P64, a Novel Major Virion DNA-Binding Protein Potentially Involved in Condensing the Spodoptera frugiperda Ascovirus 1a Genome

Abstract

We recently identified 21 structural proteins in the virion of Spodoptera frugiperda ascovirus 1a (SfAV1a), a virus with a large, double-stranded DNA genome of 157 kbp, which attacks species of the lepidopteran family Noctuidae. The two most abundant virion proteins were the major capsid protein and a novel protein (P64) of 64 kDa that contained two distinct domains not known previously to occur together. The amino-terminal half of P64 (residues 1 to 263) contained four repeats (a recently recognized motif with an unknown function) of a virus-specific two-cysteine adaptor. Adjoined to this, the carboxy-terminal half of P64 (residues 279 to 455) contained 14 copies of a highly basic, tandemly repeated motif rich in arginine and serine, having an 11-to 13-amino-acid consensus sequence, SPSQRRSTS(V/K)(A/S)RR, yielding a predicted isoelectric point of 12.2 for this protein. In the present study, we demonstrate by Southwestern analysis that SfAV1a P64 was the only virion structural protein that bound DNA. Additional electrophoretic mobility shift assays showed that P64 bound SfAV1a as well as non-SfAV1a DNA. Furthermore, we show through immunogold labeling of ultrathin sections that P64 is a component of virogenic stroma and appears to be progressively incorporated into the SfAV1a DNA core during virion assembly. As no other virion structural protein bound DNA and no basic DNA-binding proteins of lower mass are encoded by the SfAV1a genome or were identified by proteomic analysis, our results suggest that P64's function is to condense the large genome of this virus and assist in packaging this genome into its virion.

Proteomic analysis of Chilo iridescent virus

“…The gene products of six of the eleven SfAV1a homologs were also found in the proteome of SfAV1a virions (Tan et al, 2009a). A homolog of the SfAV1a virion protein P64, which was recently shown to be a major DNA binding protein with proposed DNA condensing activity (Tan et al, 2009b) is not encoded in the CIV genome.…”

Characterization of Heliothis Virescens Ascovirus 3h orf21 that Encodes a Virion Protein

“…The Using an anti-3H-21 antiserum as the primary antibody (diluted 1:5000) for 1 h at room temperature, followed by alkaline phosphatase conjugated anti-rabbit antibodies (1:5000). Further treatments as described previously [14] . Lysates of mock-infected third-instar H. armigera larvae were used as a negative control, and infected with HvAV-3h served as a positive control.…”

“…And used for the evolutional study of several related dsDNA viruses [15] . P64 is another most abundant protein in SfAV-1a as a structural protein required for packaging of the large viral genome into the virion during assembly [14] . An intriguing finding suggests that P64 interacting with other structural proteins as a unit could condense and release gDNA in the virions [26] .…”

“…The Ascoviridae family has been divided into two genera: Ascovirus, which infects various members of the family Noctuidae (Insecta: Lepidoptera), and Toursvirus, which replicates in both its lepidopteran and parasitoid vector hosts [5] . Eleven isolates of ascoviruses have been completely sequenced to date [1,2,[6][7][8][9][10][11][12][13][14] . Further researches reported that 25 ascovirus open reading frames (ORFs) were found to be conserved in all ascoviruses except the members of Toursvirus [2] .…”

“…Further researches reported that 25 ascovirus open reading frames (ORFs) were found to be conserved in all ascoviruses except the members of Toursvirus [2] . So far, researches of major capsid protein (MCP, TnAV-2a) and DNA-binding protein (P64, SfAV-1a) and ORF 117 (3H-117, HvAV-3h) has been carried out, but almost the viral genes remain to be explored [14][15][16] .…”