The NSP-interacting kinase, NIK, belongs to the five leucine-rich repeats-containing receptor-like serine/threonine kinase subfamily that includes members involved in plant development and defence. NIK was first identified by its capacity to interact with the geminivirus nuclear shuttle protein (NSP) and has been strongly associated with plant defence against geminivirus. Recent studies corroborate its function in transducing a defence signal against virus infection and describe components of the NIK-mediated antiviral signalling pathway. This mini-review describes the role of NIK as a transducer of a novel layer of plant innate defence, presents new data on NIK function, and discusses its possible involvement in plant development.

SummaryBegomovirus-associated epidemics currently threaten tomato production worldwide due to the emergence of highly pathogenic virus species and the proliferation of a whitefly B biotype vector that is adapted to tomato. To generate an efficient defence against begomovirus, we modulated the activity of the immune defence receptor nuclear shuttle protein (NSP)-interacting kinase (NIK) in tomato plants; NIK is a virulence target of the begomovirus NSP during infection. Mutation of T474 within the kinase activation loop promoted the constitutive activation of NIK-mediated defences, resulting in the down-regulation of translation-related genes and the suppression of global translation. Consistent with these findings, transgenic lines harbouring an activating mutation (T474D) were tolerant to the tomato-infecting begomoviruses ToYSV and ToSRV. This phenotype was associated with reduced loading of coat protein viral mRNA in actively translating polysomes, lower infection efficiency and reduced accumulation of viral DNA in systemic leaves. Our results also add some relevant insights into the mechanism underlying the NIK-mediated defence. We observed that the mock-inoculated T474D-overexpressing lines showed a constitutively infected wild-type transcriptome, indicating that the activation of the NIKmediated signalling pathway triggers a typical response to begomovirus infection. In addition, the gain-of-function mutant T474D could sustain an activated NIK-mediated antiviral response in the absence of the virus, further confirming that phosphorylation of Thr-474 is the crucial event that leads to the activation of the kinase.

Despite the fundamental importance of nicotinamide adenine dinucleotide (NAD+) for metabolism, the physiological roles of NAD+ carriers in plants remain unclear. We previously characterized the Arabidopsis thaliana gene (At1g25380), named AtNDT2, encoding a protein located in the mitochondrial inner membrane, which imports NAD+ from the cytosol using ADP and AMP as counter-exchange substrates for NAD+. Here, we further investigated the physiological roles of NDT2, by isolating a T-DNA insertion line, generating an antisense line and characterizing these genotypes in detail. Reduced NDT2 expression affected reproductive phase by reducing total seed yield. In addition, reduced seed germination and retardation in seedling establishment were observed in the mutant lines. Moreover, remarkable changes in primary metabolism were observed in dry and germinated seeds and an increase in fatty acid levels was verified during seedling establishment. Furthermore, flowers and seedlings of NDT2 mutants displayed upregulation of de novo and salvage pathway genes encoding NAD+ biosynthesis enzymes, demonstrating the transcriptional control mediated by NDT2 activity over these genes. Taken together, our results suggest that NDT2 expression is fundamental for maintaining NAD+ balance amongst organelles that modulate metabolism, physiology and developmental processes of heterotrophic tissues.

Thioredoxins (Trxs) modulate metabolic responses during stress conditions; however the mechanisms governing the responses of plants subjected to multiple drought events and the role of Trxs under these conditions are not well understood. Here we explored the significance of the mitochondrial Trx system in Arabidopsis following exposure to single and repeated drought events. We analyzed the previously characterized NADPH-dependent Trx reductase A and B double mutant (ntra ntrb) and two independent mitochondrial thioredoxin o1 (trxo1) mutant lines. Following similar reductions in relative water content (∼50%) Trx mutants subjected to two drought cycles displayed a significantly higher maximum quantum efficiency (F v /F m ) and were less sensitive to drought than their wild type counterparts and that all genotypes subjected to a single drought event. Trx mutant plants displayed a faster recovery after two cycles of drought, as observed by the higher accumulation of secondary metabolites and higher stomatal conductance. Our results indicate that plants exposed to multiple drought cycles are able to modulate their subsequent metabolic and physiological response, suggesting the occurrence of an exquisite acclimation in stressed Arabidopsis plants. Moreover, this differential acclimation involves the participation of a set of metabolic changes as well as redox poise alteration following stress recovery.

Optimizing in vitro shoot regeneration conditions in wheat is one of the important steps in successful micropropagation and gene transformation. Various factors such as genotypes, explants, and phytohormones affect in vitro regeneration of wheat, hindering the ability to tailor genotype-independent protocols. Novel computational approaches such as artificial neural networks (ANNs) can facilitate modeling and predicting outcomes of tissue culture experiments and thereby reduce large experimental treatments and combinations. In this study, generalized regression neural network (GRNN) were used to model and forecast in vitro shoot regeneration outcomes of wheat on the basis of 10 factors including genotypes, explants, and different concentrations of 6-benzylaminopurine (BAP), kinetin (Kin), 2,4-dichlorophenoxyacetic acid (2,4-D), indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), 1-naphthaleneacetic acid (NAA), zeatin, and CuSO4. In addition, GRNN was linked to a genetic algorithm (GA) to identify an optimized solution for maximum shoot regeneration. Results indicated that GRNN could accurately predict the shoot regeneration frequency in the validation set with a coefficient determination of 0.78. Sensitivity analysis demonstrated that shoot regeneration frequency was more sensitive to variables in the order of 2,4-D > explant > genotype < zeatin < NAA. Results of this study suggest that GRNN-GA can be used as a tool, besides experimental approaches, to develop and optimize in vitro genotype-independent regeneration protocols.

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SummaryNicotinamide adenine dinucleotide (NAD +) is an essential coenzyme required for all living organisms. In eukaryotic cells, the final step of NAD + biosynthesis is exclusively cytosolic. Hence, NAD + must be imported into organelles to support their metabolic functions. Three NAD + transporters belonging to the mitochondrial carrier family (MCF) have been biochemically characterized in plants. AtNDT1 (At2g47490), focus of the current study, At NDT2 (At1g25380), targeted to the inner mitochondrial membrane, and At PXN (At2g39970), located in the peroxisomal membrane. Although At NDT1 was presumed to reside in the chloroplast membrane, subcellular localization experiments with green fluorescent protein (GFP) fusions revealed that At NDT1 locates exclusively in the mitochondrial membrane in stably transformed Arabidopsis plants. To understand the biological function of At NDT1 in Arabidopsis, three transgenic lines containing an antisense construct of AtNDT1 under the control of the 35S promoter alongside a T‐DNA insertional line were evaluated. Plants with reduced AtNDT1 expression displayed lower pollen viability, silique length, and higher rate of seed abortion. Furthermore, these plants also exhibited an increased leaf number and leaf area concomitant with higher photosynthetic rates and higher levels of sucrose and starch. Therefore, lower expression of AtNDT1 was associated with enhanced vegetative growth but severe impairment of the reproductive stage. These results are discussed in the context of the mitochondrial localization of At NDT1 and its important role in the cellular NAD + homeostasis for both metabolic and developmental processes in plants.