Partial characterization of specific inducers of a cuticle-degrading protease from the insect pathogenic fungus Metarhizium anisopliae

Abstract

The insect pathogenic fungus Metarhizium anisopliae produces several extracellular cuticle-degrading proteases and evidence is consistent with one of these, PR1, which is a chymoelastase, being a determinant of pathogenicity. We have shown previously that PR1 production is regulated by both carbon catabolite and nitrogen metabolite repression and also by specific induction under derepressed conditions by insect cuticle. In the present work we have established that an enzymically released proteinaceous component(s) of insect cuticle is capable of inducing PR1 (based on appearance of extracellular activity). Cuticle of the desert locust Schistocerca gregaria treated with KOH to remove protein failed to induce PR1 production, whereas cuticle treated with either chloroform or ether to remove lipids still induced PR1. Cuticle digested with either PR1 or the trypsin-like PR2 of M. anisopliae released peptides mainly in the range 150-2000 Da; addition of these peptides generated by PR1 or PR2 at 3 micrograms alanine equivalents ml-1 induced PR1 production to a level similar (75%) to that obtained with untreated insect cuticle. Several amino acids and peptides which are abundant in insect cuticular protein (Ala, Gly, Ala-Ala, Ala-Ala-Ala, Ala-Pro and Pro-Ala) were tested at a range of concentrations and in restricted cultures for their ability to induce PR1. None induced the protease to the levels seen with cuticle or peptides enzymically released from cuticle, although some dimers and notably the monomers Ala and Gly gave 2-2.7-fold enhanced PR1 activity above depressed basal levels (up to 48-57% of that achieved with induced synthesis on cuticle).(ABSTRACT TRUNCATED AT 250 WORDS)