Temporal Mapping of Transcripts in Herpesvirus 6 Variants

Abstract

To define the molecular features characteristic of the early stages of infection of lymphocytes with human herpesvirus 6 (HHV-6) variant A or B, we studied the temporal regulation of expression of selected sets of viral genes. Thus, U42, U94, U89-U90, U73, and U39 are α genes since their transcripts (i) were made in the presence of inhibitors of protein synthesis and (ii) were detected 3 h after infection of untreated cells. U41, U53, U31, and U19 are β genes since their expression is inhibited by cycloheximide but not by phosphonoacetate, an inhibitor of DNA synthesis. U100 is a γ gene since its spliced transcript encoding the structural glycoprotein gp82/105 was first detected 16 h after infection of untreated cells but could not be detected in cells treated with phosphonoacetate. HHV-6 variants differ in the transcription patterns of their genes. U16-U17 originates a splice transcript and is regulated as α in HHV-6B and as β in HHV-6A. U91 generates two transcripts, amplified as 476- and 374-bp PCR fragments. The 476-bp fragment is α in HHV-6A-infected cells but β in HHV-6B-infected cells. Conversely, the 374-bp fragment is β in HHV-6A-infected cells and α in HHV-6B-infected cells. Furthermore, the spliced product of U18-U19-U20 (526 bp) is β in HHV-6A-infected cells, but only a partially spliced form (1.9 kb) was detected at late stages of infection in HHV-6B. HHV-6 transcription was also studied in nonproductive lymphoid cells, and the same transcription pattern detected during lytic infection was observed. Also, HHV-6 variants maintain the differences in U91, U16-17, and U18-U19-U20. We conclude that, as expected from the sequencing data, gene expression is generally similar in HHV-6 variants. However, transcription of selected genes in HHV-6A and HHV-6B differs with respect to temporal regulation and splicing pattern. Furthermore, the identification of viral functions expressed during the different stages of lytic replication suggests that reverse transcription-PCR for HHV-6 genes is a useful diagnostic approach to differentiate between latent and productive HHV-6 infection.

Human Herpesvirus 6 and Multiple Sclerosis: A One-Year Follow-up Study

“…In 1995, a study by Challoner et al (13) found HHV-6 in the oligodendrocytes of MS patients but not controls and found a higher concentration of infected cells in areas of demyelination compared to unaffected white matter; it was the first time that HHV-6 was related to MS. Since then, a variety of studies have investigated the possible role of HHV-6 in MS with contradictory results between those which support the possible involvement of HHV-6 in MS pathogenesis (1-5, 14, 17, 19, 21, 37, 38, 42, 44, 46), and those which refute that association (31,32,41); this controversy may be interpreted in the context of the different laboratory techniques (DNA extraction, PCR method, amount of template in the reaction), different groups of MS patients, different control populations, and different study designs (transversal, longitudinal).…”

“…The primers and probes derived from previously published data (18,45) and were designed using Primer Express Software (Applied Biosystems). As it has been previously described (32), the presence of U94 transcripts in absence of expression of the other HHV-6 genes was considered HHV-6 latent infection. Meanwhile, the expression of more than one HHV-6 gene was considered active viral infection.…”

HHV‐6 infects human aortic and heart microvascular endothelial cells, increasing their ability to secrete proinflammatory chemokines

“…The results are shown in Figure 1. Transcripts of U91, U42 (immediate-early genes) [Mirandola et al, 1998] and U22, a late gene [French et al, 1999], were present at all time points. All samples were negative when amplified without previous retrotranscription (data not shown).…”

In vitro and in vivo analysis of human herpesvirus‐6 U90 protein expression

“…It was demonstrated that U90 gene transcript was produced under immediate early conditions and was detected soon after viral infection (3 hr) [Mirandola et al, 1998]. Moreover, as a sufficient level of transcript was detected not only in infected cells but also in patients' peripheral blood mononuclear cells (PBMCs) [Yoshikawa et al, 2003], it is speculated that the U90 gene product may be detected in infected cells or PBMCs collected from ES patients.…”