Conserved motifs in the invertebrate iridescent virus 6 (IIV6) genome regulate virus transcription.

TLDR: The transcriptional class of all IIV6 genes that had not been classified until now is investigated and single nucleotide mutations in the highly conserved nucleotides at the end of the second motif showed that this motif acted as a repressor sequence for late genes in the IIV 6 genome.

About: This article is published in Journal of Invertebrate Pathology.The article was published on 2020-11-01 and is currently open access. It has received 0 citations till now.

Figures

Fig. 2. Delayed-early (DE) and late (L) gene transcripts of IIV6. Cells were infected with IIV6 in the presence of DNA (Ara-C) or protein synthesis (CHX) inhibitors. ORF-specific RT-PCR was carried out on total RNA isolated at 12 h post infection. Transcripts that were not detected in the presence of the protein synthesis inhibitor (CHX) but were not affected by the DNA synthesis inhibitor are from DE genes (A). Genes that do not show transcripts in the presence of both inhibitors as categorized as L genes (B). ORF: open reading frame; Ara-C: DNA synthesis inhibitor (cytosine arabinoside); CHX: protein synthesis inhibitor (cycloheximide). PC: Positive control, infection without addition of inhibitors.

Fig. 5. Detection and analysis of additional upstream motifs for late genes. The position of the four identified motifs in the upstream region exemplified for ORF 061R. The mutated motifs are indicated with yellow boxes. The mutations made in these motifs are illustrated in red text. Luciferase activities of deletion and site directed-mutated promoter plasmids were compared to the wild type sequence for 061R. RLU/sec; relative light units emitted per second.

Table 1 Primers for the promotor analyses.

Fig. 4. Luciferase activities of promoter reporter plasmids of immediate-early (193R) (A), delayed-early (126R) (B) and late (259R, 084L, 061R) (C) ORFs with and without the motifs shown in Fig. 3. The prom and del extensions in the plasmid names refer to plasmids with and without the motifs, respectively. RLU/sec; relative light units emitted per second.

Fig. 3. Predicted motifs within the upstream sequences of IIV6 ORFs. Genes were categorized in temporal classes based on the experimental and literature data. Conserved motifs were searched for per temporal class by analysing the 200-nt sequences upstream of the annotated IIV6 translation start sites using MEME software. Parameters were set to zero or one occurrence per sequence and the program searched the coding strand only. Single motifs were obtained for (A) immediate early (IE) and (B) delayed early (DE) genes, while late (L) genes (C and D) contained two motifs (L1) and L2).

Fig. 1. Immediate-early (IE) gene transcripts of IIV6. Cells were infected with IIV6 in the presence of DNA (Ara-C) or protein synthesis (CHX) inhibitors. ORF-specific RT-PCR was carried out on total RNA isolated at 12 h post infection. Genes that give a positive RT-PCR signal in the presence of these inhibitors are categorized as IE genes. ORF: open reading frame; Ara-C: DNA synthesis inhibitor (cytosine arabinoside); CHX: protein synthesis inhibitor (cycloheximide). PC: Positive control, infection without addition of inhibitors.

Frequently asked questions

Q1. What are the contributions in "Conserved motifs in the invertebrate iridescent virus 6 (iiv6) genome regulate virus transcription" ?

In this study, the authors investigated the transcriptional class of all IIV6 genes that had not been classified until now. Conversely, the presence of these two sequences upstream of the reporter decreased its expression. Next, upstream sequences of IIV6 L genes from which the authors removed this second motif in silico, were re-analyzed for the presence of potential conserved promoter sequences.