Autocatalytic processing of the potyvirus helper component proteinase in Escherichia coli and in vitro

Carrington, James C.; Freed, Deon D.; Sanders, Tatia C.

doi:10.1128/jvi.63.10.4459-4463.1989

Cited by 88 publications

(34 citation statements)

References 31 publications

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“…An immunoreactive product of -62 kd, which co-migrated during SDS -PAGE with the protein identified from F-ID and F-2C plants, was detected in extracts from E-6A and E-4D ( Figure 6, lanes 7 and 8). We have noted previously (Carrington et al, 1989b;Oh and Carrington, 1989) that the sequence Gly-Asn-Ser-Gly, found at positions 623 -626 in the TEV polyprotein near the center of HC-Pro, resembles the motif surrounding the active-site serine residue of serine-type proteinases (Neurath, 1984). This led to speculation that it may be part of a distinct proteolytic domain that functions to process the N terminus of HC-Pro.…”

Section: Expression Of Mutagenized Tev Sequences In Transgenic Plantsmentioning

confidence: 96%

Expression of potyviral polyproteins in transgenic plants reveals three proteolytic activities required for complete processing.

1990

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All proteins encoded by the plant potyvirus, tobacco etch virus (TEV), arise by proteolytic processing of a single polyprotein. Two virus‐encoded proteinases (NIa and HC‐Pro) that catalyze most of the proteolytic events have been characterized previously. The two proteins that are derived from the N‐terminal 87 kd region of the viral polyprotein are a 35 kd protein and HC‐Pro (52 kd). It is demonstrated in this study that a third proteolytic activity is required to process the junction between these proteins. Proteolysis at the HC‐Pro N terminus to separate these proteins occurred poorly, if at all, after in vitro synthesis of a 97 kd polyprotein, whereas cleavage of the HC‐Pro C terminus occurred efficiently by an autoprocessing mechanism. Synthesis of the same polyprotein in transgenic tobacco plants, however, resulted in complete and accurate proteolysis at both termini of HC‐Pro. A point mutation affecting an amino acid residue essential for the proteolytic activity of HC‐Pro had no effect on N‐terminal processing. Expression in transgenic plants of a construct with a large deletion in the 35 kd protein coding region resulted in partial inhibition of HC‐Pro N‐terminal cleavage, suggesting that the 35 kd protein may affect the proteolytic event but not in a catalytic role. We speculate that this cleavage event is catalyzed by either a cryptic potyviral proteinase that requires a host factor or subcellular environment for activation, or possibly a host proteinase.

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Section: Expression Of Mutagenized Tev Sequences In Transgenic Plantsmentioning

confidence: 96%

Expression of potyviral polyproteins in transgenic plants reveals three proteolytic activities required for complete processing.

1990

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“…Potyviruses exist as flexuous, rod-shaped virions comprising a positive-sense, single-stranded RNA (1ssRNA) genome coated by a virally encoded coat protein (Jagadish et al, 1991). On entry into plant cells, the 1ssRNA genome is uncoated and translated into a single polypeptide, which subsequently generates the range of potyviral proteins by autocatalysis (Carrington et al, 1989). Translation of the potyviral RNA is largely dependent on host translation factors.…”

Section: Introductionmentioning

confidence: 99%

Engineering of CRISPR/Cas9‐mediated potyvirus resistance in transgene‐free Arabidopsis plants

Pyott

Sheehan

Molnár

2016

Molecular Plant Pathology

345

184

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SummaryMembers of the eukaryotic translation initiation factor (eIF) gene family, including eIF4E and its paralogue eIF(iso)4E, have previously been identified as recessive resistance alleles against various potyviruses in a range of different hosts. However, the identification and introgression of these alleles into important crop species is often limited. In this study, we utilise CRISPR/Cas9 technology to introduce sequence‐specific deleterious point mutations at the eIF(iso)4E locus in Arabidopsis thaliana to successfully engineer complete resistance to Turnip mosaic virus (TuMV), a major pathogen in field‐grown vegetable crops. By segregating the induced mutation from the CRISPR/Cas9 transgene, we outline a framework for the production of heritable, homozygous mutations in the transgene‐free T2 generation in self‐pollinating species. Analysis of dry weights and flowering times for four independent T3 lines revealed no differences from wild‐type plants under standard growth conditions, suggesting that homozygous mutations in eIF(iso)4E do not affect plant vigour. Thus, the established CRISPR/Cas9 technology provides a new approach for the generation of Potyvirus resistance alleles in important crops without the use of persistent transgenes.

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“…55,119,122 The C-terminal proteolytic cleavage occurs in cis, hence the HC is commonly called HC-Pro. 123 Biological activity of HC-Pro, in terms of aphid transmission, is believed to be mediated by oligomers composed of at least two HC-Pro molecules. 122 The current model for the bridge hypothesis for potyvirus transmission suggests that one surface of the dimer HC-Pro is bound to the aphid stylet and another surface is bound to the potyvirus virion.…”

Section: The Helper Strategymentioning

confidence: 99%

Ipomovirus – an atypical genus in the family Potyviridae transmitted by whiteflies

Dombrovsky

Reingold

Antignus

2014

Pest Management Science

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Ipomoviruses (genus Ipomovirus) are whitefly-transmitted viruses assigned to the family Potyviridae. They are characterised by filamentous flexible particles and a positive-sense single-stranded RNA (+ssRNA) genome. The viral genome is translated into a polyprotein precursor, which is processed into mature proteins and a short overlapping open reading frame. The genus Ipomovirus contains four accepted species and one unapproved species, and two other tentative members have recently been characterised. Ipomoviruses cause serious economic losses in many important crops, including cassava, sweet potato, cucurbits, tomato and aubergine. These viruses are transmitted by whiteflies in a non-circulative, semi-persistent manner, the virions being retained on the external surface of the vectors' mouthparts for a few days or weeks. Comparison of the available complete genome sequences of different ipomoviruses revealed differences in their genome organisation and a considerable variation in their proteins and conserved motifs that may reflect functional differences. This review summarises the current knowledge of the members within the genus Ipomovirus, focusing on genome organisation, taxonomic classification and the mechanism by which they are transmitted.

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Autocatalytic processing of the potyvirus helper component proteinase in Escherichia coli and in vitro

Cited by 88 publications

References 31 publications

Expression of potyviral polyproteins in transgenic plants reveals three proteolytic activities required for complete processing.

Expression of potyviral polyproteins in transgenic plants reveals three proteolytic activities required for complete processing.

Engineering of CRISPR/Cas9‐mediated potyvirus resistance in transgene‐free Arabidopsis plants

Ipomovirus – an atypical genus in the family Potyviridae transmitted by whiteflies

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