Characterization of human telomeric repeat sequences from human herpesvirus 6 and relationship to replication

Abstract

Here we examine by polymerase chain reaction amplification followed by cloning and sequence analyses selected regions of the human herpesvirus 6 (HHV-6) genome which contain human telomeric repeats (TTA-GGG). We determine the relative number, arrangement and orientation of the repeats in the unit length genome, in concatemeric replicative intermediates and in heterogeneous (het) regions. We also examine distribution of the repeats in the entire genome (159 kb) and their orientation relative to DNA packaging motifs and the origin of lytic replication. In the prototype orientation the HHV-6 repeat is the related complement, TAACCC. We show that tandem arrays of this repeat are present in the right and left long direct repeats (DR L and DR R, 8 kb each) which bound the long unique sequence (U L, 143 kb). Within each DR there is a left terminal imperfect tandem array and a right terminal perfect tandem array (58 copies). In DR they are each adjacent to DNA packaging motifs, pacl and pac2, described for herpes simplex virus and human cytomegalovirus, in the arrangement pacl-imperfect repeat-7.2 kb-perfect repeat-pac2. Five independent clones were isolated and sequence determined from junctions of concatemeric replicative intermediates which showed adjacent pac2 and pacl motifs surrounded by telomeric repeats. Favoured cleavage sites for unit length genomes were indicated which avoided cleavage within the repeats. Analyses of the complete genome showed no tandem repeats within U L but did show a polar distribution of monomeric copies and related sequences around the origin of replication, with an effect on the overall base composition. The implications for virus replication are discussed.Human herpesvirus 6 (HHV-6) is one of the most recently characterized of the family of seven human herpesviruses. Like other herpesviruses it can establish a latent or persistent infection which remains for the lifetime of the host and can reactivate during immunosuppression. HHV-6 infects up to 90 % of the population as infants where infection is asymptomatic or causes exanthem subitum, a mild skin rash (Yamanishi et al., 1988;Okuno et al., 1989). The virus has a cellular tropism for CD4 + T lymphocytes and there is evidence for the monocyte as a possible site for latency. HHV-6 strains were first isolated from blood samples from AIDS patients where the virus had reactivated (Salahuddin et al., 1986;Downing et al., 1987). Given the similar cellular tropisms of human immunodeficiency virus

Detection of channel catfish virus DNA in latently infected catfish.

“…The unusual nature of these junctional clones is not understood. However, previous studies investigating the DNA structure of human herpesvirus type 6 (HHV-6) in infected cells also describe aberrant junctional clones generated by PCR amplification of the HHV-6 DR R -to-DR L junction (Gompels & Macaulay, 1995 ;Thomson et al, 1994). The studies offer no explanation for the anomalous junctions other than attributing them to possible PCR artefacts.…”

The DR1 and DR6 First Exons of Human Herpesvirus 6A Are Not Required for Virus Replication in Culture and Are Deleted in Virus Stocks That Replicate Well in T-Cell Lines

“…The unit-length DNA molecules are approximately 160 kb, composed of a 143-kb unique (U) segment flanked by left and right direct repeats (DR L and DR R , respectively) (19,24,27,46). The DRs are of sizes 8 to 10 kb in different viral isolates (2,19,24,46).…”

Detection and application of circular (concatemeric) DNA as an indicator of koi herpesvirus infection

“…On the other hand, the DR junction of CCV contained 11 to 17 bp of the right terminus of UL sequences following the DR R region (Gray et al 1999). While previous studies attributed the existence of nucleotides between DR regions to PCR artifacts (Thomson et al 1994, Gompels & Macaulay 1995, Gray et al 1999, our sequence analysis of the DR-PCR amplicon in the timecourse experiment showed that the same nucleotide sequences were present in all of the sequenced samples (data not shown). Further investigation is therefore necessary to examine the configuration of the DR junction of KHV.…”

“…3). In the case of HHV-6 which has a 10 kbp DR L and DR R region at the end of each genome terminus , the DR junction was sequenced using a PCR product, and aberrant junction sequences due to several nucleotide insertions or deletions were observed (Thomson et al 1994, Gompels & Macaulay 1995. On the other hand, the DR junction of CCV contained 11 to 17 bp of the right terminus of UL sequences following the DR R region (Gray et al 1999).…”

“…This genome structure is the same as that of human herpesvirus 6 (HHV-6), which forms a long continuous DNA molecule, or head-to-tail concatemer, containing multiple copies of the viral genome (Thomson et al 1994, Gompels & Macaulay 1995. CCV has 18.5 kbp DR L and DR R regions at each of the genome termini, and the intracellular genome has an 'endless' (circular or concatemer) configuration in acutely infected brown bullhead cells (Cebrian et al 1983).…”