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A putative family 3 glycosyl hydrolase (GH) gene showed significant differential expression in resistant Sinapis alba, compared with the susceptible Brassica juncea, as part of the initial responses during interaction with the necrotroph Alternaria brassicicola. To understand the mechanism of induction, the promoter was isolated and deletion analysis carried out. All the promoter fragments were fused with the β-glucuronidase gene and the expressions were studied in stable B. juncea transgenics and transiently transformed Nicotiana tabacum. Analysis of the expression of the promoter showed the presence of functional abscisic acid (ABA)-, jasmonic acid (JA)-, and salicylic acid (SA)-responsive cis elements. Interestingly, the promoter was found to be induced in both S. alba and B. juncea upon challenge with A. brassicicola but, in S. alba, SA had an inhibitory effect on the pathogen-induced expression of the gene whereas, in B. juncea, SA did not have any negative effect. Therefore, the SA-mediated inhibition in S. alba indicates that the induction is probably through JA or ABA signaling. The difference in the mechanism of induction of the same promoter in the resistant and susceptible plants is probably due to the differential hormonal responses initiated upon challenge with A. brassicicola. [ABSTRACT FROM AUTHOR]

Indian mustard (Brassica juncea) faces significant yield loss due to the 'Black Spot Disease,' caused by a fungus Alternaria brassicicola. In plants, NAC transcription factors (NAC TFs) are known for their roles in development and stress tolerance. One such NAC TF , NAC 62 , was induced during A. brassicicola challenge in Sinapis alba, a non-host resistant plant against this fungus. Sequence analyses of BjuNAC62 from B. juncea showed that it belonged to the membrane-bound class of transcription factors. Gene expression study revealed differential protein processing of NAC62 between B. juncea and S. alba on pathogen challenge. Furthermore, NAC62 processing to 25 kDa protein was found to be unique to the resistant plant during pathogenesis. Conditional expression of BjuNAC62ΔC , which lacks its transmembrane domain, in B. juncea showed improved tolerance to A. brassicicola. BjuNAC62ΔC processing to 25 kDa product was also observed in tolerant transgenic plants. Additionally, transgenic plants showed induced expression of genes associated with defense-related phytohormone signaling pathways on pathogen challenge. Again, altered phenotypes suggest a possible developmental effect of BjuNAC62∆C in transgenic plants. The overall results suggest that the processing of BjuNAC62 might be playing a crucial role in resistance response against Black Spot disease by modulating defense-associated genes. • Arabidopsis orthlogue of ANAC062 is present in Brassica juncea and Sinapis alba. • This NAC TF , NAC62 , possesses a C-terminal transmembrane domain (TMD). • NAC62 is preferentially induced during Black Spot disease in S. alba. • Processing of NAC62 to 25 kDa is unique to S. alba and associated with resistance • Conditional expression of NAC62 ∆ C in B. juncea provides tolerance to this disease. [ABSTRACT FROM AUTHOR]

The enhanced expression of PLD genes and the quantitative increase in the level of phosphatidic acid (PA) in resistant Sinapis alba compared to the downregulated or unchanged expression of the PLD genes and decreased PA level in susceptible Brassica juncea when challenged with Alternaria brassicicola , indicated a positive relationship between PA-mediated signalling and resistance against this necrotrophic pathogen. Furthermore, spraying PA onto the susceptible species B. juncea increased resistance and enhanced expression of ABA-responsive genes, which was comparable to the expression of these genes in S. alba when challenged with A. brassicicola . • The enhanced expression of several PLD genes in resistant S. alba was observed compared to B. juncea upon challenge with A. brassicicola . • An increase in the PA level in S. alba compared to B. juncea following interaction with the pathogen. • Leaves of B. juncea incubated on PA-soaked filter paper did not show resistance. • Spraying B. juncea leaves with PA led to resistance as well as to the enhanced expression of ABA-responsive genes. • A temporal increase in the PA level is associated with increased resistance against A. brassicicola . [ABSTRACT FROM AUTHOR]

Abstract: This work addresses the changes in the phytohormonal signature in the recognition of the necrotrophic fungal pathogen Alternaria brassicicola by susceptible Brassica juncea and resistant Sinapis alba. Although B. juncea, S. alba and Arabidopsis all belong to the same family, Brassicaceae, the phytohormonal response of susceptible B. juncea towards this pathogen is unique because the latter two species express non-host resistance. The differential expression of the PR1 gene and the increased level of salicylic acid (SA) indicated that an SA-mediated biotrophic mode of defence response was triggered in B. juncea upon challenge with the pathogen. Compared to B. juncea, resistant S. alba initiated enhanced abscisic acid (ABA) and jasmonic acid (JA) responses following challenge with this pathogen, as revealed by monitoring the expression of ABA-related genes along with the concentration of ABA and JA. Furthermore, these results were verified by the exogenous application of ABA on B. juncea leaves prior to challenge with A. brassicicola, which resulted in a delayed disease progression, followed by the inhibition of the pathogen-mediated increase in SA response and enhanced JA levels. Therefore, it seems that A. brassicicola is steering the defence response towards a biotrophic mode by mounting an SA response in susceptible B. juncea, whereas the enhanced ABA response of S. alba not only counteracts the SA response but also restores the necrotrophic mode of resistance by enhancing JA biosynthesis. [Copyright &y& Elsevier]

• Lysin motif receptor-like kinase 4 isolated from Brassica juncea , is an inactive kinase, localized to the plasma membrane. • Overexpression (OE) of LYK4 enhanced tolerance against Alternaria brassicicola. • OE also exhibited a trichome dense phenotype with increased methyl jasmonate responsiveness. • Several transcription factors involved in defense and trichome development were significantly upregulated in the OE lines. • LYK4 has a dual functional role in defense and trichome development. Lysin motif receptor-like kinases (LYKs) are involved in the recognition of chitin and activation of plant immune response. In this study, we found LYK4 to be strongly induced in resistant Sinapis alba compared with susceptible Brassica juncea on challenge with Alternaria brassicicola. In silico analysis and in vitro kinase assay revealed that despite the presence of canonical protein kinase fold, B.juncea LYK4 (BjLYK4) lacks several key residues of a prototype protein kinase which renders it catalytically inactive. Transient expression analysis confirmed that fluorescently tagged BjLYK4 localizes specifically to the plasma membrane. Overexpression (OE) of BjLYK4 in B. juncea enhanced tolerance against A. brassicicola. Interestingly, the OE lines also exhibited a novel trichome dense phenotype and increased jasmonic acid (JA) responsiveness. We further showed that many chitin responsive WRKY transcription factors and JA biosynthetic genes were strongly induced in the OE lines on challenge with the pathogen. Moreover, several JA inducible trichome developmental genes constituting the WD-repeat/bHLH/MYB activator complex were also upregulated in the OE lines compared with vector control and RNA interference line. These results suggest that BjLYK4 plays an essential role in chitin-dependent activation of defense response and chitin independent trichome development likely by influencing the JA signaling pathway. [ABSTRACT FROM AUTHOR]

Plants have evolved a specific class of transcription factors (TF) known as NAC TFs (NAM, ATAF and CUC), which have diverse functions including responses to abiotic and biotic stresses. To understand the role of NAC TFs in plant defence response against 'Black Spot' disease caused by Alternaria brassicicola in oilseed mustard (Brassica juncea), the NAC TFs identified from transcriptome data of non-host resistant Sinapis alba challenged with the pathogen were used, since no complete resistant variant of oilseed mustard is available. Subsequent phylogenetic analysis of the 38 NAC TFs, identified from S. alba and the corresponding NAC sequences from B. juncea clustered the NAC TFs into 4 groups. Upon challenge with A. brassicicola , expression dynamics of 13 selected NAC TFs led to the identification of 6 NAC TFs (NAC19, NAC36, NAC55, NAC62, NAC72, and NAC81) which were significantly induced in resistant S. alba , ABA (abscisic acid) treated B. juncea B85 (which gives tolerance) and Alternaria tolerant B. juncea (PAB9511), compared to susceptible B. juncea (B85). Further study revealed that the selected NACs were induced upon desiccation and/or wound treatment as well as by methyl-jasmonate, indicating a mutually shared function of these NAC s in both abiotic and biotic stresses in B. juncea and S. alba. Finally, our data support that these NACs despite being evolutionarily conserved in mediating desiccation and wound response among B. juncea and S. alba , show variation in early signalling upon recognition of the pathogen, which is a major issue for differential expression of these genes. • The study includes A. brassicicola resistant S. alba and susceptible B. juncea. • Transcriptome data available in public domain were used to identify NAC sequences. • A. brassicicola induced NAC TFs were identified and validated. • These NAC TFs were induced during desiccation and wound stresses in B. juncea. • Some unique dual stress responsive NAC TFs were identified from this study. [ABSTRACT FROM AUTHOR]