Papain-like protease p29 as a symptom determinant encoded by a hypovirulence-associated virus of the chestnut blight fungus

Craven, M G; Pawlyk, Diane M.; Choi, Gil H.; Nuss, Donald L.

doi:10.1128/jvi.67.11.6513-6521.1993

Cited by 92 publications

(38 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Investigations on hypovirus variation in C. parasitica isolates on the grafted trees may assist determinations on which CHV1 regions are or are not associated with certain fungal colony phenotypes. Studies have indicated that p29 in ORF A of CHV1-EP713 is associated with the suppression of pigment production and other symptoms in C. parasitica (Craven et al 1993). Suzuki et al (1999) mapped the p29 symptom-determinant domain to a region extending from Phe(25) through Gln(73).…”

Section: Discussionmentioning

confidence: 99%

Nucleotide sequence identification of Cryphonectria hypovirus 1 infecting Cryphonectria parasitica on grafted American chestnut trees 12–18 years after inoculation with a hypovirulent strain mixture

Griffin¹,

Robbins²,

Hogan³

et al. 2004

Forest Pathology

View full text Add to dashboard Cite

Summary Natural blight cankers on grafted American chestnut Castanea dentata trees were inoculated with one French‐derived hypovirulent Cryphonectria parasitica strain, with a predominantly orange‐pigmented colony phenotype and three Italian hypovirulent C. parasitica strains, with a predominantly white colony phenotype, in 1982 and 1983. Cryphonectria hypovirus 1 (CHV1) dsRNA was extracted from the four inoculated strains and from 11 predominantly white or orange pigmented C. parasitica isolates recovered from blight‐controlled cankers on the grafted trees during 1994–2000; cDNAs were made by reverse transcriptase‐polymerase chain reaction for two hypovirus regions: (i) an 844‐bp region in the helicase domain of open reading frame B (ORF B), designated OB and (ii) an 894‐bp region that included part of the 5′ non‐coding region and part (p29) of ORF A (5′OA). Nucleotide sequence analysis indicated that hypovirus isolates from the grafted trees and Italian inoculated strains had high homologies (identities) to each other and high homologies for OB (98.7–99.9%) and 5′OA (99.2–99.6%) to reference hypovirus, CHV1‐Euro7; homologies to reference hypovirus, CHV1‐EP713, were low (≤89.8%). In contrast, the hypovirus from the predominantly pigmented, French‐derived inoculated strain had high homologies for OB (99.1%) and 5′OA (99.7%) to reference CHV1‐EP713, and low homologies (87.7–89.7%) to the predominantly pigmented grafted‐tree isolates. These results provide strong evidence that hypoviruses present in the two predominantly pigmented isolates and nine predominantly white isolates of C. parasitica from the grafted trees were of Italian inoculated hypovirulent strain origin. For the Phe(25) through Gln(73) phenotype‐determinant region in p29 of 5′OA, hypovirus isolate amino acid sequences were dependent on whether they were CHV1‐Euro7 or CHV1‐EP713 hypoviruses, but predominantly white and predominantly pigmented phenotypes within CHV1‐Euro7 or within CHV1‐EP713 had identical amino acid sequences. Grafted‐tree hypovirus isolates infecting the predominantly pigmented C. parasitica isolates may originate from chance transmission and replication of a CHV1‐Euro7 hypovirus variant or variant mixture, possibly present in one or more of the Italian inoculated hypovirulent strains, such as Ep 49.

show abstract

Section: Discussionmentioning

confidence: 99%

Nucleotide sequence identification of Cryphonectria hypovirus 1 infecting Cryphonectria parasitica on grafted American chestnut trees 12–18 years after inoculation with a hypovirulent strain mixture

Griffin¹,

Robbins²,

Hogan³

et al. 2004

Forest Pathology

View full text Add to dashboard Cite

show abstract

“…Like many viral leader proteases, p29 and p48 also serve important functional roles. Although the p29 protease is not essential for viral replication (11), it serves as a suppressor of the C. parasitica antiviral RNA silencing response to increase viral RNA accumulation (12)(13)(14)(15). Unlike p29, p48 is not dispensable for viral replication.…”

mentioning

confidence: 99%

Mutagenesis of the Catalytic and Cleavage Site Residues of the Hypovirus Papain-Like Proteases p29 and p48 Reveals Alternative Processing and Contributions to Optimal Viral RNA Accumulation

Jensen

Nuss

2014

J Virol

View full text Add to dashboard Cite

The positive-stranded RNA genome of the prototypic virulence-attenuating hypovirus CHV-1/EP713 contains two open reading frames (ORF), each encoding an autocatalytic papain-like leader protease. Protease p29, derived from the N-terminal portion of ORF A, functions as a suppressor of RNA silencing, while protease p48, derived from the N-terminal portion of ORF B, is required for viral RNA replication. The catalytic and cleavage site residues required for autoproteolytic processing have been functionally mapped in vitro for both proteases but not confirmed in the infected fungal host. We report here the mutagenesis of the CHV-1/EP713 infectious cDNA clone to define the requirements for p29 and p48 cleavage and the role of autoproteolysis in the context of hypovirus replication. Mutation of the catalytic cysteine and histidine residues for either p29 or p48 was tolerated but reduced viral RNA accumulation to ca. 20 to 50% of the wild-type level. Mutation of the p29 catalytic residues caused an accumulation of unprocessed ORF A product p69. Surprisingly, the release of p48 from the ORF B-encoded polyprotein was not prevented by mutation of the p48 catalytic and cleavage site residues and was independent of p29. The results show that, while dispensable for hypovirus replication, the autocatalytic processing of the leader proteases p29 and p48 contributes to optimal virus RNA accumulation. The role of the predicted catalytic residues in autoproteolytic processing of p29 was confirmed in the infected host, while p48 was found to also undergo alternative processing independent of the encoded papain-like protease activities. IMPORTANCEHypoviruses are positive-strand RNA mycoviruses that attenuate virulence of their pathogenic fungal hosts. The prototypic hypovirus CHV-1/EP713, which infects the chestnut bight fungus Cryphonetria parasitica, encodes two papain-like autocatalytic leader proteases, p29 and p48, that also have important functions in suppressing the RNA silencing antiviral defense response and in viral RNA replication, respectively. The mutational analyses of the CHV-1/EP713 infectious cDNA clone, reported here, define the requirements for p29 and p48 cleavage and the functional importance of autoproteolysis in the context of hypovirus replication and exposed an alternative p48 processing pathway independent of the encoded papain-like protease activities. These findings provide additional insights into hypovirus gene expression, replication, and evolution and inform ongoing efforts to engineer hypoviruses for interrogating and modulating fungal virulence.

show abstract

“…EP155p29 carries the coding region of p29 protein from CHV1-EP713 hypovirus under gpd1 promoter (G. H. Craven et al 1993). EP155p29stop also carries the p29 sequence under gpd1 promoter, but has three stop codons inserted at the 5'-end of the p29 coding region.…”

Section: Methodsmentioning

confidence: 99%

“…Translated ORF A produces a p69 protein that undergoes autocatalysis into p29 and p40 proteins. Major hypovirulence symptoms, such as loss of pigmentation, lack of conidiation and female sterility, are determined by the p29 protein (Craven et al 1993;Suzuki et al 1999;Suzuki et al 2003). The p29 protein shows significant homology to HC-Pro protease from plant potyviruses (Fukuzawa et al 2010;Suzuki et al 1999).…”

Section: Cryphonectria Parasitica As a Model Organismmentioning

confidence: 99%

“…The p29 protein shows significant homology to HC-Pro protease from plant potyviruses (Fukuzawa et al 2010;Suzuki et al 1999). Early studies also showed that deletion of p29 from the CHV1 genome does not impede its ability to replicate, but C. parasitica strains infected with CHV1Δp29 regain normal levels of pigmentation, asexual sporulation, and laccase activity (Craven et al 1993). There is no complete picture about molecular function of p29, but recent findings indicate that p29, similarly to HC-Pro, serves as suppressor of RNA silencing (RNAi) (Fukuzawa et al 2010;Segers et al 2006;Segers et al 2007).…”

Section: Cryphonectria Parasitica As a Model Organismmentioning

confidence: 99%

See 1 more Smart Citation

Transcriptome analysis of programmed cell death associated with allorecognition in chestnut blight fungus Cryphonectria parasitica

Belov¹

View full text Add to dashboard Cite

i The "dialectics" of this situation is that the evolutionary narratives certainly are oversimplified "myths" that have the unfortunate (and, in modern studies, unintended) teleological flavor (as in "selected for" or, worse, "selected for the purpose of"), yet the language of these narratives seems best suited to describing evolution and formulating falsifiable hypotheses that propel further research. At present, we hardly can give up these stories […] precisely because they are necessary means for the advancement of research, even though they tend to leave a scientist […] with feelings of uneasiness and dissatisfaction. It seems important not to forget that evolutionary narratives effectively are semantic devices that are constructed to structure and simplify our thinking about evolution and to facilitate the generation of hypotheses. These narratives should be prudently distrusted and by no account should be construed as "accurate representations of reality" (whatever that might mean […]). AbstractIn this thesis, I analysed transcriptional profiles of the chestnut blight fungus, Cryphonectria parasitica, during vegetative incompatibility (allorecognition) reactions due to differences at the vic3 locus. Out of a total 13944 identified expressed transcripts, including 2334 novel ones, only 1411 were differentially expressed during vic3 incompatibility reaction. Functional enrichment analysis showed increased expression of genes involved in detoxification and stress response (e.g.s Cytochrome p450, Glutathione S-transferase), and toxin biosynthesis. Surprisingly, even though the test strains were both the same mating type (MAT-2), genes involved in sexual reproduction (mf2-1, mf2-2and mat-2) showed the most dramatic increase in expression during allorecognition response. Further qPCR analysis showed that activation of mating pheromone genes occurs during incompatible reactions involving five of the six known vic incompatibility loci. The only exception was vic4, which elicits a weak incompatibility and showed almost no change in pheromone gene expression. Analysis suggests that mf2-1, mf2-2 and mat-2 expression is triggered by activation of asexual sporulation. Genes encoded at the vic3 locus, vic3a and vic3b, were upregulated in barraging samples along with seven HET-domain genes located at other regions of the genome. Among the seven HET genes activated, one is located at the vic1 locus and previously implicated in vic1incompatibility. Activation of these same HET domain genes also occurred in other vic incompatible pairings. For example, upregulation of dev3a and vic1a genes occurred during incompatibility reactions associated with each of the six known vic loci. This suggests that some HET genes serve as universal allorecognition factors. Furthermore, this data indicates that each incompatibility locus uses a set of several HET genes to iii activate Programmed Cell Death (PCD). In addition, I analysed the effects on barrage formation of p29, a protein-coding region from Cryphonectria hypovirus 1 (CHV1).Ex...

show abstract

Papain-like protease p29 as a symptom determinant encoded by a hypovirulence-associated virus of the chestnut blight fungus

Cited by 92 publications

References 40 publications

Nucleotide sequence identification of Cryphonectria hypovirus 1 infecting Cryphonectria parasitica on grafted American chestnut trees 12–18 years after inoculation with a hypovirulent strain mixture

Nucleotide sequence identification of Cryphonectria hypovirus 1 infecting Cryphonectria parasitica on grafted American chestnut trees 12–18 years after inoculation with a hypovirulent strain mixture

Mutagenesis of the Catalytic and Cleavage Site Residues of the Hypovirus Papain-Like Proteases p29 and p48 Reveals Alternative Processing and Contributions to Optimal Viral RNA Accumulation

Transcriptome analysis of programmed cell death associated with allorecognition in chestnut blight fungus Cryphonectria parasitica

Contact Info

Product

Resources

About