Fig. 3: P7 interacts with NbMPK3/6 both in vivo and in vitro. | Nature Communications

Fig. 3: P7 interacts with NbMPK3/6 both in vivo and in vitro.

From: A viral protein activates the MAPK pathway to promote viral infection by downregulating callose deposition in plants

Fig. 3

a GST pull-down analysis of the interaction between NbMPK3 and P7. P7 was incubated with NbMPK3/6-GST or GST. Input and pull-down products were analysed by western blot with anti-His and anti-GST antibodies. The molecular weights of GST, NbMPK3-GST, NbMPK6-GST and P7-His are 28, 70, 72 and 27 kDa, respectively. b LCI assay was used to detect interaction between P7 and NbMPK3/6. Luminescence signals were recorded in N. benthamiana leaves at 48 hpi. nLUC-SGT1 and cLUC-RAR1 served as positive controls. nLUC and cLUC served as negative controls. c BiFC assay to detect interaction of P7 with NbMPK3/6. P7-nYFP was coexpressed with NbMPK3-cYFP, NbMPK6-cYFP, AtPIP2A-cYFP or cYFP. NbMPK3-cYFP was coexpressed with AtPIP2A-nYFP or nYFP. NbMPK6-cYFP was coexpressed with AtPIP2A-nYFP or nYFP. YFP signals in N. benthamiana leaves were recorded at 48 hpi (bar = 50 µm). AtPIP2A-cYFP, AtPIP2A-nYFP, nYFP and cYFP served as negative controls. d Subcellular location of P7 interacting with NbMPK3/6 detected by western blot using a polyclonal YFP antibody. T total protein; M membrane protein; C cytoplasmic protein; N nuclear protein. H+-ATP served as a plasma membrane marker. Actin served as a cytoplasmic marker. H3 served as a nuclear marker. The molecular weights of P7-nYFP, NbMPK3-cYFP and NbMPK6-cYFP are 27, 55 and 57 kDa, respectively. For (a, c, d) the experiments were repeated three times with similar results. Source data are provided as a Source Data file.

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