Genomic Flexibility of Human Endogenous Retrovirus Type K

Abstract

Human endogenous retrovirus type K (HERV-K) proviruses are scattered throughout the human genome, but as no infectious HERV-K virus has been detected to date, the mechanism by which these viruses replicated and populated the genome remains unresolved. Here, we provide evidence that, in addition to the RNA genomes that canonical retroviruses package, modern HERV-K viruses can contain reverse-transcribed DNA (RT-DNA) genomes. Indeed, reverse transcription of genomic HERV-K RNA into the DNA form is able to occur in three distinct times and locations: (i) in the virus-producing cell prior to viral release, yielding a DNA-containing extracellular virus particle similar to the spumaviruses; (ii) within the extracellular virus particle itself, transitioning from an RNA-containing particle to a DNA-containing particle; and (iii) after entry of the RNA-containing virus into the target cell, similar to canonical retroviruses, such as murine leukemia virus and HIV. Moreover, using a resuscitated HERV-K virus construct, we show that both viruses with RNA genomes and viruses with DNA genomes are capable of infecting target cells. This high level of genomic flexibility historically could have permitted these viruses to replicate in various host cell environments, potentially assisting in their many integration events and resulting in their high prevalence in the human genome. Moreover, the ability of modern HERV-K viruses to proceed through reverse transcription and package RT-DNA genomes suggests a higher level of replication competency than was previously understood, and it may be relevant in HERV-Kassociated human diseases. IMPORTANCERetroviral elements comprise at least 8% of the human genome. Of all the endogenous retroviruses, HERV-K viruses are the most intact and biologically active. While a modern infectious HERV-K has yet to be found, HERV-K activation has been associated with cancers, autoimmune diseases, and HIV-1 infection. Thus, determining how this virus family became such a prevalent member of our genome and what it is capable of in its current form are of the utmost importance. Here, we provide evidence that HERV-K viruses currently found in the human genome are able to proceed through reverse transcription and historically utilized a life cycle with a surprising degree of genomic flexibility in which both RNA-and DNA-containing viruses were capable of mediating infection.

Susceptibility of Human Endogenous Retrovirus Type K to Reverse Transcriptase Inhibitors

“…We previously showed that the retroviral genome of HK2 virus particles is made up of RNA and recently reverse-transcribed cDNA (rt-cDNA) (53). Therefore, we measured the levels of viral RNA/cDNA in cell-free HK2 viral particles by RT-PCR, which can measure the levels of both viral RNA and rt-cDNA.…”

“…We previously observed that HK2 RNA transcripts could be reverse transcribed intracellularly as well as in viral particles as noted above (53). We therefore measured the amount of HK2 env DNA in NCCIT cells after treatment with AZT.…”

“…The reverse transcription step of HK2 bears some resemblance to that of spumaviruses, a genus of the retroviridae family, which replicates through an infectious DNA form (52). Viral RNA genomes transcribed from HK2 proviruses can be reverse transcribed into cDNA within the cells, in alreadyreleased VLPs, and/or after particle infection (53). We previously noted that unlike conventional retroviruses, which after infection are integrated into the host genome, contemporary HK2 cDNAs autointegrate into 1-long terminal repeat (1-LTR) and 2-long terminal repeat (2-LTR) episomes (51).…”

Human Endogenous Retrovirus Type K (HERV-K) Particles Package and Transmit HERV-K–Related Sequences

“…Although the Bogota probe seems to favor episomal replication when transmitted in an autonomous form, infection and retroviral integration of HK2 into the genome of an infected cell has been demonstrated using the resurrected HK2 viruses HERV-K CON and Phoenix (53,54), as well as the autointegration of HERV-K CON , to form episomes (59). Thus, HK2 is, in principle, able to integrate into the genome of human cells, and it is surprising that the majority of HK2 viruses led to transmissible viral genomes that resulted in circular forms rather than genomic integrations.…”

“…Subsequently, two groups resurrected infectious viruses using existing HK2 fragments of the human genome (52,53); molecular recombination among present-day HK2 proviruses produced viruses competent to replicate, providing proof of principle of the possibility of HK2 replication in ancient and perhaps modern humans (53). Our laboratory has recently demonstrated that HK2 viral genomes found in HK2 particles are able to proceed through reverse transcription and have utilized a life cycle with a degree of genomic flexibility in which both RNA-and DNA-containing viruses are capable of mediating infection (54). Finally, we looked for evidence of HK2 transmission in modern humans by analyzing the viral genome found in viruses produced by living patients (9).…”