The conserved aromatic residue W¹²² is a determinant of potyviral coat protein stability, replication, and cell‐to‐cell movement in plants

Abstract

Coat proteins (CPs) play critical roles in potyvirus cell-to-cell movement. However, the underlying mechanism controlling them remains unclear. Here, we show that substitutions of alanine, glutamic acid, or lysine for the conserved residue tryptophan at position 122 (W 122) in tobacco vein banding mosaic virus (TVBMV) CP abolished virus cell-to-cell movement in Nicotiana benthamiana plants. In agroinfiltrated N. benthamiana leaf patches, both the CP and RNA accumulation levels of three W 122 mutant viruses were significantly reduced compared with those of wild-type TVBMV, and CP accumulated to a low level similar to that of a replication-deficient mutant. The results of polyprotein transient expression experiments indicated that CP instability was responsible for the significantly low CP accumulation levels of the three W 122 mutant viruses. The substitution of W 122 did not affect CP plasmodesmata localization or virus particle formation; however, the substitution significantly reduced the number of virus particles. The wild-type TVBMV CP could complement the reduced replication and abolished cell-to-cell movement of the mutant viruses. When the codon for W 122 was mutated to that for a different aromatic residue, phenylalanine or tyrosine, the resultant mutant viruses moved systemically and accumulated up to 80% of the wild-type TVBMV level. Similar results were obtained for the corresponding amino acids of W 122 in the watermelon mosaic virus and potato virus Y CPs. Therefore, we conclude that the aromatic ring in W 122 in the core domain of the potyviral CP is critical for cell-to-cell movement through the effects on CP stability and viral replication.

Identification of genetic determinants of tomato brown rugose fruit virus that enable infection of plants harbouring the Tm‐22 resistance gene

“…The chimeric viruses/MPs were separately transformed into Agrobacterium tumefaciens GV3101 cells. Agrobacterium cultures harbouring different plasmids were prepared as previously described (Yan et al, 2021a) and diluted to OD 600 = 0.2 for virus inoculation and protein transient expression. The agroinfiltrated plants were maintained in a growth room at 23 °C under a 16 hr light/8 hr dark cycle, and photographed under white light using a digital camera (Canon 800D).…”

“…Western blotting was performed as previously described (Yan et al, 2021a). Total protein was extracted from N. benthamiana leaf tissue…”

Pepper Mottle Virus and Its Host Interactions: Current State of Knowledge

“…In contrast, C-terminal regions of TuMV, PVY, and TEV CPs were shown to be associated with viral cell-to-cell and long-distance movement [90,92,94]. The aromatic residue tryptophan at core domain (W 122 ) of CP in tobacco vein banding mosaic virus, which is highly conserved residue among potyviruses, plays a role in maintaining stability of CP during viral replication and this involvement in viral cell-to-cell movement was also observed with the same residue of watermelon mosaic virus and PVY [96]. An additional role as the pathogenicity determinant for CP has been reported in PVY [97].…”

Engineering of the complementary mutation site in tobacco mosaic virus p126 to develop a stable attenuated mutant for cross-protection

“…The plants inoculated with the empty vector pCB301-Rz were used as negative controls. To determine viral cell-tocell movement, the Agrobacterium cultures were diluted to an OD 600 of 0.0001 and individually infiltrated into N. benthamiana leaves (Yan et al 2020;Yan et al 2021). GFP fluorescence from TMV-GFP and its mutants was observed using a Zeiss LSM510 META laser scanning microscope at 60 hpai.…”