Cloning, Characterization, and Phylogenetic Analysis of a Shrimp White Spot Syndrome Virus Gene That Encodes a Protein Kinase

Abstract

An open reading frame (ORF) that encodes a 715-amino-acid polypeptide was found in an 8421-bp EcoRI fragment of the shrimp white spot syndrome virus (WSSV) genome. The polypeptide shows significant homology to eukaryotic serine/threonine protein kinase (PK) and contains the major conserved subdomains for eukaryotic protein kinases. Coupled in vitro transcription and translation generated a protein having an apparent molecular mass of about 87 kDa according to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. For transcriptional analysis of the pk gene, total RNA was isolated from WSSV-infected shrimp at different times after infection. Northern blot analysis with pk-specific riboprobe found a major and a minor transcript of 2.7 and 5.7 kb, respectively. Rapid amplification of the 5' cDNA ends of the major 2.7-kb pk transcript showed that there were two transcriptional initiation sites located at nucleotide residues -38(G) and -39(G) relative to the ATG translational start codon. Temporal expression analysis by RT-PCR indicated that the transcription of the pk gene started 2 h after infection and continued for at least 60 h. Phylogenetic analysis showed that WSSV protein kinase does not have any close relatives and does not fall into any of the major protein kinase groups.

Transcriptional analysis of the white spot syndrome virus major virion protein genes

“…The earlier onset of transcription of vp15 (Fig. 1) could be regulated by this TATA-box, as in other WSSV genes which have been shown or are likely to be expressed early in infection (rr1, rr2, pk and dnapol) (Tsai et al, 2000a;Liu et al, 2001;Chen et al, 2002). The transcriptional start and stop sites of the vps were determined by 59 and 39 RACE.…”

“…Probably part of the 59 end of the mRNAs is missing in these studies , because intact 59 ends of mRNA are rarely recovered from cDNA libraries. Polyadenylation of the WSSV genes rr1, rr2, pk and dnapol is, respectively, 12, 13, 16 and 17 nt downstream of the polyadenylation signal (Liu et al, 2001;Tsai et al, 2000a;Chen et al, 2002). All identified WSSV polyadenylation sites are within the range of regular polyadenylation in eukaryotic mRNAs, which is typically located 15 to 25 nt downstream of the sequence AAUAAA (Fitzgerald & Shenk, 1981).…”

“…Transcriptional analysis has been performed on WSSV genes encoding the large and small subunit of ribonucleotide reductase (rr), protein kinase (pk), the chimeric thymidine kinase-thymidylate kinase (tk-tmk) and DNA polymerase (dnapol). These genes were identified in P. monodon 2 or 4 h post-WSSV infection (Liu et al, 2001;Tsai et al, 2000a, b;Chen et al, 2002), indicating that these genes are of the early type. The transcription of rr1, rr2, pk and dnapol is initiated 20-28 nt downstream of a TATA-box, suggesting a functional role for the TATA-box during early transcription.…”

CHARACTERISTICS OF VIRAL PROTEIN, VP-15, OF WHITESPOT SYNDROME VIRUS ISOLATED FROM INFECTED TIGER SHRIMP Penaeus monodon (Fabricius, 1798)

“…However, the complete sequence of WSSV has recently been reported by Van-Hulten et al (2002). Marks et al (2003) reported that a VP-15 was the only major protein structure gene with a consensus TATA-box compared to VP-19, VP-24, and VP-28, in which the earlier onset of transcription of VP-15 could be regulated by this TATA-box, as in other WSSV genes which have been shown or are likely to be expressed early in an infection (Tsai et al, 2000;Liu et al, 2001;Chen et al, 2002). Van-Hulten et al (2002) stated that VP-15 is a very basic protein and resembles histone protein which functions as a DNA-binding protein in the WSSV nucleocapsid.…”

A phage-displayed peptide can inhibit infection by white spot syndrome virus of shrimp

“…In 2000, two major structural protein genes of WSSV (VP28 and VP26) were identified (van Hulten et al, 2000a), followed by genes for ribonucleotide reductase, endonuclease, protein kinase and other structural proteins (van Hulten et al, 2000b(van Hulten et al, , 2002Witteveldt et al, 2001;Liu et al, 2001;Chen et al, 2002;Zhang et al, 2002). In 2001, the entire 305 kb genome sequence was reported Yang et al, 2001).…”