Cell-free translation of polysomal messenger RNA isolated from healthy and rust-infected wheat leaves

Abstract

Polysomes have been isolated from healthy wheat leaves ( Triticum vulgare L., van. W2691) and from those inoculated with the wheat rust fungus ( Puccinia graminis f. sp. tritici, race 126-ANZ-6, 7). When these are incubated with the soluble components of a cell-free protein synthesizing system from wheat embryos, translation of polysomal messenger RNA resumes. Analytical gel electrophoresis has shown that the newly synthesized polypeptides range in mol. wt from 10 000 to 90 000 daltons. In this reconstituted protein synthesizing system, some size classes of polypeptides synthesized by polysomes from inoculated leaves greatly outnumber those synthesized by the polysomes from healthy leaves. The polysomes from healthy and inoculated leaves consist of messenger RNA-bound ribosomes which contain polypeptide chains that have been initiated in vitro. The time course of protein synthesis and the size of the polypeptides synthesized have suggested that in the cell-free system, most of the in vivo initiated polypeptide chains are fully elongated. Experiments with a polypeptide chain initiation inhibitor (aurintricarboxylic acid) have revealed that under these conditions, there is little or no reinitiation of new chains. Thus, the size classes of newly synthesized polypeptides reflect the size classes of polysomal messenger RNA molecules. These results have revealed remarkable changes in the wheat leaf polysomal messenger RNA populations at 3 days after inoculation with the rust fungus.