Improving photosynthesis and crop productivity by accelerating recovery from photoprotection

Crop leaves in full sunlight dissipate damaging excess absorbed light energy as heat. When sunlit leaves are shaded by clouds or other leaves, this protective dissipation continues for many minutes and reduces photosynthesis. Calculations have shown that this could cost field crops up to 20% of their potential yield. Here, we describe the bioengineering of an accelerated response to natural shading events in Nicotiana (tobacco), resulting in increased leaf carbon dioxide uptake and plant dry matter productivity by about 15% in fluctuating light. Because the photoprotective mechanism that has been altered is common to all flowering plants and crops, the findings provide proof of concept for a route to obtaining a sustainable increase in productivity for food crops and a much-needed yield jump

Transient genome-wide interactions of the master transcription factor NLP7 initiate a rapid nitrogen-response cascade

Dynamic reprogramming of gene regulatory networks (GRNs) enables organisms to rapidly respond to environmental perturbation. However, the underlying transient interactions between transcription factors (TFs) and genome-wide targets typically elude biochemical detection. Here, we capture both stable and transient TF-target interactions genome-wide within minutes after controlled TF nuclear import using time-series chromatin immunoprecipitation (ChIP-seq) and/or DNA adenine methyltransferase identification (DamID-seq). The transient TF-target interactions captured uncover the early mode-of-action of NIN-LIKE PROTEIN 7 (NLP7), a master regulator of the nitrogen signaling pathway in plants. These transient NLP7 targets captured in root cells using temporal TF perturbation account for 50% of NLP7-regulated genes not detectably bound by NLP7 in planta. Rapid and transient NLP7 binding activates early nitrogen response TFs, which we validate to amplify the NLP7-initiated transcriptional cascade. Our approaches to capture transient TF-target interactions genome-wide can be applied to validate dynamic GRN models for any pathway or organism of interest. Conventional methods cannot reveal transient transcription factors (TFs) and targets interactions. Here, Alvarez et al. capture both stable and transient TF-target interactions by time-series ChIP-seq and/or DamID-seq in a cell-based TF perturbation system and show NLP7 as a master TF to initiate a rapid nitrogen-response cascade

\u3ci\u3ePhotosystem II Subunit S\u3c/i\u3e overexpression increases the efficiency of water use in a field-grown crop

Insufficient water availability for crop production is a mounting barrier to achieving the 70% increase in food production that will be needed by 2050. One solution is to develop crops that require less water per unit mass of production. Water vapor transpires from leaves through stomata, which also facilitate the influx of CO2 during photosynthetic assimilation. Here, we hypothesize that Photosystem II Subunit S (PsbS) expression affects a chloroplastderived signal for stomatal opening in response to light, which can be used to improve wateruse efficiency. Transgenic tobacco plants with a range of PsbS expression, from undetectable to 3.7 times wild-type are generated. Plants with increased PsbS expression show less stomatal opening in response to light, resulting in a 25% reduction in water loss per CO2 assimilated under field conditions. Since the role of PsbS is universal across higher plants, this manipulation should be effective across all crops

Global Analysis of Arabidopsis/Downy Mildew Interactions Reveals Prevalence of Incomplete Resistance and Rapid Evolution of Pathogen Recognition

Interactions between Arabidopsis thaliana and its native obligate oomycete pathogen Hyaloperonospora arabidopsidis (Hpa) represent a model system to study evolution of natural variation in a host/pathogen interaction. Both Arabidopsis and Hpa genomes are sequenced and collections of different sub-species are available. We analyzed ∼400 interactions between different Arabidopsis accessions and five strains of Hpa. We examined the pathogen's overall ability to reproduce on a given host, and performed detailed cytological staining to assay for pathogen growth and hypersensitive cell death response in the host. We demonstrate that intermediate levels of resistance are prevalent among Arabidopsis populations and correlate strongly with host developmental stage. In addition to looking at plant responses to challenge by whole pathogen inoculations, we investigated the Arabidopsis resistance attributed to recognition of the individual Hpa effectors, ATR1 and ATR13. Our results suggest that recognition of these effectors is evolutionarily dynamic and does not form a single clade in overall Arabidopsis phylogeny for either effector. Furthermore, we show that the ultimate outcome of the interactions can be modified by the pathogen, despite a defined gene-for-gene resistance in the host. These data indicate that the outcome of disease and disease resistance depends on genome-for-genome interactions between the host and its pathogen, rather than single gene pairs as thought previously

Structural Elucidation and Functional Characterization of the Hyaloperonospora arabidopsidis Effector Protein ATR13

The oomycete Hyaloperonospora arabidopsidis (Hpa) is the causal agent of downy mildew on the model plant Arabidopsis thaliana and has been adapted as a model system to investigate pathogen virulence strategies and plant disease resistance mechanisms. Recognition of Hpa infection occurs when plant resistance proteins (R-genes) detect the presence or activity of pathogen-derived protein effectors delivered to the plant host. This study examines the Hpa effector ATR13 Emco5 and its recognition by RPP13-Nd, the cognate R-gene that triggers programmed cell death (HR) in the presence of recognized ATR13 variants. Herein, we use NMR to solve the backbone structure of ATR13 Emco5, revealing both a helical domain and a disordered internal loop. Additionally, we use site-directed and random mutagenesis to identify several amino acid residues involved in the recognition response conferred by RPP13-Nd. Using our structure as a scaffold, we map these residues to one of two surface-exposed patches of residues under diversifying selection. Exploring possible roles of the disordered region within the ATR13 structure, we perform domain swapping experiments and identify a peptide sequence involved in nucleolar localization. We conclude that ATR13 is a highly dynamic protein with no clear structural homologues that contains two surface-exposed patches of polymorphism, only one of which is involved in RPP13-Nd recognition specificity

Genetic, Biochemical, and Structural Studies of the Hyaloperonospora arabidopsidis Effector ATR13 and its Cognate Arabidopsis thaliana R-gene, RPP13

The oomycete Hyaloperonospora arabidopsidis (Hpa) is the causal agent of downy mildew on the model plant Arabidopsis thaliana. Since oomycetes are a particularly important agricultural pest, this pathosystem is an ideal setting for exploration of interactions between host and microbe, as genome sequences for both are available and possess a high level of genetic diversity between naturally occurring populations. Plant recognition of Hpa infection occurs when resistance proteins (R-genes) in the plant host recognize pathogen-derived effectors, which are proteins delivered to the host. One such protein, the Hpa effector ATR13 Emco5, is examined in this study. Herein, we use NMR to solve the backbone structure of a highly disordered protein, ATR13 Emco5, revealing a loosely packed protein possessing a great deal of flexibility. In addition to solving this structure, we use site-directed and random mutagenesis to expose several amino acid residues involved in the recognition response conferred by RPP13-Nd, the cognate R-gene that triggers programmed cell death (HR) in the presence of recognized ATR13 variants. Using our structure as a scaffold, we map these residues to one of two surface-exposed patches composed of residues that are under diversifying selection. Exploring possible roles of the disordered region within the ATR13 structure, we perform domain swapping experiments and identify a peptide sequence involved in nucleolar localization. We conclude that ATR13 is a highly dynamic protein with weak structural similarities to RAN-GTP that contains two surface-exposed patches, only one of which is involved in RPP13 recognition specificity. Furthermore, we have identified two potential protein targets of ATR13 (the RCC1 and PRP39 families). Finally, and we have performed several different EMS mutagenesis screens leading to the isolation of three lines of A. thaliana plants disrupted in RPP13 signaling

Genetic, Biochemical, and Structural Studies of the Hyaloperonospora arabidopsidis Effector ATR13 and its Cognate Arabidopsis thaliana R-gene, RPP13

The oomycete Hyaloperonospora arabidopsidis (Hpa) is the causal agent of downy mildew on the model plant Arabidopsis thaliana. Since oomycetes are a particularly important agricultural pest, this pathosystem is an ideal setting for exploration of interactions between host and microbe, as genome sequences for both are available and possess a high level of genetic diversity between naturally occurring populations. Plant recognition of Hpa infection occurs when resistance proteins (R-genes) in the plant host recognize pathogen-derived effectors, which are proteins delivered to the host. One such protein, the Hpa effector ATR13 Emco5, is examined in this study. Herein, we use NMR to solve the backbone structure of a highly disordered protein, ATR13 Emco5, revealing a loosely packed protein possessing a great deal of flexibility. In addition to solving this structure, we use site-directed and random mutagenesis to expose several amino acid residues involved in the recognition response conferred by RPP13-Nd, the cognate R-gene that triggers programmed cell death (HR) in the presence of recognized ATR13 variants. Using our structure as a scaffold, we map these residues to one of two surface-exposed patches composed of residues that are under diversifying selection. Exploring possible roles of the disordered region within the ATR13 structure, we perform domain swapping experiments and identify a peptide sequence involved in nucleolar localization. We conclude that ATR13 is a highly dynamic protein with weak structural similarities to RAN-GTP that contains two surface-exposed patches, only one of which is involved in RPP13 recognition specificity. Furthermore, we have identified two potential protein targets of ATR13 (the RCC1 and PRP39 families). Finally, and we have performed several different EMS mutagenesis screens leading to the isolation of three lines of A. thaliana plants disrupted in RPP13 signaling