A viral histone H4 is encoded in a polydnavirus called Cotesia plutellae bracovirus (CpBV), which is symbiotic to an endoparasitoid wasp, C. plutellae. Compared to general histone H4, the viral H4 possesses an extra N-terminal tail containing 38 amino acid residues, which has been presumed to control host gene expression in an epigenetic mode. This study addressed the mutational analysis of extra N-terminal amino acid residues of a viral histone H4 and their epigenetic control efficacy. Mutational analysis was performed by serially deleting each of the nine amino acid residues from N-terminal tail of a viral histone H4. Transient expression of each truncated mutants (K1M-K19) in diamondback moth, Plutella xylostella, was performed by microinjection of a recombinant expression vector and confirmed by RT-PCR. Under transient expression, we analysed the effect of these mutations on target gene, transferin. Interestingly, we found that truncated mutants (K1M-K15) did not inhibit the expression of target gene but mutations thereafter (K6M-K9M) significantly alter its expression. As expected these truncated mutants (K1M-K5M) also inhibit hemocyte nodule formation and development of Plutella xylostella. This suggest that lysine residue (K6) in the N-terminal tail is very crucial for the epigenetic control efficacy of viral histone H4.

A viral histone H4 (=CpBV-H4) is encoded in a polydnavirus, Cotesia plutellae bracovirus, and symbiotically associated with an endoparasitoid wasp, C. plutellae. It has an extended N-terminal tail consisting of 38 amino acid residues, compared to the host H4 and this extended N-terminal tail has been postulated to play a crucial role in an epigenetic control of gene expression. The (SSH) suppression subtractive hybridization analysis was analyzed in transcriptome by short-read sequencing technology. The SSH analysis provided several target and nontarget genes of a viral histone H4. In this study, we analyzed the effect CpBV-H4 on the expression of two target genes serpins and histone lysine N-methyl transferase. Transient expression of CpBV-H4 by microinjecting recombinant expression vector to non parasitized larvae of Plutella xylostella showed that it was expressed up to 70 h. Under this transient expression condition, we analyzed the effect of CpBV-H4 on the expression of target genes by RT-PCR at different time points. Interestingly, the CpBV-H4 significantly inhibited the expression of target genes after 44 h, while the truncated CpBV-H4 deleting the N-terminal tail did not show the inhibitory activity.

Polydnaviruses (PDVs) are a group of insect viruses and symbiotic to some endoparasitoid wasps classified in to Braconidae and Ichneumonidae. Though a lot of PDV genes are identified and analyzed in the host-parasitoid molecular interactions, PDV replication is still far from our understanding. PDVs are replicated in the wasp ovary during late pupal stage. A PDV, Cotesia plutellae bracovirus (CpBV), is symbiotic to Cotesia plutellae. The C. Plutellae ovary was analyzed in transcriptome by 454 pyrosequencing. The ovarian transcriptome provided several major DNA polymerases including Pol α, Pol δ, and Pol ε. All contigs matched to these polymerases were expressed in C. plutellae. Especially DP1 contig homologous to Pol α was highly expressed during late pupal and female adult stages. Its RNA interference significantly suppressed CpBV viral titre in the ovary. This study suggests a hint that CpBV replication uses a host DNA polymerase, in which Pol α may play a specific role in the viral replication in the ovary.

Parasitism by an endoparasitoid wasp, Cotesia plutellae, results in significant immunosuppression of the diamondback moth, Plutella xylostella. Parasitized larvae significantly suffered higher susceptibility to a microbial biopesticide, Bacillus thuringiensis (Bt) than nonparasitized (NP) larvae. To find out an immunosuppressive agent causing the enhanced Bt efficacy, viral ankyrin (=vankyrin) genes encoded in C. plutellae bracovirus (CpBV) were analyzed by transient expression in NP larvae. CpBV segments containing different vankyrins were microinjected to NP larvae and expressed their encoded vankyrins. Expression of some vankyrins significantly inhibited immune response and enhanced Bt efficacy. This study suggests that expression of vankyrins suppress a cellular immune response and lose Bt tolerance of P. xylostella larvae.

An endoparasitoid wasp, Cotesia plutellae, contains a polydnavirus called C. plutellae bracovirus (CpBV) and induces various physiological alterations of parasitized host along with expressions of viral genes. Two host translation inhibitory factors (HTIFs) encoded in CpBV specifically inhibit host mRNAs at post-transcriptional level. They are expressed in late larval stage of Plutella xylostella parasitized by C. plutellae. To understand their late expression control, promoter region of an HTIF gene called CpBV15α was cloned by inverse PCR. The cloned HTIF upstream region (1,113 bp) possessed a putative JH response element (JHRE) and other promoter elements. The putative promoter region was rejoined with an open reading frame of enhanced green fluorescence protein (EGFP). When the recombinant vector construct was injected into early third instar larvae of nonparasitized P. xylostella, it was expressed in fourth larval instar at 72 h after injection, compared to relatively early expression in 24 h after injection of control construct containing a baculovirus immediate-early promoter. However, recombinant EGFP construct lost the late expression pattern when its promoter region was incomplete by truncating JHRE region. PYR application inhibited EGFP expression of the recombinant construct, but gave little influence on truncated constructs. Interestingly, when the complete promoter construct was injected to pupal stage, its late expression pattern was lost and showed early expression pattern. However, an addition of PYR to pupae, which had been injected with the complete promoter construct, inhibited the reporter gene expression. These results suggest that late expression of a HTIF (CpBV15α) is controlled by its promoter, which is sensitive to host JH titer.

A polydnavirus, Cotesia plutellae bracovirus (CpBV), is symbiotic to an endoparasitoid wasp, C.plutellae, which specifically parasitizes young larvae of the diamondback moth, Plutella xylostella. A recent study on CpBV replication by analysis of ovary transcriptome of C.Plutellae suggests several candidate coat protein genes. This study was conducted to confirm the coat protein genes by analyzing coat proteins of CpBV viral particles by a tandem mass MALDI-TOF. Immunoprecipitation of ovary protein extract with a polyclonal CpBV antibody captured three proteins named as p35, p60, and p70. More number of coat proteins were resolved in a protein extract directly from viral particles. All candidate coat proteins are analyzed in amino acid sequences by MALDI-TOF. A comprehensive analysis of viral proteomics and ovary transcriptome determined novel viral coat proteins from CpBV

Parasitization by an endoparasitoid wasp, Cotesia plutellae, extends a larval period of Plutella xylostella and inhibits a larva-to-pupa metamorphosis. To determine antimetamorphic parasitic factor(s) in this host-parasitoid interaction, an effect of its symbiotic polydnavirus, Cotesia plutellae bracovirus (CpBV), was investigated by injecting purified virus particles to nonparasitized larvae of P. xylostella. Larvae injected with CpBV exhibited antimetamophosis in a viral dose-dependent manner. Also, the susceptibility to the viral injection was increased at young larval stages. Parasitized or virus-injected larvae shwed significant decrease in cell size of prothoracic gland and reduction in expression of ecdysone receptor (EcR) gene. However, they increased and maintained expression of insulin receptor (InR) gene. Twenty four CpBVsegments were individually injected to nonparasitized larvae. Only two segments (S22 and S27) had significant antimetamorphic effect. Subsequent RNA interference using double stranded RNA (dsRNA) was performed in each of encoded genes in each segment. Protein tyrosine phosphatase, ELP, and three hypothetical genes were determined to be antimetamorphic factors.

A polydnavirus, Cotesia plutellae bracovirus (CpBV), possesses segmented genome located on chromosome(s) of an endoparasitoid wasp, C. plutellae. An episomal viral segment (CpBV-S3) consists of 11,017 bp encoding two putative open reading frames (ORFs). ORF301 shows amino acid sequence homologies (28~50%) with RNase T2s of various organisms. It also contains BEN domain in C-terminal region. ORF302 is a hypothetical gene, which is also found in other bracoviruses. Both genes were expressed in larvae of Plutella xylostella parasitized by C. plutellae. ORF301 and ORF302 were transiently expressed in hemocyte, fat body, gut, and epidermis of P. xylostella. To analyze effects of these genes on the parasitism, the segment of CpBV-S3 was injected to non parasitized larvae of P. xylostella, in which the two genes were expressed at least for four days post-injection. The P. xylostella larvae injected with CpBV-S3 exhibited significant immunosuppression, such as reduction in total hemocyte population, suppression of immune associated genes including cecropin, pro-phenoloxidase (PO) and serpin1, and impairment in nodule formation behavior of hemocytes in response to bacterial challenge. Each gene expression in the treated larvae was inhibited by co-injecting respective double strand RNA (dsRNA) specific to each ORF. Injection of dsRNA of ORF301 could rescue the immunosuppression by the viral segmenttreated larvae, but not by ORF302 specific dsRNA. The larval injected with CpBV-S3 exhibited an enhanced susceptibility to baculovirus infection. These results indicate that ORF301 of CpBV-S3, which containing BEN domain, suppresses both cellular and humoral immune responses in P. xylostella.

A cysteine- rich protein encoded by Cotesia plutellae bacovirus (CpBV) was identified in the parasitized Plutella xylostella. The gene, called CpBV-CRP, encodes 189 amino acids with a signal peptide of 20 residues at N-terminus determined by bioinformatic analysis, suggesting a secretory protein. High CpBV-CRP expression in the parasitized P. xylostella was observed at early days after parasitization and decreased with the course of parasitization. Expression of CpBV-CRP was tissue-specific in the fat body/epidermis, but not in hemocyte and gut. Its physiological function was analyzed by transient expression of a CpBV segment containing CpBV-CRP. The treated larvae underwent an immunosuppression in terms of hemocyte-spreading behavior. When the treated larvae were also co-injected with dsRNA against CpBV-CRP, the suppressed hemocyte behavior was significantly recovered. This study reports a cysteine-rich protein encoded in CpBV genome and its physiological function to be an immunosuppressant.

Inhibitor <SUB>K</SUB>B (I<SUB>K</SUB>B)-like gene has been found ill the genome of Cotesia plutellae bracovirus (CpBV), which is the obligatory symbiont of an endoparsitoid wasp, C. plutellae. The open reading frame of CpBV-I<SUB>K</SUB>B was 417 bp and encoded 138 amino acids. Four ankyrin repeat domains were found in CpBV-I<SUB>K</SUB>B, which shared high homology with other known polydnavirus I<SUB>K</SUB>Bs. Considering a presumptive cellular I<SUB>K</SUB>B based on Drosophila Cactus, CpBV-I<SUB>K</SUB>B exhibited a truncated structure with deletion of signal-receiving domains, which suggested its irreversible inhibitory role in NF<SUB>K</SUB>B signal transduction pathway of the parasitized host in response to the wasp parasitization. CpBV-I<SUB>K</SUB>B was expressed only in the parasitized diamondback moth, Plutella xylostella. Its expression was estimated by quantitative RT-PCR during parasitization period, showing a constitutive expression pattern from the first day of parasitization. An indirect functional analysis of CpBV-I<SUB>K</SUB>B was conducted and suggested a hypothesis of host antivirus inhibition.

폴리드나바이러스는 일부 내부기생봉의 공생 바이러스로서 기생봉 기생과정중 피기생자의 면역억제와 발육지연에 중요한 역할을 담당한다. 프루텔고치벌(Cotesia plutellae)유래 브라코바이러스(CpBV)는 피기생자의 발육을 교란시키는 주요 원인자로서 작용하는 폴리드나바이러스의 일종이다. 본 연구는 이 CpBV가 비자연기주에게도 발육교란을 유발할 수 있는 지 조사하였다. 이를 위해 프루텔고치별에 의해 기생되지 않는 비기주인 파밤나방(Spodoptera exigua)을 이용하였다. 이용된 CpBV는 프루델고치벌 난소받침(calyx)추출물에서 얻었다. 이 추출물은 CpBV항체에 대해서 뚜렷한 항원-항체반응성으로 CpBV 보유가 입증되었다. 고치벌 암컷 한 마리 CpBV 추출물을 후기 4령의 파밤나방 혈강내로 주입시켰다. CpBV 주입된 파밤나방 유충은 유충기간이 연장되고, 체중증가 가 억제되며, 궁극적으로 변태가 억제되었다. 이러한 CpBV의 발육억제 효과는 이 바이러스의 항체를 함께 주입하여 줌으로 해소되었다. 여기서 항체 단독으로는 파방나방 유충 발육에 아무런 영향을 주지 않았다. 이 결과는 CpBV가 공생 기생봉의 비기주체에 대해서 발육교란 효과를 줄 수 있다는 것을 제시하고 있다.