Significance: Based on several studies, including ours, we estimate that 40–70 million individuals carry a chromosomally integrated copy of the human herpesvirus 6 (HHV-6) genome in every cell of their body. This condition is referred to as inherited chromosomally integrated HHV-6 (iciHHV-6). The regions targeted for integration are telomeres, which play important roles in the self-renewal capacity of cells. Whether iciHHV-6 is associated with disease remains unknown. After conducting a population screen ( n = 20,000), our results indicate that the prevalence of angina is three times greater in iciHHV-6+ subjects relative to iciHHV-6− ones. Furthermore, iciHHV-6+ subjects have shorter telomeres, a result that may explain, at least in part, how iciHHV-6 may contribute to the development of angina.

The majority of human herpesvirus 6 (HHV-6) congenital infections (86%) originate from germ line transmission of chromosomally integrated HHV-6 (ciHHV-6). To determine whether transplacentally acquired HHV-6 could derive from the transmission of reactivated maternal ciHHV-6, we identified mother-infant pairs in which infants had proven transplacentally acquired HHV-6 and mothers had documented ciHHV-6, and we sequenced and compared the HHV-6 gB gene sequences for each pair. Our data indicate that the gB gene sequence found in each cord blood specimen was identical to that of the corresponding mother but divergent from that of other known HHV-6 isolates. These results are consistent with transplacentally acquired HHV-6 originating from the transmission of reactivated ciHHV-6.

Human herpesvirus 6 (HHV-6) is a ubiquitous virus with which infections have been associated with pathologies ranging from delayed bone marrow engraftment to a variety of neurological diseases. The lack of a standardized assay that can be used to detect and estimate HHV-6 DNA contents in various clinical specimens can lead and has led to discordant results among investigators and on the potential association of HHV-6 to diseases. To identify the most reliable and sensitive assays, an identical set of 11 coded serum samples spiked with various quantities of the HHV-6A variant (range, 4 to 400,000 genome copies/ml) was sent to eight independent laboratories around the world. Each laboratory was asked to estimate the HHV-6 DNA content by use of its own protocols and assays. Among the various assays, three TaqMan-based real-time PCR assays yielded quantities that were closest to the quantity of HHV-6 that had been spiked. To provide better homogeneity between the results from the different laboratories working on HHV-6, we propose that investigators interested in quantifying HHV-6 in clinical samples adopt one of these assays.

Human herpesvirus 6A (HHV-6A) and 6B (HHV-6B) are ubiquitous betaherpesviruses that infects humans within the first years of life and establishes latency in various cell types. Both viruses can integrate their genomes into telomeres of host chromosomes in latently infected cells. The molecular mechanism of viral integration remains elusive. Intriguingly, HHV-6A, HHV-6B and several other herpesviruses harbor arrays of telomeric repeats (TMR) identical to human telomere sequences at the ends of their genomes. The HHV-6A and HHV-6B genomes harbor two TMR arrays, the perfect TMR (pTMR) and the imperfect TMR (impTMR). To determine if the TMR are involved in virus integration, we deleted both pTMR and impTMR in the HHV-6A genome. Upon reconstitution, the TMR mutant virus replicated comparable to wild type (wt) virus, indicating that the TMR are not essential for HHV-6A replication. To assess the integration properties of the recombinant viruses, we established an in vitro integration system that allows assessment of integration efficiency and genome maintenance in latently infected cells. Integration of HHV-6A was severely impaired in the absence of the TMR and the virus genome was lost rapidly, suggesting that integration is crucial for the maintenance of the virus genome. Individual deletion of the pTMR and impTMR revealed that the pTMR play the major role in HHV-6A integration, whereas the impTMR only make a minor contribution, allowing us to establish a model for HHV-6A integration. Taken together, our data shows that the HHV-6A TMR are dispensable for virus replication, but are crucial for integration and maintenance of the virus genome in latently infected cells. Herpesviruses are ubiquitous pathogens that persist in the host for life. Two human herpesviruses (HHV-6A and HHV-6B) can integrate their genetic material into the telomeres of host chromosomes. Integration also occurs in germ cells, resulting in individuals that harbor the virus in every single cells of their body and transmit it to their offspring, a condition that affects about 1% of the human population. We set to elucidate the integration mechanism that allows these viruses to maintain their genome in infected cells. Intriguingly, HHV-6A, HHV-6B and several other herpesviruses harbor telomere sequences at the end of their genome. Removal of these sequences in the genome of HHV-6A revealed that the viral telomeres are crucial for the integration of this human herpesvirus. In addition, we demonstrate that the telomere sequences at the right and left end of the virus genome play different roles in the integration process. Taken together, our data sheds light on the integration mechanism that allows HHV-6A to integrate into somatic cells and to enter into the germ line.

Integrons are genetic elements that are able to capture genes by a site-specific recombination mechanism. Integrons contain a gene coding for a λ-like integrase that carries out site-specific recombination by interacting with two different target sites; the attI site and the palindromic sequence attC (59 base element). Cassette integrations usually involve the attI site, while cassette excisions use attC. Therefore, the integrase should bind both sites to cleave DNA and perform site-specific recombination reactions. We have used purified maltose-binding protein fused with the integrase (MBP-IntI1) and native IntI1 protein and gel retardation assays with fragments containing the complete and partial attI1 site to show formation of four complexes in this region. Chemical modification of specific nucleotides within the attI1 site was used to investigate their interference with binding of the integrase protein. We attribute IntI1 specific binding to four regions in the attI1 site and a GTTA consensus sequence is found in three of the four regions. Interference by modified guanine and thymine residues in the DNA major groove and adenine residues in the minor groove were observed, indicating that the integrase interacts with both sides of the helix. Binding of IntI1 to attC is also discussed.

Human herpesviruses (HHV) are stealth pathogens possessing several decoy or immune system evasion mechanisms favoring their persistence within the infected host. Of these viruses, HHV-6 is among the most successful human parasites, establishing lifelong infections in nearly 100% of individuals around the world. To better understand this host-pathogen relationship, we determined whether HHV-6 could interfere with the development of the innate antiviral response by affecting interferon (IFN) biosynthesis. Using inducible cell lines and transient transfection assays, we have identified the immediate-early 1 (IE1) protein as a potent inhibitor of IFN-β gene expression. IE1 proteins from both HHV-6 variants were capable of suppressing IFN-β gene induction. IE1 prevents IFN-β gene expression triggered by Sendai virus infection, double-stranded RNA (dsRNA) and dsDNA transfection, or the ectopic expression of IFN-β gene activators such as retinoic inducible gene I protein, mitochondrial antiviral signaling protein, TBK-1, IκB kinase ε (IKKε), and IFN regulatory factor 3 (IRF3). While the stability of IFN-β mRNA is not affected, IE1-expressing cells have reduced levels of dimerized IRF3 and nucleus-translocated IRF3 in response to activation by TBK-1 or IKKε. Using nuclear extracts and gel shift experiments, we could demonstrate that in the presence of IE1, IRF3 does not bind efficiently to the IFN-β promoter sequence. Overall, these results indicate that the IE1 protein of HHV-6, one of the first viral proteins synthesized upon viral entry, is a potent suppressor of IFN-β gene induction and likely contributes to favor the establishment of and successful infection of cells with this virus.