Human endogenous retrovirus (HERV) sequences comprise approximately 8% of human DNA (5
). Although almost all HERV genomes appear to lack intact open reading frames (ORFs) and are therefore likely defective in replication, some of them, such as HERV-K113, have complete ORFs for all viral proteins (52
). Nonetheless, HERV-K113 is poorly expressed and is not capable of replication (6
). Recently, using bioinformatics approaches, two groups reconstructed infectious HERV-K sequences (18
). In one approach, a consensus sequence of HERV-K113 and closely related well-preserved HERV-Ks was determined and termed HERV-KCON
). HERV-K molecular clones and their derivatives that encode these reconstructed sequences have become powerful tools to examine the biology of these ancient retroviruses.
While HERV-K is categorized as a betaretrovirus because of its sequence similarity to mouse mammary tumor virus (MMTV), in contrast to other betaretroviruses, which assemble particles in the cytoplasm (type B/D), HERV-K forms particles at the plasma membrane (PM) (type C), like human immunodeficiency virus type 1 (HIV-1) and murine leukemia virus (MLV) (8
). Particle formation of retroviruses is driven by the precursor polyprotein Gag. HIV-1 Gag consists of four major domains, matrix (MA), capsid (CA), nucleocapsid (NC), and p6, as well as two spacer peptides, SP1 and SP2 (4
). Using HERV-KCON
) and another HERV-K113 derivative (termed oricoHERV-K113) in which 5 postinsertion mutations were reverted (21
), recent studies unambiguously determined the domain organization of HERV-K Gag (21
). HERV-K Gag also has four domains, MA, CA, NC, p15, as well as three short peptide sequences, SP1, QP1, and QP2 (21
). In HIV-1, MA is required for Gag targeting and binding to the PM. CA and NC domains are essential for Gag multimerization. The p6 domain contains a late domain motif, Pro-Thr-Ala-Pro (PTAP), which recruits the cellular ESCRT complexes that facilitate release of virions. These domains give rise to individual mature Gag proteins upon proteolytic cleavage mediated by viral protease, which occurs during or immediately after virus particle release. The functions of these domains are less well understood for HERV-K Gag than in HIV-1 Gag, although some functional motifs are shared by both Gag proteins (21
). Notably, a single postinsertion mutation, which was corrected in both HERV-KCON
and oricoHERV-K113, is responsible for the defect in assembly of HERV-K113 (26
). Thus, these restored HERV-K113 sequences serve as good models for studying assembly of HERV-K Gag.
Since all human cells harbor HERV-K genomes and potentially express HERV-K proteins, HERV-K or some of its components might be coexpressed with HIV-1 in the same cell. In this regard, it is notable that a number of studies showed a correlation between increased expression of HERV-K and HIV-1 infection. Several groups have detected antibody responses to HERV-K in a majority of HIV-1-positive patients but not in uninfected donors (41
). Similarly, T cell responses to HERV-K were observed in HIV-1-infected patients but not in healthy donors (19
). Furthermore, upregulation of HERV-K RNA was detected (14
) in plasma samples of HIV-1-infected individuals, in HIV-1-infected T cells (17
), and in HIV-1 Tat-transfected T cells (23
). In addition to RNA, HERV-K Gag proteins were also observed to increase upon HIV-1 infection (15
) and HIV-1 Tat transfection (23
). These reports collectively suggest that HIV-1 infection enhances the HERV-K expression in T cells. Thus, it is possible that HERV-K Gag induced by HIV-1 infection coexists with HIV-1 Gag in the same host cells. However, the impact of such coexpression on HIV-1 replication is unknown.
Coinfection and coexistence of two different lentiviruses in the same host have been observed naturally (2
). In phylogenetic analysis, some of the primate lentiviruses were found to be recombinants of two distinct parental viruses, which must have arisen from coinfection of these two viruses in a single cell (3
). In cells coinfected with two such distinct parental viruses, two different Gag proteins would be expressed in the same cells, which raises a possibility that different but related Gag proteins coassemble into the same virions. Indeed, in cells coexpressing both HIV-1 and HIV-2, HIV-1 and HIV-2 Gag proteins colocalized at the PM and coassembled into the same virions (10
). Gag proteins of more distantly related retroviruses also coassemble when modified by addition of a heterologous membrane binding motif (7
) or exchange of CA (1
). However, coassembly between native Gag proteins of retroviruses of different genera, such as HIV-1 and HERV-K, has not been observed. Because diverse retroviruses rely on similar host factors, such as phosphatidylinositol (4,5)-bisphosphate [PI(4,5)P2
] and ESCRT proteins, consequences that arise from coexpression of HIV-1 and HERV-K Gag proteins in the same cell might rather be competition for cellular cofactors and inhibition of assembly of one virus.
We hypothesized that if expression of HERV-K Gag is induced by HIV-1 infection, the HERV-K Gag might compete with HIV-1 Gag in the same host cell. In this study, we report that HERV-K Gag indeed inhibited the HIV-1 release efficiency and infectivity when Gag from HERV-KCON was coexpressed with HIV-1. Unexpectedly, HERV-K Gag colocalized with HIV-1 Gag at the PM and coassembled into the same virion. We found that both membrane binding and NC-mediated RNA binding of Gag are required for coassembly between HIV-1 and HERV-K Gag as well as for inhibition of progeny virus release and infectivity. To our knowledge, this is the first example in which a retroviral Gag coassembles with, and inhibits assembly of, another retrovirus Gag protein coexpressed in the same cells.