In vitro association between the helper component–proteinase of zucchini yellow mosaic virus and cuticle proteins of Myzus persicae

Abstract

Potyviruses, as typical non-persistently transmitted viruses, are carried within the stylets of aphids. Cuticle proteins (CuPs), which are a major component of the insect cuticle, were examined for in vitro binding to the potyviral helper component-proteinase (HC-Pro). Proteins in 8 M urea extracts from Myzus persicae were separated by SDS-PAGE, electroblotted onto membranes and identified as CuPs by using specific antibodies to M. persicae CuP. Blotted M. persicae protein extracts were overlaid with two HC-Pros, differing by the presence of K or E in the KLSC domain. The HC-Pro with KLSC, known to assist transmission, was found to bind M. persicae proteins, whereas the HC-Pro with ELSC, being deficient in assisting transmission, did not. To identify CuPs that react with HC-Pro, protein extracts were separated by two-dimensional gel electrophoresis. Nine proteins reacting with HC-Pro were sequenced by mass spectrometry. Sequences of peptides in four proteins, of molecular masses between 22 and 31 kDa, were identified as CuPs according to comparison with sequences in GenBank. The putative CuPs from M. persicae that bind HC-Pro are potentially of interest in locating receptors for virions bound to HC-Pro in aphids' stylets.

The helper-component protease transmission factor of tobacco etch potyvirus binds specifically to an aphid ribosomal protein homologous to the laminin receptor precursor

“…The refined identification assigned to each protein spot is indicated. ZYMV, to identify HCPro interactors in the cuticle of an aphid vector (Dombrovsky et al, 2007). A similar approach was adopted in this research using purified transmission-active TEV HCPro protein as bait.…”

“…Following reports describing the retention site for transmission of non-persistent viruses as the distal tip of the maxillary stylets (Wang et al, 1996;Martin et al, 1997), the interaction with HCPro was assayed with extracts obtained from aphid heads, after 2D separation of the proteins. To maximize the chance of detecting new products, a different extraction procedure compared with that used in the study with ZYMV (Dombrovsky et al, 2007) was adopted, as our purpose was to expand the range of potential receptors.…”

“…In any case, identifying elements in the vector that participate in the transmission process is a prerequisite for a better comprehension of the mechanisms governing dissemination of plant viruses. The present work is a start in this direction, aiming to expand the list of aphid proteins interacting with the potyviral HCPro, in addition to other proteins identified previously (Dombrovsky et al, 2007). The fact that, in both studies, HCPro mutants failed to interact with aphid proteins might indicate that more than one protein could be involved in the transmission process.…”

“…Overcoming this hindrance, a study using a far-Western blotting approach in the aphid Myzus persicae (Sulzer) produced a set of cuticular proteins that were able to interact with the HCPro of zucchini yellow mosaic virus (ZYMV) (Dombrovsky et al, 2007). In this study, the absence of interaction with a mutated version of HCPro unable to assist ZYMV transmission by a change in a KLSC motif (similar to the KITC motif of TEV) suggested a correlation between binding to cuticle proteins and transmission.…”

“…Variants of HCPro that are non-functional for transmission due to mutations in a conserved KITC motif have been described (Atreya et al, 1992). In the case of tobacco etch virus (TEV), this motif has been implicated in binding to aphid stylets (Blanc et al, 1998).The number of virus particles involved in transmission has been estimated to be quite low in the case of potyviruses (Moury et al, 2007), which may explain why it has been so difficult to identify putative receptors in aphids.Overcoming this hindrance, a study using a far-Western blotting approach in the aphid Myzus persicae (Sulzer) produced a set of cuticular proteins that were able to interact with the HCPro of zucchini yellow mosaic virus (ZYMV) (Dombrovsky et al, 2007). In this study, the absence of interaction with a mutated version of HCPro unable to assist ZYMV transmission by a change in a KLSC motif (similar to the KITC motif of TEV) suggested a correlation between binding to cuticle proteins and transmission.…”