Polydnaviridae - a proposed family of insect viruses with segmented, double-stranded, circular DNA genomes.
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Two related viral genes are located on a single superhelical DNA segment of the multipartite Campoletis sonorensis virus genome.
Campoletis sonorensis virus (CsV) (Polydnaviridae) is a large eucaryotic DNA virus with a structurally complex genome consisting of 28 or more superhelical (SH) DNA segments. Little is known of the relationship between different SH DNAs, but some SH DNAs do cross hybridize, indicating a relatedness between certain SH DNAs. In a previous study of viral expression in parasitized Heliothis virescens larvae, several partially homologous viral mRNAs were also identified (G. W. Blissard, S. B. Vinson, and M. D. Summers, 1986, J. Virol. 57, 318-327). To study the organization of the viral genome and the relationship between two partially homologous viral mRNAs, we analyzed cDNA clones and a cloned SH DNA segment of the CsV genome. Of the two CsV mRNAs examined (1.6 and 1.0 kb), both mRNAs were abundant at 48 hr after parasitization and the 1.6-kb mRNA was detected as early as 2 hr after parasitization. Nucleotide sequence analyses of cDNA clones representing the two partially homologous CsV mRNAs (1.6 and 1.0 kb) show that the two CsV mRNAs share five regions of imperfect homology (68 to 88%) which include a large part of each mRNA. These data indicate that the two mRNAs are transcribed from two separate but closely related CsV genes. Comparison of predicted amino acid sequences shows that the two related viral genes encode proteins with divergent amino acid sequences. Northern and Southern hybridization analyses using cloned cDNAs as probes showed that one CsV mRNA (1.6 kb) is homologous to CsV SH DNAs W, R, and M, while the other mRNA (1.0 kb) shows strong homology only to SH DNA W. By cloning and Southern hybridization mapping of the 15.8-kbp SH DNA W, we demonstrate that the genes for the related 1.6- and 1.0-kb mRNAs are located on different regions of this single 15.8-kbp SH DNA. Alignment of nucleotide sequences from a cloned viral genomic DNA and a cDNA demonstrates that the CsV gene encoding the 1.6-kb mRNA is a spliced gene containing at least two introns. Conservation of splice junctions between the two mRNAs suggests that the 1.0-kb mRNA is also spliced. These data represent the detailed analysis of two closely related CsV mRNAs abundantly expressed in parasitized H. virescens larvae, the first observation of related viral genes in a eucaryotic DNA virus, the demonstration of splicing in the Polydnaviridae, and the cloning and mapping of one of the largest SH DNA segments of the CsV genome.
Base Sequence↗
Polydnavirus genomes reflect their dual roles as mutualists and pathogens.
Symbionts often exhibit significant reductions in genome complexity while pathogens often exhibit increased complexity through acquisition and diversification of virulence determinants. A few organisms have evolved complex life cycles in which they interact as symbionts with one host and pathogens with another. How the predicted and opposing influences of symbiosis and pathogenesis affect genome evolution in such instances, however, is unclear. The Polydnaviridae is a family of double-stranded (ds) DNA viruses associated with parasitoid wasps that parasitize other insects. Polydnaviruses (PDVs) only replicate in wasps but infect and cause severe disease in parasitized hosts. This disease is essential for survival of the parasitoid's offspring. Thus, a true mutualism exists between PDVs and wasps as viral transmission depends on parasitoid survival and parasitoid survival depends on viral infection of the wasp's host. To investigate how life cycle and ancestry affect PDVs, we compared the genomes of Campoletis sonorensis ichnovirus (CsIV) and Microplitis demolitor bracovirus (MdBV). CsIV and MdBV have no direct common ancestor, yet their encapsidated genomes share several features including segmentation, diversification of virulence genes into families, and the absence of genes required for replication. In contrast, CsIV and MdBV share few genes expressed in parasitized hosts. We conclude that the similar organizational features of PDV genomes reflect their shared life cycle but that PDVs associated with ichneumonid and braconid wasps have likely evolved different strategies to cause disease in the wasp's host and promote parasitoid survival.
Animals↗
Origin and evolution of polydnaviruses by symbiogenesis of insect DNA viruses in endoparasitic wasps.
During oviposition, many endoparasitic wasps inject virus-like particles into their insect hosts that enable these parasitoids to evade or directly suppress their hosts' immune system, especially encapsulation by hemocytes. These particles are defined as virions that belong to viruses of the two genera that comprise the family Polydnaviridae, bracoviruses (genus Bracovirus) transmitted by braconid wasps, and ichnoviruses (genus Ichnovirus) transmitted by ichneumonid wasps. Structurally, bracovirus virions resemble nudivirus and baculovirus virions (family Baculoviridae), and ichnovirus virions resemble those of ascoviruses (family Ascoviridae). Whereas nudiviruses, baculoviruses and ascoviruses replicate their DNA and produce progeny virions, polydnavirus DNA is integrated into and replicated from the wasp genome, which also directs virion synthesis. The structural similarity of polydnavirus virions to those of viruses that attack the wasps' lepidopteran hosts, along with polydnavirus transmission and replication biology, suggest that these viruses evolved from insect DNA viruses by symbiogenesis, the same process by which mitochondia and chloroplasts evolved from bacteria. Molecular evidence supporting this hypothesis comes from similarities among structural proteins of ascoviruses and the Campoletis sonorensis ichnovirus. Implications of this hypothesis are that polydnaviruses evolved from viruses, but are no longer viruses, and that DNA packaged into polydnavirus virions is not viral genomic DNA per se, but rather wasp genomic DNA consisting primarily of wasp genes and non-coding DNA. Thus, we suggest that a better understanding of polydnaviruses would result by viewing these not as viruses, but rather as a wasp organelle system that evolved to shuttle wasp genes and proteins into hosts to evade and suppress their immune response.
Amino Acid Sequence↗
RNA interference silences Microplitis demolitor bracovirus genes and implicates glc1.8 in disruption of adhesion in infected host cells.
The family Polydnaviridae consists of ds-DNA viruses that are symbiotically associated with certain parasitoid wasps. PDVs are transmitted vertically but also are injected by wasps into hosts where they cause several physiological alterations including immunosuppression. The PDV genes responsible for mediating immunosuppression and other host alterations remain poorly characterized in large measure because viral mutants cannot be produced to study gene function. Here we report the use of RNA interference (RNAi) to specifically silence the glc1.8 and egf1.0 genes from Microplitis demolitor bracovirus (MdBV) in High Five cells derived from the lepidopteran Trichoplusia ni. Dose-response studies indicated that MdBV infects High Five cells and blocks the ability of these cells to adhere to culture plates. This response was very similar to what occurs in two classes of hemocytes, granular cells, and plasmatocytes, after infection by MdBV. Screening of monoclonal antibody (mAb) markers that distinguish different classes of lepidopteran hemocytes indicated that High Five cells cross-react with three mAbs that recognize granular cells from T. ni. Double-stranded RNA (dsRNA) complementary to glc1.8 specifically silenced glc1.8 expression and rescued the adhesive phenotype of High Five cells. Reciprocally, dsRNA complementary to egf1.0 silenced egf1.0 expression but had no effect on adhesion. The simplicity and potency of RNAi could be extremely useful for analysis of other PDV genes.
Animals↗
PTP-H2 and PTP-H3 from Microplitis demolitor Bracovirus localize to focal adhesions and are antiphagocytic in insect immune cells.
Viruses in the family Polydnaviridae are symbiotically associated with parasitoid wasps. Wasps inject polydnaviruses (PDVs) when laying an egg into their insect host, and expression of viral gene products causes several physiological alterations, including immunosuppression, that allow the wasp's progeny to develop. As with other PDVs, most Microplitis demolitor bracovirus (MdBV) genes are related variants that form gene families. The largest MdBV gene family includes 13 members that encode predicted proteins related to protein tyrosine phosphatases (PTPs). Sequence analysis during the present study indicated that five PTP family members (PTP-H2, -H3, -N1, and -N2) have fully conserved catalytic domains, whereas other family members exhibited replacements, deletions, or rearrangements of amino acids considered essential for tyrosine phosphatase activity. Expression studies indicated that most MdBV PTP genes are expressed in virus-infected host insects, with transcript abundance usually being highest in hemocytes. MdBV-infected hemocytes also exhibited higher levels of tyrosine phosphatase activity than noninfected hemocytes. We produced expression constructs for four of the most abundantly expressed PTP family members and conducted functional studies with hemocyte-like Drosophila S2 cells. These experiments suggested that recombinant PTP-H2 and PTP-H3 are functional tyrosine phosphatases whereas PTP-H1 and PTP-J1 are not. PTP-H2 and -H3 localized to focal adhesions in S2 cells, and coexpression with another MdBV gene product, Glc1.8, resulted in complete inhibition of phagocytosis.
Animals↗
Glc1.8 from Microplitis demolitor bracovirus induces a loss of adhesion and phagocytosis in insect high five and S2 cells.
Polydnaviridae is a unique family of DNA viruses that are symbiotically associated with parasitoid wasps. Upon oviposition, wasps inject these viruses into their hosts, where they cause several physiological alterations, including suppression of the cellular immune response. Here we report that expression of the glc1.8 gene from Microplitis demolitor bracovirus (MdBV) causes a loss of adhesion by two hemocyte-like cell lines, namely, High Five cells from the lepidopteran Trichoplusia ni and S2 cells from the dipteran Drosophila melanogaster. The expression of recombinant Glc1.8 also greatly reduced the ability of these cells to phagocytize foreign targets. Glc1.8 is characterized by a signal peptide at its N terminus, an extracellular domain comprised of five nearly perfect tandem repeats of 78 amino acids, and a C-terminal hydrophobic domain that encodes a putative membrane anchor sequence. The expression of a Glc1.8 mutant lacking the anchor sequence resulted in a secreted protein that had no effect on adhesion or phagocytosis. In contrast, sequential deletion of the repeats in the extracellular domain resulted in a progressive reduction in immunosuppressive activity. Since each repeat and its associated glycosylation sites are nearly identical, these results suggested that adhesion-blocking activity depends more on the overall number of repeats in the extracellular domain than on the specific determinants within each repeat. While it severely compromised adhesion and phagocytic functions, Glc1.8 did not cause cell death. Collectively, these results indicate that Glc1.8 is a major pathogenic determinant of MdBV that is involved in suppression of the insect cellular immune response.
Animals↗
Segment W of Campoletis sonorensis virus: expression, gene products, and organization.
Campoletis sonorensis virus (CsV, Polydnaviridae) is a segmented double-stranded DNA virus which has an apparently symbiotic relationship with the parasitic wasp, Campoletis sonorensis. CsV replicates in the oviducts of the parasitic wasp and is injected into the wasp's host, Heliothis virescens (Lepidoptera; Noctuiidae), during oviposition. In the parasitized lepidopteran host, the virus has a dramatic effect on host physiology and viral gene products are believed to play an essential role in the survival of the parasitic wasp's egg and larva. In the current study, we used Northern blot analyses to examine expression from segment W in the parasitized host and in the parasitic wasp. Segment W hybridized primarily to two relatively abundant mRNAs (1.6 and 1.0 kb) from the parasitized host. These 1.6- and 1.0-kb mRNAs, which were previously shown to be transcribed from two closely related genes (WHv1 and WHv2) on segment W (G. W. Blissard, O. P. Smith, and M. D. Summers, 1987, Virology 160, 120-134) increased in relative abundance between 2 and 24 hr postparasitization (pp) and were detected throughout parasitization (8 days). To study the proteins encoded by these closely related genes, the open reading frame from each of the related genes was cloned into a baculovirus expression vector. By pulse labeling in the presence and absence of tunicamycin, we examined secretion and glycosylation of these CsV proteins in infected lepidopteran cells (Spodoptera frugiperda). Expression of segment W in the oviducts of the female wasp was also examined. Segment W hybridized to at least five CsV mRNAs on Northern blots of poly(A) mRNA from C. sonorensis oviducts. To identify specific CsV mRNAs and map putative viral genes expressed in wasp oviduct tissues, segment W was used to screen a cDNA library of C. sonorensis oviduct mRNAs. Three cDNAs were used to identify CsV mRNAs by Northern blot analyses and to map the locations of three putative CsV genes on segment W. Cross-hybridization within the CsV genome was examined with cloned segment W and with the three cloned cDNAs.
Amino Acid Sequence↗
Characteristics of pathogenic and mutualistic relationships of ascoviruses in field populations of parasitoid wasps.
Ascoviruses are disseminated among larvae in lepidopteran populations by parasitic wasps during oviposition. Ascovirus relationships with these wasps vary from pathogenic to mutualistic, and experimentally can be shown possibly to be commensal non-pathogenic virus having little or no effect. Most ascoviruses are pathogens that female wasps vector mechanically. Other ascoviruses have a more intimate relationship with their wasp vectors in that their genome is stably maintained in all wasp nuclei through several generations by vertical transmission. In this relationship, these viruses are mutualistic, enhancing the successful development of the wasp larvae by suppressing lepidopteran defence mechanisms. The DpAV4 ascovirus is a mutualist in certain Diadromus wasps but is pathogenic or not when vectored by other species of this genus. These various biologies suggest that ascovirus/wasp relationships depend on wasp regulatory factors that control virus replication. Thus, certain ascoviruses can potentially have either a pathogenic, mutualistic, or non-pathogenic relationship with a specific wasp vector, the type of relationship being dependent upon the species system in which the relationship evolved. Finally, because ascoviruses appear to be related to ichnoviruses (Polydnaviridae), the DpAV4/Diadromus system constitutes a possible interesting intermediate between the pathogenic ascoviruses and symbiotic viruses that evolved to be ichnoviruses.
Animals↗
Commensal and mutualistic relationships of reoviruses with their parasitoid wasp hosts.
During evolution, certain endoparasitoid wasps have developed mechanisms to suppress the defence systems of their hosts. For this purpose, these species, all of which belong to the families Ichneumonidae and Braconidae, inject various kinds of virus-like particles. The most studied of these particles are classified as polydnaviruses (family Polydnaviridae) which are symbiotic viruses. Over the past decade, it has also been shown that several wasp species harbour reoviruses (family Reoviridae), and that two of these suppress host defence, allowing the development of the parasitoid eggs. In this paper, we summarize the key features of these viruses and their relationships with their wasp hosts. Five reoviruses are known that appear to be non-pathogenic for the wasps. Three of these, McRVLP, HeRV, OpRVLP, use their wasp hosts as vectors, and do not appear to be involved in host defence suppression. The fourth, DpRV-1, is a commensal reovirus detected in most field populations of the wasp, Diadromus pulchellus. This reovirus is always found associated with an ascovirus, DpAV-4a, which is indispensable for host immune suppression. Although DpRV-1 has not been shown to directly increase D. pulchellus parasitic success, it may contribute to this success by retarding DpAV-4a replication in the wasp. The fifth reovirus, DpRV-2, occurs in a specific population of D. pulchellus in which DpRV-1 and DpAV-4 are absent. This virus has a mutualistic relationship with its wasp host, as its injection by females during oviposition is essential for host immunosuppression. Interestingly, these viruses belong to several different reovirus genera.
Amino Acid Sequence↗
Campoletis sonorensis Endoparasitic Wasps Contain Forms of C. sonorensis Virus DNA Suggestive of Integrated and Extrachromosomal Polydnavirus DNAs.
Campoletis sonorensis virus (CsV) (Polydnaviridae) previously was detected only in the calyx epithelial cells and lumen of the oviducts from female C. sonorensis (Ichneumonidae) endoparasitic wasps (Norton et al., Cell Tissue Res. 162:195-208, 1975). Using dot-blot hybridizations, we detected low amounts of CsV DNA in male and female wasp head and thorax tissues and in male abdominal tissues. Low amounts of extrachromosomal viral DNA were detected in Southern blots of undigested male wasp DNA and in male DNA purified by isopycnic centrifugation. High-molecular-weight male wasp DNA digested with any of several restriction endonucleases and hybridized with cloned viral DNAs from CsV superhelices B and Q under stringent conditions contained CsV-specific DNA fragments that differed significantly in size and number from the hybridizing fragments detected in comparably digested viral DNA. Identical offsize restriction fragments were detected in digested female head and thorax DNA. These data suggest that at least CsV DNAs B and Q are integrated in C. sonorensis cellular DNA and that the virus may be transmitted through the germline.
Journal Article↗
Identification and comparison of Campoletis sonorensis virus transcripts expressed from four genomic segments in the insect hosts Campoletis sonorensis and Heliothis virescens.
The Campoletis sonorensis virus (CsV; Polydnaviridae) genome consists of at least 28 closed circular superhelical (SH) DNAs. In this study we used complete clones of four SH DNAs to analyze viral transcription both in the adult parasitic wasp host Campoletis sonorensis (Ichneumonidae) and in the lepidopteran host, Heliothis virescens (Noctuidae). CsV genes are expressed in parasitized H. virescens, but no viral transcripts had been characterized from C. sonorensis until this study. The clones of the SH DNAs B, H, M, and O1 were used to probe Northern blots of poly(A)+ RNA isolated from C. sonorensis reproductive tissue and from parasitized H. virescens larvae. All four SH DNAs hybridized to viral transcripts. SH-H,-M, and -O1 hybridized to messages expressed in both hosts. SH-B and -M hybridized to transcripts that were detected only in either C. sonorensis reproductive tissue or parasitized H. virescens larvae. These results suggest that some CsV genes are expressed in a host-specific manner. In a previous study we identified a family of imperfectly conserved tandemly repeated 540-bp repeat elements on SH-B,-H and -O1 (D. A. Theilmann and M. D. Summers, 1987 J. Virol. 61; 2589-2598). Hybridization of the 540-bp repeat regions to Northern blots showed that they were all homologous to viral transcripts. A cDNA clone of a mRNA that is transcribed from the 540-bp repeat region of SH-B was isolated from a lambda gt 10 library and completely sequenced. The sequence data revealed that the 540-bp repeat element was contained within the open reading frame of this gene. These results indicate that transcribed sequences homologous to the 540-bp repeat elements represent a second gene family to be identified within the CsV genome.
Amino Acid Sequence↗
Expression and hemocyte-targeting of a Campoletis sonorensis polydnavirus cysteine-rich gene in Heliothis virescens larvae.
The polydnavirus associated with the parasitic wasp Campoletis sonorensis is injected into the lepidopteran insect, Heliothis virescens, during parasitization, after which viral gene products suppress the cellular immune system of the hosts. Four related cysteine-rich polydnavirus gene have been identified in parasitized H. virescens larvae and grouped into a family. In this study, we investigated the expression and hemocyte targeting of the cysteine-rich VHv1.4 protein. Full-length and truncated VHv1.4 proteins were produced in a bacterial expression system, and the purified proteins were used to raise polyclonal antisera. In immunoblots the VHv1.4 protein was detected in parasitized insects as early as 6 h and throughout the entire course of parasitism. The VHv1.4 protein appeared predominantly in the plasma fraction of hemolymph from parasitized larvae, suggesting that this protein is secreted. The VHv1.4 protein expressed from a recombinant baculovirus was secreted in two lepidopteran cell lines and in larvae injected with the recombinant virus. Digestion with endoglycosidases suggests that the VHv1.4 protein is glycosylated at multiple N-glycosylation sites. Immunofluorescence assays showed that the VHv1.4 protein binds to the hemocytes, most notably the granulocytes, in H. virescens larvae. After binding, the VHv1.4 protein was internalized, probably by endocytosis. Specific binding of the VHv1.4 to granulocytes implies an important function in the suppression of host cellular encapsulation response.
Animals↗
Cloning and expression of a gene encoding a Campoletis sonorensis polydnavirus structural protein.
Polydnaviruses are the only known group of mutualistic viruses. They are required for successful parasitization in many braconid and ichneumonid parasitoids. The intimacy of this mutualistic association is indicated by the integration and vertical transmission of polydnaviruses in wasp genomes and by their asymptomatic, developmentally regulated replication. The evolution of this mutualism raises several interesting issues that require a better understanding of the viral genome and viral replication. To develop probes for virus replication and morphogenesis, we have begun to characterize several viral structural proteins. A 699 bp cDNA encoding the p12 viral structural protein was cloned and sequenced. The p12 gene localizes to viral segment Y and encodes a predicted protein of 92 amino acids that does not encode a signal peptide and is unrelated to known peptide or nucleic acid sequences. The p12 mRNA is detected at the onset of virus replication. mRNA titers increase with increasing rates of virus replication. Polyclonal antisera raised against histidine-tagged p12 protein expressed in bacteria reacted specifically with the p12 polypeptide in Western blots of CsPDV virions. The p12 polypeptide was not detected in non-replicative wasp or lepidopteran tissues by Western blot analyses but was readily detected in protein extracts of wasp ovaries. The data indicate that the p12 gene is a viral gene encoding a virion protein and provides a specific probe for virus replication that will be useful for studying the evolution of this group of mutualistic viruses.
Amino Acid Sequence↗
Plasmatocyte spreading peptide does not induce Microplitis demolitor polydnavirus-infected plasmatocytes to spread on foreign surfaces.
Capsule formation by the moth Pseudopulsia includens requires that plasmatocytes change from being nonadhesive cells in circulation to strongly adhesive cells capable of attaching to the foreign target and one another. This change in adhesive state is induced by Plasmatocyte Spreading Peptide (PSP1); a 23 amino acid peptide isolated from P. includens plasma. Plasmatocytes from hosts parasitized by Microplitis demolitor remain in a nonadhesive state after infection by Microplitis demolitor polydnavirus (MdPDV). This alteration in plasmatocyte function prevents P. includens from encapsulating the developing parasitoid. In the current study, we examined whether MdPDV infection eliminates PSP1-responsive plasmatocytes from circulation or disrupts the ability of PSP1 to induce adhesion and spreading of plasmatocytes to foreign surfaces. In vivo experiments revealed that infection of P. includens by MdPDV induced an increase in the total number of hemocytes in circulation but reduced the proportion of hemocytes in circulation that were plasmatocytes. However, plasmatocytes normally capable of responding to PSP1 were not eliminated from circulation. Both in vivo and in vitro experiments indicated that plasmatocytes inoculated with MdPDV lost the capacity to respond to PSP1 4-6 h post-infection. Infection of P. includens with MdPDV reduced expression levels of prepro-PSP1 mRNA in hemocytes but did not appear to alter expression levels in fat body.
Animals↗
Polydnavirus of Campoletis chlorideae: characterization and temporal effect on host Helicoverpa armigera cellular immune response.
Polydnavirus was isolated from oviduct calyx in the parasitoid wasp Campoletis chlorideae (Hymenoptera: Ichneumonidae), and termed CcIV. The virus particles consist of fusiform nucleocapsids surrounded by two unit membrane envelopes. The DNAs purified from these viruses were found diversified in molecular weight and existed in nonequimolar concentrations. At least 20 different-sized bands were present after electrophoresis, and they ranged from approximately 3 to 26 kb. Persistence and gene expression of CcIV were examined in parasitized and virus-injected Helicoverpa armigera larvae. Viral DNA could be detected in the hemocyte of H. armigera at 30 min post-parasitization (p.p.), and persisted for 6 days. While no viral DNA increase was found, CcIV transcripts were first detected in host hemocytes at day 1 p.p. and continued for 5 days. Similar transcripts were observed in hemocytes from larvae that had been injected with calyx fluid or CcIV 24 h earlier. CcIV viral DNAs hybridized only with certain first-strand cDNAs from hemocytes, suggesting that only part of the CcIV genome was expressed in H. armigera. The pattern of CcIV gene expression was consistent with that of the inhibition of encapsulation for Sephadex G-10 and parasitoid eggs by host larvae. The recovery of host immune response at day 4 p.p. indicated that CcIV exhibited a partial and temporal effect on the host immune system and the developing parasitoid appeared to avoid encapsulation via different mechanisms.
Animals↗
Identification of host translation inhibitory factor of Campoletis sonorensis ichnovirus on the tobacco budworm, Heliothis virescens.
Parasitization of a wasp, Campoletis sonorensis, against the larvae of Heliothis virescens depresses synthesis of specific host proteins related to growth and immunity. It has been suggested that the inhibition of host gene expression is targeted at a posttranscriptional level. This study aimed to verify the identity of host translation inhibitory factor (HTIF) derived from wasp parasitization. To identify HTIF, the proteins in the parasitized host were fractionated using different protein purification methods, and each fraction's HTIF activity was assessed. In the course of the protein purification steps, HTIF activity was highly correlated with the fractions containing VHv 1.4 protein, which has a conserved cysteine-motif and is encoded in C. sonorensis ichnovirus (CsIV). Purified VHv 1.4 protein using an immunoaffinity column exhibited a significant HTIF effect, while the heat-inactivated VHv 1.4 did not. Both recombinant VHv 1.4 and VHv 1.1 (another cys-motif protein encoded in CsIV) proteins were synthesized in Sf 9 cells through a baculovirus expression system. The purified recombinant VHv 1.4 and VHv 1.1 exhibited significant HTIF activities in a nanomolar range. However, VHv1.4 protein showed about four times higher HTIF activity than did VHv 1.1 protein. Both HTIFs acted directly on translation machinery because they inhibited a cell-free in vitro translation system using rabbit reticulocyte lysate. Both HTIFs are likely to discriminate specific target mRNAs because they inhibited translation of RNA extracts from the Tn 368 cell line, but not from Sf 9 cells. In addition, they inhibited translation of RNAs from fat body, hemocytes, and testis, but not from epidermis, gut, labial gland, and nerve tissues of H. virescens. These results indicate that both cys-motif proteins of VHv 1.4 and VHv 1.1 play a role as HTIF in C. sonorensis parasitization.
Animals↗
Microplitis demolitor bracovirus inhibits phagocytosis by hemocytes from Pseudoplusia includens.
The braconid wasp Microplitis demolitor carries Microplitis demolitor bracovirus (MdBV) and parasitizes the larval stage of several noctuid moths. A key function of MdBV in parasitism is suppression of the host's cellular immune response. Prior studies in the host Pseudoplusia includens indicated that MdBV blocks encapsulation by preventing two types of hemocytes, plasmatocytes and granulocytes, from adhering to foreign targets. The other main immune response mediated by insect hemocytes is phagocytosis. The goal of this study was to determine which hemocyte types were phagocytic in P. includens and to assess whether MdBV infection affects this defense response. Using the bacterium Escherichia coli and inert polystyrene beads as targets, our results indicated that the professional phagocyte in P. includens is granulocytes. The phagocytic responses of granulocytes were very similar to those of High Five cells that prior studies have suggested are a granulocyte-like cell line. MdBV infection dose-dependently disrupted phagocytosis in both cell types by inhibiting adhesion of targets to the cell surface. The MdBV glc1.8 gene encodes a cell surface glycoprotein that had previously been implicated in disruption of adhesion and encapsulation responses by immune cells. Knockdown of glc1.8 expression by RNA interference (RNAi) during the current study rescued the ability of MdBV-infected High Five cells to phagocytize targets. Collectively, these results indicate that glc1.8 is a key virulence determinant in disruption of both adhesion and phagocytosis by insect immune cells.
Animals↗