The nucleotide sequences of wild-type coxsackievirus A9 strains imply that an RGD motif in VP1 is functionally significant

Abstract

We have shown previously that, compared to other enteroviruses, the coxsackievirus A9 (CAV-9) prototype strain, Griggs, contains a C-terminal extension to the capsid protein VP1 and that within this extension there is an RGD (arginine-glycine-aspartic acid) motif. To determine whether these features are found in other CAV-9 strains and therefore analyse whether they are likely to be functionally important, we have determined the nucleotide sequence of the appropriate region from five strains, isolated over a 25 year period. The results indicate that there is considerable diversity between the strains and there is little correlation between nucleotide sequence identity and date of isolation. All isolates exhibit the VP 1 extension and although its amino acid sequence is otherwise variable, the RGD motif is common to all. This conservation of sequence, within a region which can otherwise vary, implies that the RGD sequence must be functionally significant. The VP1 extension shows similarity to sequences found in footand-mouth-disease virus strains and to part of the precursor of the cellular protein, human transforming growth factor fl, and the possible significance of these observations is discussed.

Molecular analysis of human parechovirus type 2 (formerly echovirus 23).

“…No L protein appears to exist in the parechoviruses, as sequences close to the predicted N terminus of the polyprotein have been observed in the VP0 protein purified from virus particles (Stanway et al, 1994). This contradicts the original prediction of a short L, based on the presence in HPeV1 of a consensus sequence for myristoylation 12 amino acids downstream of the initiation codon and the observation of myristoylation of VP0 in most picornaviruses (Chow et al, 1987 ;Hyypia$ et al, 1992). A potential myristoylation site also exists in HPeV2, but this has the sequence GXXXT, while the sequence in other picornaviruses that show this modification is GXXXS.…”

“…One µg HPeV2 RNA was reverse transcribed using an oligo(dT) "( primer and aliquots were subjected to PCR amplification, with primers based either on the limited HPeV2 sequence available or on the genomic HPeV1 sequence (Hyypia$ et al, 1992 ;Stanway et al, 1994). The procedure was as previously described (Gama et al, 1989).…”

“…In HPeV1, two of the polyprotein cleavage sites (VP0\VP3, VP3\VP1) have been identified directly by amino acid sequence determination of purified capsid proteins, and the others were predicted by protein alignments (Hyypia$ et al, 1992 ;Stanway et al, 1994). The sequences flanking the probable cleavage sites of HPeV1 and HPeV2 are shown in Table 2.…”

“…The primers used were based on previously obtained sequences from HPeV2 cDNA clones covering the VP1 region (Stanway et al, 1994), part of the 5h UTR and part of 2BC, and on the published HPeV1 sequence (Hyypia$ et al, 1992). Previous attempts to obtain cDNA clones spanning the first 72 nucleotides of the HPeV1 and HPeV2 genomes had proved unsuccessful and the published HPeV1 sequence for this region was obtained by sequencing the RNA genome directly using reverse transcriptase (Hyypia$ et al, 1992). This region from both HPeV1 and HPeV2 was successfully cloned and sequenced by heating the RNA to 95 mC (to denature secondary structure) prior to reverse transcription, then amplifying by PCR with a primer based on nucleotides 1-16 of HPeV1 RNA.…”

“…A processing cascade, brought about by virus-encoded proteases, gives a number of functionally important precursors and the individual proteins. These include the structural proteins, encoded by the 5h-proximal region of the open reading frame, and non-F. Ghazi and others F. Ghazi and others resulting from the lack of cleavage of VP0 into VP4 and VP2 (Hyypia$ et al, 1992 ;Stanway et al, 1994). Partial sequence analysis of another picornavirus, echovirus 23, suggested that it is a close relative of HPeV1 (Stanway et al, 1994), and it has now been renamed human parechovirus type 2 (HPeV2).…”