Figure 5.

Activation of the SIPK in Tobacco Cells Treated with Harpin_Psph.

Tobacco cells treated with 1 μM harpin_Psph were harvested at the times after infiltration indicated and used to prepare protein extracts.

(A) Kinase activity determined by an in-gel kinase assay using MBP as the substrate.

(B) Protein extracts analyzed by immunoblotting using an antiserum (Ab) recognizing the _pTE_pY motif of activated MAPK.

(C) Protein extracts were prepared from tobacco cells treated with buffer (lane 1) or harpin_Psph (lanes 2 and 3) for 5 min. For immunoprecipitation, the tobacco SIPK-specific antibody (Ab-p48N) (Zhang et al., 1998) was added alone (lanes 1 and 2) or together with the competitor peptide used for antibody production (lane 3). Kinase activity of immunoprecipitated material was assayed using MBP as the substrate. The phosphorylated MBP was visualized by autoradiography.