An aspartic proteinase gene family in the filamentous fungus Botrytis cinerea contains members with novel features

Abstract

Botrytis cinerea, an important fungal plant pathogen, secretes aspartic proteinase (AP) activity in axenic cultures. No cysteine, serine or metalloproteinase activity could be detected. Proteinase activity was higher in culture medium containing BSA or wheat germ extract, as compared to minimal medium. A proportion of the enzyme activity remained in the extracellular glucan sheath. AP was also the only type of proteinase activity in fluid obtained from B. cinerea-infected tissue of apple, pepper, tomato and zucchini. Five B. cinerea genes encoding an AP were cloned and denoted Bcap1–5. Features of the encoded proteins are discussed. BcAP1, especially, has novel characteristics. A phylogenetic analysis was performed comprising sequences originating from different kingdoms. BcAP1 and BcAP5 did not cluster in a bootstrap-supported clade. BcAP2 clusters with vacuolar APs. BcAP3 and BcAP4 cluster with secreted APs in a clade that also contains glycosylphosphatidylinositol-anchored proteinases from Saccharomyces cerevisiae and Candida albicans. All five Bcap genes are expressed in liquid cultures. Transcript levels of Bcap1, Bcap2, Bcap3 and Bcap4 are subject to glucose and peptone repression. Transcripts from all five Bcap genes were detected in infected plant tissue, indicating that at least part of the AP activity in planta originates from the pathogen.

Identification of soluble secreted proteins from appressoria of Colletotrichum higginsianum by analysis of expressed sequence tags

“…gloeosporioides) and other fungal plant and animal pathogens (Hube et al, 1997;Plummer et al, 2004). Secreted proteases could degrade plant cell wall proteins during penetration or interfere with host-defence-related proteins (ten Have et al, 2004), although in G. cingulata an aspartyl protease expressed in appressoria was dispensable for pathogenicity (Plummer et al, 2004). We also identified a putative pectin methyl esterase that could assist host penetration.…”

pH controls both transcription and post-translational processing of the protease BcACP1 in the phytopathogenic fungus Botrytis cinerea

“…Among these enzymes are pectinases, cellulases and xylanases that are involved in polysaccharide degradation (St Leger et al, 1997;Cotton et al, 2003;Olivieri et al, 2004;Brito et al, 2006). Several classes of proteases have also been described (Billon-Grand et al, 2002;Bindschedler et al, 2003;Ten Have et al, 2004), whose proposed role would be to facilitate host tissue penetration and colonization by degrading structural plant cell wall proteins, and by releasing amino acids, which represent the pathogen's main source of nitrogen and sulfur during infection (Rauscher et al, 1995).…”

Effect of different carbon sources on proteases secreted by the fungal pathogen Sclerotinia sclerotiorum during Phaseolus vulgaris infection

“…In S. sclerotiorum and Botrytis cinerea the proteolytic activity seems to be correlated with the development of symptoms (Movahedi and Heale, 1990) and the enzymes are considered important virulence factors (Cotton et al, 2002;Rollins, 2003). Proteases are also regarded as antagonists of antifungal proteins secreted as part of the defense responses by the host (Poussereau et al, 2001a;ten Have et al, 2004).…”

“…S. sclerotiorum uses an infection strategy based on the acidification of the environment via the production of oxalic acid (Magro et al, 1984) and the secretion of numerous cell-wall degrading enzymes. The action of these enzymes, such as endopolygalacturonases, cellulases, pectin lyase, xylanase, β-galactosidase, and proteases, is believed to facilitate intercellular fungal growth and destabilize host cell integrity (ten Have et al, 2004).…”