Arogenate dehydratases: unique roles in light-directed development during the seed-to-seedling transition in Arabidopsis thaliana.

The seed-to-seedling transition is impacted by changes in nutrient availability and light profiles, but is still poorly understood. Phenylalanine affects early seedling development; thus, the roles of arogenate dehydratases (ADTs), which catalyze phenylalanine formation, were studied in germination and during the seed-to-seedling transition by exploring the impact of light conditions and specific hormone responses in adt mutants of Arabidopsis thaliana. ADT gene expression was assessed in distinct tissues and for light-quality dependence in seedlings for each of the six-member ADT gene family. Mutant adt seedlings were evaluated relative to wild type for germination, photomorphogenesis (blue, red, far red, white light, and dark conditions), anthocyanin accumulation, and plastid development-related phenotypes. ADT proteins are expressed in a light- and tissue-specific manner in transgenic seedlings. Among the analyzed adt mutants, adt3, adt5, and adt6 exhibit significant defects in germination, hypocotyl elongation, and root development responses during the seed-to-seedling transition. Interestingly, adt5 exhibits a light-dependent disruption in plastid development, similar to a phyA mutant. These data indicate interactions between photoreceptors, hormones, and regulation of phenylalanine pools in the process of seedling establishment. ADT5 and ADT6 may play important roles in coordinating hormone and light signals for normal early seedling development.

Plasticity of Arabidopsis rosette transcriptomes and photosynthetic responses in dynamic light conditions.

With the high variability of natural growth environments, plants exhibit flexibility and resilience in regard to the strategies they employ to maintain overall fitness, including maximizing light use for photosynthesis, while simultaneously limiting light-associated damage. We measured distinct parameters of photosynthetic performance of Arabidopsis thaliana plants under dynamic light regimes. Plants were grown to maturity then subjected to the following 5-day (16 h light, 8 h dark) regime: Day 1 at constant light (CL) intensity during light period, representative of a common lab growth condition; Day 2 under sinusoidal variation in light intensity (SL) during the light period that is representative of changes occurring during a clear sunny day; Day 3 under fluctuating light (FL) intensity during the light period that simulates sudden changes that might occur with the movements of clouds in and out of the view of the sun; Day 4, repeat of CL; and Day 5, repeat of FL. We also examined the global transcriptome profile in these growth conditions based on obtaining RNA-sequencing (RNA-seq) data for whole plant rosettes. Our transcriptomic analyses indicated downregulation of photosystem I (PSI) and II (PSII) associated genes, which were correlated with elevated levels of photoinhibition as indicated by measurements of nonphotochemical quenching (NPQ), energy-dependent quenching (qE), and inhibitory quenching (qI) under both SL and FL conditions. Furthermore, our transcriptomic results indicated downregulation of tetrapyrrole biosynthesis associated genes, coupled with reduced levels of chlorophyll under both SL and FL compared with CL, as well as downregulation of photorespiration-associated genes under SL. We also noticed an enrichment of the stress response gene ontology (GO) terms for genes differentially regulated under FL when compared with SL. Collectively, our phenotypic and transcriptome analyses serve as useful resources for probing the underlying molecular mechanisms associated with plant acclimation to rapid light intensity changes in the natural environment.

Nuclear-encoded sigma factor 6 (SIG6) is involved in the block of greening response in Arabidopsis thaliana.

PREMISE: Light is critical in the ability of plants to accumulate chlorophyll. When exposed to far-red (FR) light and then grown in white light in the absence of sucrose, wild-type seedlings fail to green in a response known as the FR block of greening (BOG). This response is controlled by phytochrome A through repression of protochlorophyllide reductase-encoding (POR) genes by FR light coupled with irreversible plastid damage. Sigma (SIG) factors are nuclear-encoded proteins that contribute to plant greening and plastid development through regulating gene transcription in chloroplasts and impacting retrograde signaling from the plastid to nucleus. SIGs are regulated by phytochromes, and the expression of some SIG factors is reduced in phytochrome mutant lines, including phyA. Given the association of phyA with the FR BOG and its regulation of SIG factors, we investigated the potential regulatory role of SIG factors in the FR BOG response. METHODS: We examined FR BOG responses in sig mutants, phytochrome-deficient lines, and mutant lines for several phy-associated factors. We quantified chlorophyll levels and examined expression of key BOG-associated genes. RESULTS: Among six sig mutants, only the sig6 mutant significantly accumulated chlorophyll after FR BOG treatment, similar to the phyA mutant. SIG6 appears to control protochlorophyllide accumulation by contributing to the regulation of tetrapyrrole biosynthesis associated with glutamyl-tRNA reductase (HEMA1) function, select phytochrome-interacting factor genes (PIF4 and PIF6), and PENTA1, which regulates PORA mRNA translation after FR exposure. CONCLUSIONS: Regulation of SIG6 plays a significant role in plant responses to FR exposure during the BOG response.

Isolation, characterization, cloning and bioinformatics analysis of a novel receptor from black cut worm (Agrotis ipsilon) of Bacillus thuringiensis vip 3Aa toxins.

Black cutworm (BCW) is an economically important lepidopteran insect. The control of this insect by a Bt toxin and the understanding of the interaction between the Bt toxin and its receptor molecule were the objectives of this research work. A gene coding for a Vip3A receptor molecule was identified, characterized, and cloned, from the brush border membrane vesicles (BBMV) of the BCW. The nucleotide sequence analysis of the cloned putative Vip3A-receptor gene revealed that the gene was 1.3-kb long and exhibited no homology with any gene in the gene bank. We succeeded in identifying and characterizing most of the Vip3A-receptor gene sequence; and the nucleotide sequence analysis of the cloned putative Vip3A-receptor gene (accession no. KX858809) revealed about 92% of the expected sequence was recovered, which exhibited no homology with any gene in the GenBank.

Developing transgenic wheat to encounter rusts and powdery mildew by overexpressing barley chi26 gene for fungal resistance.

BACKGROUND: The main aim of this study was to improve fungal resistance in bread wheat via transgenesis. Transgenic wheat plants harboring barley chitinase (chi26) gene, driven by maize ubi promoter, were obtained using biolistic bombardment, whereas the herbicide resistance gene, bar, driven by the CaMV 35S promoter was used as a selectable marker. RESULTS: Molecular analysis confirmed the integration, copy number, and the level of expression of the chi26 gene in four independent transgenic events. Chitinase enzyme activity was detected using a standard enzymatic assay. The expression levels of chi26 gene in the different transgenic lines, compared to their respective controls, were determined using qRT-PCR. The transgene was silenced in some transgenic families across generations. Gene silencing in the present study seemed to be random and irreversible. The homozygous transgenic plants of T4, T5, T6, T8, and T9 generations were tested in the field for five growing seasons to evaluate their resistance against rusts and powdery mildew. The results indicated high chitinase activity at T0 and high transgene expression levels in few transgenic families. This resulted in high resistance against wheat rusts and powdery mildew under field conditions. It was indicated by proximate and chemical analyses that one of the transgenic families and the non-transgenic line were substantially equivalent. CONCLUSION: Transgenic wheat with barley chi26 was found to be resistant even after five generations under artificial fungal infection conditions. One transgenic line was proved to be substantially equivalent as compared to the non-transgenic control.

The khmer software package: enabling efficient nucleotide sequence analysis.

The khmer package is a freely available software library for working efficiently with fixed length DNA words, or k-mers. khmer provides implementations of a probabilistic k-mer counting data structure, a compressible De Bruijn graph representation, De Bruijn graph partitioning, and digital normalization. khmer is implemented in C++ and Python, and is freely available under the BSD license at  https://github.com/dib-lab/khmer/.

Development of transgenic wheat (Triticum aestivum L.) expressing avidin gene conferring resistance to stored product insects.

BACKGROUND: Wheat is considered the most important cereal crop all over the world. The wheat weevil Sitophilus granarius is a serious insect pests in much of the wheat growing area worldwide and is responsible for significant loss of yield. Avidin proteins has been proposed to function as plant defense agents against insect pests. RESULTS: A synthetic avidin gene was introduced into spring wheat (Triticum aestivum L.) cv. Giza 168 using a biolistic bombardment protocol. The presence and expression of the transgene in six selected T0 transgenic wheat lines were confirmed at the molecular level. Accumulation of avidin protein was detected in transgenic plants compared to non-transgenic plants. Avidin transgene was stably integrated, transcribed and translated as indicated by Southern blot, ELISA, and dot blot analyses, with a high level of expression in transgenic wheat seeds. However, no expression was detected in untransformed wheat seeds. Functional integrity of avidin was confirmed by insect bioassay. The results of bioassay using transgenic wheat plants challenged with wheat weevil revealed 100 % mortality of the insects reared on transgenic plants after 21 days. CONCLUSION: Transgenic wheat plants had improved resistance to Sitophilus granarius.