Identification and characterization of inulinases by bioinformatics analysis of bacterial glycoside hydrolases family 32 (GH32).

The glycoside hydrolase family contains enzymes that break the glycosidic bonds of carbohydrates by hydrolysis. Inulinase is one of the most important industrial enzymes in the family of Glycoside Hydrolases 32 (GH32). In this study, to identify and classify bacterial inulinases initially, 16,002 protein sequences belonging to the GH32 family were obtained using various databases. The inulin-effective enzymes (endoinulinase and exoinulinase) were identified. Eight endoinulinases (EC 3.2.1.7) and 4318 exoinulinases (EC 3.2.1.80) were found. Then, the localization of endoinulinase and exoinulinase enzymes in the cell was predicted. Among them, two extracellular endoinulinases and 1232 extracellular exoinulinases were found. The biochemical properties of 363 enzymes of the genus Arthrobacter, Bacillus, and Streptomyces (most abundant) showed that exoinulinases have an acid isoelectric point up to the neutral range due to their amino acid length. That is, the smaller the protein (336 aa), the more acidic the pI (4.39), and the larger the protein (1207 aa), the pI is in the neutral range (8.84). Also, a negative gravitational index indicates the hydrophilicity of exoinulinases. Finally, considering the biochemical properties affecting protein stability and post-translational changes studies, one enzyme for endoinulinase and 40 enzymes with desirable characteristics were selected to identify their enzyme production sources. To screen and isolate enzyme-containing strains, now with the expansion of databases and the development of bioinformatics tools, it is possible to classify, review and analyze a lot of data related to different enzyme-producing strains. Although, in laboratory studies, a maximum of 20 to 30 strains can be examined. Therefore, when more strains are examined, finally, strains with more stable and efficient enzymes were selected and introduced for laboratory activities. The findings of this study can help researchers to select the appropriate gene source from introduced strains for cloning and expression heterologous inulinase, or to extract native inulinase from introduced strains.

Proteomic analysis of wheat contrasting genotypes reveals the interplay between primary metabolic and regulatory pathways in anthers under drought stress.

Reproductive stage is very sensitive to various forms of environmental stresses such as drought stress. The proteomic analysis of anther during pollen development in response to drought stress was performed using a label-free quantitative shotgun proteomic technique to define the underlying molecular principles in two contrasting wheat genotypes Shiraz (susceptible) and D-10 (tolerant). Drought stress resulted in around two-fold decline in seed setting capacity and pollen viability in the Shiraz genotype compared to D-10. A Partial Least Square Discriminant Analysis (PLS-DA) of proteomic data revealed the abundance of 131 differentially abundant proteins significantly contributing in separation of drought tolerant and susceptible genotypes under normal and stress conditions. Proteins involved in cellular respiration, carbohydrate metabolism, RNA metabolism, and vesicle trafficking showed completely different responses in two genotypes. These proteins may maintain hexose pool and energy level and control regulation of transcription and transport. Furthermore, different members of functional groups such as protein biosynthesis and degradation, chromatin organization, and cytoskeleton dynamics were differentially abundant in response to stress in both genotypes which suggest their function in both genotypes to maintain minimum pollen viability/ fertility under drought stress. In conclusion, our findings revealed various metabolic and regulatory pathways underlying survival strategies required for pollen fertility and viability. SIGNIFICANCE: Drought caused by global climate change decreases cereal grain productivity worldwide. Yield losses due to water stress have been reported for major small grain cereal including wheat. Our findings highlighted the importance of key proteins in wheat adaptation to drought stress at reproductive stage. The obtained data showed that differentially abundant proteins in drought tolerant wheat genotype was remarkably associated with cellular respiration, carbohydrate metabolism, RNA metabolism, and vesicle trafficking. These results revealed fundamental data to elucidate the complexity of pollen fertility and viability under drought stress condition in wheat.

Plant isomiRs: origins, biogenesis, and biological functions.

MicroRNAs (miRNAs) are small endogenous non-coding RNAs in eukaryotes which regulate the expression of numerous genes post-transcriptionally, thereby playing critical roles in cells and organismal development. The high-throughput sequencing technologies enable the effective detection and annotation of miRNAs. Several miRNA variants with heterogeneous ends, lengths, and sequences can be generated from a single miRNA locus. Discovery of these miRNA variants, also known as miRNA isoforms or isomiRs, has made our understanding of the cells' miRNome deeper than previously pictured. Despite their wide presence in multiple datasets, the different possible origins and true biological significance of isomiRs are yet to be uncovered. Several recent emerging studies suggest that isomiRs are biologically active and non-randomly formed. This review aims to provide a comprehensive insight into the origins and biological importance of isomiRs, highlighting the enormous complexity of miRNA regulatory networks which broadens our knowledge about the post-transcriptional gene regulation in plants.

MicroRNAs regulate the main events in rice drought stress response by manipulating the water supply to shoots.

MicroRNAs (miRNAs) are small endogenous regulatory RNAs that are involved in a variety of biological processes related to proliferation, development, and response to biotic and abiotic stresses. miRNA profiles of rice (Oryza sativa L. cv. IR64.) leaves in a partial root zone drying (PRD) system were analysed using a high-throughput sequencing approach to identify miRNAs associated with drought signalling. The treatments performed in this study were as follows: well-watered ("wet" roots, WW), wherein both halves of the pot were watered daily; drought ("dry" roots, DD), wherein water was withheld from both halves of the pot; and well-watered/drought ("wet" and "dry" roots, WD), wherein one half of each pot was watered daily, the same as in WW, and water was withheld from the other part, the same as in DD. High-throughput sequencing enabled us to detect novel miRNAs and study the differential expression of known miRNAs. A total of 209 novel miRNAs were detected in this study. Differential miRNA profiling of the DD, WD and WW conditions showed differential expression of 159 miRNAs, among which 83, 44 and 32 miRNAs showed differential expression under both DD and WD conditions. The detection of putative targets of the differentially expressed miRNAs and investigation of their functions showed that most of these genes encode transcription factors involved in growth and development, leaf morphology, regulation of hormonal homeostasis, and stress response. The most important differences between the DD and WD conditions involved regulation of the levels of hormones such as auxin, cytokinin, abscisic acid, and jasmonic acid and also regulation of phosphor homeostasis. Overall, differentially expressed miRNAs under WD conditions were found to differ from those under DD conditions, with such differences playing a role in adaptation and inducing the normal condition. The mechanisms involved in regulating hormonal homeostasis and involved in energy production and consumption were found to be the most important regulatory pathways distinguishing the DD and WD conditions.

Drought responsive microRNAs in two barley cultivars differing in their level of sensitivity to drought stress.

MicroRNAs (miRNAs) are known to be involved in the regulation of gene expression, including that of genes involved in the response to stress. Here, a comparison has been drawn between the miRNA profiles of a drought susceptible, 'Morocco 9-75', and a drought tolerant, 'Yousef', barley cultivars. Leaf water content, shoot dry matter and chlorophyll content decreased in 'Morocco 9-75' more considerably compared with 'Yousef' under drought stress. Furthermore, lower stomatal conductance and higher leaf temperature were observed in 'Morocco 9-75' compared with 'Yousef'. Based on the criteria set for differential abundance, 118 of conserved and novel miRNAs were identified as being responsive to soil water status. Although drought stress resulted in an altered abundance of more miRNAs in 'Morocco 9-75' than in 'Yousef', drought stress was generally associated with an increased miRNA abundance in 'Yousef' and a decreased abundance in 'Morocco 9-75'. An in silico analysis identified 645 genes as putative targets for the drought-responsive miRNAs in 'Yousef' and 3735 in 'Morocco 9-75'. Gene ontology analysis showed that drought stress was associated with the altered abundance of miRNAs targeting growth, development, the juvenile to adult transition and hormone signaling. Some miRNAs which became more abundant in 'Yousef' are thought to target genes intimately involved in development and stress adaptation. In 'Morocco 9-75', drought stress induced changes in the abundance of miRNAs associated with genes affecting growth, development, the juvenile to adult transition and ABA signaling. The data imply that miRNAs may affect the tolerance/sensitivity of barley to drought stress by modulating the expression of a wide set of genes and induction of some physiological changes.

The contrasting microRNA content of a drought tolerant and a drought susceptible wheat cultivar.

Drought stress represents one of the most common stresses affecting the productivity of crop plants. A rather recently discovered component of the plant response to drought is the cellular population of microRNAs. Here, the microRNA content was revealed of two bread wheat cultivars contrasting strongly with respect to the ability to withstand drought stress. A total of 1813 miRNAs was identified, grouped into 106 families. Some 104 of these miRNAs were predicted to match 212 novel miRNA precursors. In the drought tolerant cultivar (SM), 105 (33 known and 72 novel) miRNAs were altered in abundance by the imposition of drought stress, while the equivalent number in the more sensitive cultivar (SW) was 51 (20 and 31). An in silico analysis predicted that these miRNAs target at least 1959 genes in SM and 1111 in SW, suggesting their broad contribution to the drought stress response. Among the target genes were several known stress-related genes, encoding, for example, superoxide dismutase, various MYB transcription factors, various ABA signaling proteins and various MADS-box transcription factors. In many cases, the more susceptible cultivar SW behaved in a contrasting manner. The suggestion is that miRNAs represent an important aspect of the drought stress response, post-transcriptionally regulating a range of stress-related genes.