Molecular and Functional Analyses of a Maize Autoactive NB-LRR Protein Identify Precise Structural Requirements for Activity
Fig 3
Rp1-D21 triggers a hypersensitive response phenotype when transiently expressed in N. benthamiana.
(A) Rp1-D21, Rp1-D and Rp1-dp2 proteins fused with a C-terminal EGFP tag were agro-infiltrated into N. benthamiana, with an empty vector (EV) as a negative control. A representative leaf was photographed at 3 days post infiltration (dpi, left). Total protein was extracted from agro-infiltrated leaves at 30 hours post infiltration (hpi), and anti-GFP antibody was used to detect the expression of the fused proteins (right). The sizes of the proteins were labeled on the right. Equal loading of protein samples was shown by Ponceau-S staining of the Rubisco subunit (below, right). (B) Rp1-D21, Rp1-D and Rp1-dp2 proteins fused with a C-terminal 3×HA tag were agro-infiltrated into N. benthamiana, with empty vector (EV) as a negative control. A representative leaf was photographed at 3 dpi (left). Total protein was extracted from agro-infiltrated leaves at 30 hpi, and anti-HA antibody was used to detect the expression of the fused proteins. Equal loading of protein samples was shown by Ponceau-S staining of Rubisco subunit (below, right). These experiments were repeated three times with the same results.