Figure 1.

Effect of 2-ME on the Synthesis of Phaseolin.

Leaf protoplasts prepared from transgenic tobacco expressing T343F phaseolin ([A] and [B]) or Δ418 phaseolin (C) were pulse-labeled with [³⁵S]Met and [³⁵S]Cys for 1 h and chased for the indicated times. Immunoprecipitations were with anti-phaseolin antiserum. Analysis was by SDS-PAGE and fluorography. The positions of intact phaseolin (arrowheads), phaseolin vacuolar fragments (vertical bars), deglycosylated phaseolin (asterisk), and molecular mass markers (numbers at right; in kD) are indicated.

(A) Chase was performed in the absence or presence of 2-ME at the indicated concentrations. Phaseolin was immunoprecipitated from protoplast or incubation medium homogenates.

(B) Lanes 1 to 6, chase was performed in the absence (−) or presence (+) of 20 mM 2-ME. Phaseolin was immunoprecipitated from soluble (sol) or microsomal (micr) fractions prepared from protoplast homogenates. Lanes 7 and 8, phaseolin was immunoprecipitated from an aliquot of the subcellular fraction shown in lane 6 and incubated with (+) or without (−) endoglycosidase H (eH).

(C) Chase was performed in the absence (−) or presence (+) of 20 mM 2-ME. Phaseolin was immunoprecipitated from protoplast homogenates.