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Nitric oxide (NO) orchestrates a plethora of incongruent plant immune responses, including the reprograming of global gene expression. However, the cognate molecular mechanisms remain largely unknown. Here we show a zinc finger transcription factor (ZF-TF), SRG1, is a central target of NO bioactivity during plant immunity, where it functions as a positive regulator. NO accumulation promotes SRG1 expression and subsequently SRG1 occupies a repeated canonical sequence within target promoters. An EAR domain enables SRG1 to recruit the corepressor TOPLESS, suppressing target gene expression. Sustained NO synthesis drives SRG1 S -nitrosylation predominantly at Cys87, relieving both SRG1 DNA binding and transcriptional repression activity. Accordingly, mutation of Cys87 compromises NO-mediated control of SRG1-dependent transcriptional suppression. Thus, the SRG1-SNO formation may contribute to a negative feedback loop that attenuates the plant immune response. SRG1 Cys87 is evolutionary conserved and thus may be a target for redox regulation of ZF-TF function across phylogenetic kingdoms. Upon pathogen infection plants accumulate nitric oxide which subsequently regulates defence gene expression. Here, the authors show that S -nitrosylation of the zinc finger transcription factor SRG1 affects transcriptional suppression and contributes to activation of defence responses.

Nitric oxide signalling is a central feature of plant biology. Specificity in this process is achieved through the chemistries of interacting molecules and mechanisms for selective S-nitrosylation/denitrosylation of protein targets. Abstract Reactive nitrogen species (RNS) and their cognate redox signalling networks pervade almost all facets of plant growth, development, immunity, and environmental interactions. The emerging evidence implies that specificity in redox signalling is achieved by a multilayered molecular framework. This encompasses the production of redox cues in the locale of the given protein target and protein tertiary structures that convey the appropriate local chemical environment to support redox-based, post-translational modifications (PTMs). Nascent nitrosylases have also recently emerged that mediate the formation of redox-based PTMs. Reversal of these redox-based PTMs, rather than their formation, is also a major contributor of signalling specificity. In this context, the activities of S-nitrosoglutathione (GSNO) reductase and thioredoxin h5 (Trxh5) are a key feature. Redox signalling specificity is also conveyed by the unique chemistries of individual RNS which is overlaid on the structural constraints imposed by tertiary protein structure in gating access to given redox switches. Finally, the interactions between RNS and ROS (reactive oxygen species) can also indirectly establish signalling specificity through shaping the formation of appropriate redox cues. It is anticipated that some of these insights might function as primers to initiate their future translation into agricultural, horticultural, and industrial biological applications.

Methylated chemicals are widely used as key intermediates for the syntheses of pharmaceuticals, fragrances, flavors, biofuels and plastics. In nature, the process of methylation is commonly undertaken by a super-family of S-adenosyl methionine-dependent enzymes known as methyltransferases. Herein, we describe a novel high throughput enzyme-coupled assay for determining methyltransferase activites. Adenosylhomocysteine nucleosidase, xanthine oxidase, and horseradish peroxidase enzymes were shown to function in tandem to generate a fluorescence signal in the presence of S-adenosyl-L-homocysteine and Amplex Red (10-acetyl-3,7-dihydroxyphenoxazine). Since S-adenosyl-L-homocysteine is a key by-product of reactions catalyzed by S-adenosyl methionine-dependent methyltransferases, the coupling enzymes were used to assess the activities of RI methyltransferase and a salicylic acid methyltransferase from in the presence of S-adenosyl methionine. For the RI methyltransferase, the assay was sensitive enough to allow the monitoring of DNA methylation in the nanomolar range. In the case of the salicylic acid methyltransferase, detectable activity was observed for several substrates including salicylic acid, benzoic acid, 3-hydroxybenzoic acid, and vanillic acid. Additionally, the synthesis of the relatively expensive and unstable cosubstrate, S-adenosyl methionine, catalyzed by methionine adenosyltransferase could be incorporated within the assay. Overall, the assay offers an excellent level of sensitivity that permits continuous and reliable monitoring of methyltransferase activities. We anticipate this assay will serve as a useful bioanalytical tool for the rapid screening of S-adenosyl methionine-dependent methyltransferase activities.

Nitric oxide (NO), more benign than its more reactive and damaging related molecules, reactive oxygen species (ROS), is perfectly suited for duties as a redox signalling molecule. A key route for NO bioactivity is through S-nitrosation, the addition of an NO moiety to a protein Cys thiol (-SH). This redox-based, post-translational modification (PTM) can modify protein function analogous to more well established PTMs such as phosphorylation, for example by modulating enzyme activity, localization, or protein–protein interactions. At the heart of the underpinning chemistry associated with this PTM is sulfur. The emerging evidence suggests that S-nitrosation is integral to a myriad of plant biological processes embedded in both development and environmental relations. However, a role for S-nitrosation is perhaps most well established in plant–pathogen interactions.

• Nitric oxide (NO) regulates the deployment of a phalanx of immune responses, chief among which is the activation of a constellation of defence-related genes. However, the underlying molecular mechanisms remain largely unknown. The Arabidopsis thaliana zinc finger transcription factor (ZF-TF), S-nitrosothiol (SNO) Regulated 1 (SRG1), is a central target of NO bioactivity during plant immunity. Here we characterize the remaining members of the SRG gene family. • Both SRG2 and, especially, SRG3 were positive regulators of salicylic acid-dependent plant immunity. Analysis of SRG single, double and triple mutants implied that SRG family members have additive functions in plant immunity and, surprisingly, are under reciprocal regulation. • SRG2 and SRG3 localized to the nucleus and functioned as ethylene-responsive element binding factor-associated amphiphilic repression (EAR) domain-dependent transcriptional repressors: NO abolished this activity for SRG3 but not for SRG2. Consistently, loss of GSNOR function, resulting in increased (S)NO concentrations, fully suppressed the disease resistance phenotype established from SRG3 but not SRG2 overexpression. Remarkably, SRG3 but not SRG2 was S-nitrosylated in vitro and in vivo. • Our findings suggest that the SRG family has separable functions in plant immunity, and, surprisingly, these ZF-TFs exhibit reciprocal regulation. It is remarkable that, through neofunctionalization, the SRG family has evolved to become differentially regulated by the key immune-related redox cue, NO.

Phytophthora species can infect hundreds of different plants, including many important crops, causing a number of agriculturally relevant diseases. A key feature of attempted pathogen infection is the rapid production of the redox active molecule nitric oxide (NO). However, the potential role(s) of NO in plant resistance against Phytophthora is relatively unexplored. Here we show that the level of NO accumulation is crucial for basal resistance in Arabidopsis against Phytophthora parasitica. Counterintuitively, both relatively low or relatively high NO accumulation leads to reduced resistance against P. parasitica. S‐nitrosylation, the addition of a NO group to a protein cysteine thiol to form an S‐nitrosothiol, is an important route for NO bioactivity and this process is regulated predominantly by S‐nitrosoglutathione reductase 1 (GSNOR1). Loss‐of‐function mutations in GSNOR1 disable both salicylic acid accumulation and associated signalling, and also the production of reactive oxygen species, leading to susceptibility towards P. parasitica. Significantly, we also demonstrate that secreted proteins from P. parasitica can inhibit Arabidopsis GSNOR1 activity. Perturbations in Arabidopsis nitric oxide homeostasis promote disease susceptibility towards the oomycete pathogen Phytophthora parasitica.

As a gaseous biological signaling molecule, nitric oxide (NO) regulates many physiological processes in plants. Over the last decades, this low molecular weight compound has been identified as a key signaling molecule to regulate plant stress responses, and also plays an important role in plant development. However, elucidation of the molecular mechanisms for NO in leaf development has so far been limited due to a lack of mutant resources. Here, we employed the NO-deficient mutant nia1nia2 to examine the role of NO in leaf development. We have found that nia1nia2 mutant plants displayed very different leaf phenotypes as compared to wild type Col-0. Further studies have shown that reactive oxygen species (ROS) levels are higher in nia1nia2 mutant plants. Interestingly, ROS-related enzymes ascorbate peroxidase (APX), catalases (CAT), and peroxidases (POD) have shown decreases in their activities. Our transcriptome data have revealed that the ROS synthesis gene RBOHD was enhanced in nia1nia2 mutants and the photosynthesis-related pathway was impaired, which suggests that NO is required for chloroplast development and leaf development. Together, these results imply that NO plays a significant role in plant leaf development by regulating ROS homeostasis.

Researchers of super-diversity, a state of intense cultural heterogeneity common mostly in urban centers across the globe, have strongly emphasized the potential for rigorous qualitative studies of this social phenomenon to supplement perspectives in the field of linguistic anthropology by focusing on \"what is lived and expressed in the everyday\" (Blommaert & Rampton, 2011, p. 11) and analyzing \"the degree to and ways in which today's migrants maintain identities, activities, and connections linking them with communities outside\" (Vertovec, 2007, p. 1043). Moreover, in studies of language and literacy socialization in contexts of super-diversity, it has become highly significant to explore the patterns of \"inter-generational language socialization within families\" (Blommaert & Rampton, 2011, p. 14), focusing on the direction of influence (parent to child, child to parent, grandparent to child, sibling to sibling, etc.), as well as the context of its occurrence (whether in domestic, recreational, community, or religious circles). This study contributes to the growing literature on super-diversity and more specifically to the field of linguistic anthropology and research on immigrant mothers. Bringing the Pakistani immigrant community in Toronto into focus, the present study demonstrates how Pakistani immigrant mothers, who are themselves in the midst of negotiating linguistic and cultural transitions, negotiate and construct their familial identities, while living in an increasingly diverse city like Toronto in a time when notions of diversity, multiculturalism, and multilingualism are themselves shifting. Analysis of multiple dimensions of immigrant mothers' stories highlights the relational aspect of identity negotiation (Bucholtz & Hall, 2005) in the context of immigration. Attending closely to the narratives of their past and present experiences with language learning and use, this study demonstrates how these mothers have negotiated their familial identities in relation to both their own positioning (Davies & Harré 1990) as immigrant mothers, as well as through the positioning of others' in their stories. Moreover, close analysis of the multiple instances of identity negotiation through the way immigrant mothers talk about language and the socialization of their children confirms their own as well as their children's fluid, multiple and hybrid identities. Their negotiations of `new' immigrant identities into `a third space' draw on an amalgam of cultural and linguistic resources available in a city like Toronto. These findings on one hand problematize and transgress traditionally held notions of language and identity that have been focusing on English versus ethnic language and culture debate; on the other hand, they offer us a window into how immigrant identities are negotiated in superdiverse contexts. Finally, I consider how the narratives constructed in an interactional context like the focus group discussions that constituted an important methodological tool in this study offer broader options for analysis of identity negotiation by providing special insights into how perspectives can be negotiated collaboratively among group members.

Leaf primordia initiation takes place at the flanks of SAM and then passes through common developmental stages. Very different final leaf shapes and sizes result from varying the timing and further patterning events within these developmental stages. Similar final leaf shapes may also result from very distinct early events. Begonia section Gireoudia is a recently radiated group of species with highly divergent leaf forms. I have used a classical genetic approach and candidate gene approach to explain the evolution of leaf form in this genus. These results suggest that convergent evolution of peltate leaves may be through changes at different loci. Key developmental regulators KNOX and ARP genes are reported to be involved in the evolution of leaf form in different species. I have shown that in at least one species ARP is linked to the evolution of peltate leaf form. In a second species there is no link between STM-like KNOX genes and leaf dissection. Estimates of the rate of evolution of ARP CDS showed that different domains of the genes are under different selection pressures. Myb domain2 of ARP genes is under positive selection and variable between two copies of ARP genes in Begonia. Results of complementation tests with Begonia ARP genes in Arabidopsis show that ARPs from Begonia are functionally equivalent to Arabidopsis AS1 genes and one of the two ARP genes in Begonia may be a dominant negative. Expression analysis based on insitu hybridization in compound, peltate and simple leaved Begonias is described. There is no variation in expression patterns between peltate, non peltate or compound leaved Begonia species for BARP1 and KNB1 genes.