Heat shock protein synthesis and restriction fragment length polymorphism in a virulent and a weakly virulent strain of Leptosphaeria maculans

Abstract

Two strains of Leptosphaeria maculans, differing in virulence on Brassica napus and with distinct cultural characteristics, were tested for their heat shock response. Following exposure to elevated temperatures, both the strains responded by rapidly synthesizing a set of heat shock proteins (HSPs). Protein synthesis profiles, visualized by labelling with [35S]methionine and resolution on one-dimensional Laemmli gels, revealed a classical heat shock specific pattern: three major thermally induced polypeptides corresponding to the 90-, 80-, and 70-kDa HSPs were observed in both strains. At the optimum heat shock temperature of 40 °C, 11 and 5 new polypeptides appeared labelled in the virulent and weakly virulent strains, respectively. At 45 °C only the three prominent high molecular mass polypeptides were synthesized by the virulent strain, while the weakly virulent strain synthesized these and also showed sustained synthesis of several other normal cellular proteins. Protein synthesis was completely abolished on transfer to 48 °C. Qualitative and quantitative differences were detected in the overall polypeptide profiles of the two strains. Southern blots of genomic DNA of the two strains, digested with three restriction endonucleases and probed with a fragment of the Neurospora crassa hsp70 gene, revealed restriction fragment length polymorphisms with all the three enzymes. Key words: Leptosphaeria maculans, virulent–avirulent pathotypes, heat shock proteins, RFLP, hsp70.