origin of serum gp70 is more heterogeneous than previously thought, and that distinct retroviral gp70s are differentially regulated in physiological vs inflammatory conditions.

FIGURE 1

Primer sequences used in RT-PCR and genomic PCR for amplification of subgroups of xenotropic and polytropic gp70s. The primer sequences in the hypervariable region A (VRA) specific for the four subgroups of xenotropic gp70s (A and B) and in a proline-rich domain (PRD) specific for PT and mPT gp70s (C) are underlined. R: A/G; S: C/G; Y: T/C.

FIGURE 2

Predicted amino-acid sequences of different retroviral gp70s. Nucleotide sequence analysis of cDNA prepared from two different NZB-X1 (Clone 35 and NZB 179) viruses revealed that both NZB-X1 and NZB-X2 gp70s belong to the Xeno-I subgroup, but their predicted amino acid sequences differ by three amino-acid residues at the C-terminus, which are highlighted in bold. The GenBank accession number for NZB-X1 gp70 cDNA sequence is EU334447. NZB-X2, PT and mPT gp70 sequences are derived from NZB-9-1, MX27 and MX33 proviruses, respectively (26, 37), and Xeno-II, Xeno-III and Xeno-IV gp70 sequences from RP24-240L12, RP24-114A21 and RP23-110C17 BAC clones, respectively. The hypervariable regions A and B (VRA and VRB) and a proline-rich domain (PRD) are shaded. Identities are indicated by dashes. Numbers indicate amino-acid positions.

FIGURE 3

RT-PCR and genomic PCR analyses for four subgroups of xenotropic gp70 mRNAs and proviruses in different strains of mice. (A) The presence of Xeno-I, Xeno-II, Xeno-III and Xeno-IV gp70 mRNAs in liver from male mice of different strains was determined by RT-PCR with a forward primer covering the tRNA primer-binding site and reverse primers specific for the four different subgroups of xenotropic viruses (Fig. 1A). Since the reverse primer designed for the amplification of Xeno-III gp70 also amplifies Xeno-II gp70, the results obtained with this primer are indicated as Xeno-II/III. (B) Genomic DNA from different strains of female (F) and male (M) mice was analyzed for the presence of Xeno-I, Xeno-II, Xeno-III and Xeno-IV gp70 sequences by PCR with a common forward primer (Xeno277F) and the same reverse primers specific for the four different subgroups of xenotropic viruses used for the detection of gp70 mRNAs.