Separation and identification of candidate protein elicitors from the cultivation medium of Leptosphaeria maculans inducing resistance in Brassica napus

Abstract

The Dothideomycete Leptosphaeria maculans, a worldwide fungal pathogen of oilseed rape (Brassica napus), secretes a broad spectrum of molecules into the cultivation medium during growth in vitro. Here, candidate elicitor molecules, which induce resistance in B. napus to L. maculans, were identified in the cultivation medium. The elicitation activity was indicated by increased transcription of pathogenesis-related gene 1 (PR1) and enhanced resistance of B. napus plants to the invasion of L. maculans. The elicitation activity was significantly lowered when the cultivation medium was heated to 80°C. Active components were further characterized by specific cleavage with the proteolytic enzymes trypsin and proteinase K and with glycosidases α-amylase and β-glucanase. The elicitor activity was eliminated by proteolytic digestion while glycosidases had no effect. The filtered medium was fractionated by either ion-exchange chromatography or isoelectric focusing. Mass spectrometry analysis of the most active fractions obtained by both separation procedures revealed predominantly enzymes that can be involved in the degradation of plant cell wall polysaccharides. This is the first study searching for L. maculans-specific secreted elicitors with a potential to be used as defense-activating agents in the protection of B. napus against L. maculans in agriculture. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:918-928, 2016.

Arabinogalactan Protein-Like Proteins From Ulva lactuca Activate Immune Responses and Plant Resistance in an Oilseed Crop

“…Besides our results correspond to the results of Cluzet et al (2004) , who described the increased expression of CHI , PR1 , and PR10 genes after treatments with various Ulva extracts. The activation of PR1 transcription was also reported in B. napus cotyledons after treatment with oligosaccharide elicitor isolated from L. maculans mycelium ( Kim et al, 2013 ) and protein elicitor isolated from L. maculans cultivation medium ( Nováková et al, 2016 ). The high production of hydrogen peroxide together with a strong induction of SA and ET signaling pathways explains significant inhibitory effects of AGPs on the infection development.…”

Current Status and Challenges in Identifying Disease Resistance Genes in Brassica napus

“…With better genome assemblies and higher gene prediction accuracies, we will soon have better insights into the structures and functions of effector proteins, which allows more efficient screening and identification of R genes (Gibriel et al, 2016 ). Elicitor candidates for L. maculans (Wilson and Howlett, 2005 ; Nováková et al, 2016a ), and effector candidates for S. sclerotiorum (Guyon et al, 2014 ), are both responsible for inducing HR. RNAi studies established that the transcription factor LmStuA in L. maculans regulates expression of effector genes (Soyer et al, 2015 ).…”