■ Abstract Apoptosis is a genetically programmed, morphologically distinct form of cell death that can be triggered by a variety of physiological and pathological stimuli. Studies performed over the past 10 years have demonstrated that proteases play critical roles in initiation and execution of this process. The caspases, a family of cysteine-dependent aspartate-directed proteases, are prominent among the death proteases. Caspases are synthesized as relatively inactive zymogens that become activated by scaffold-mediated transactivation or by cleavage via upstream proteases in an intracellular cascade. Regulation of caspase activation and activity occurs at several different levels: ( a) Zymogen gene transcription is regulated; ( b) antiapoptotic members of the Bcl-2 family and other cellular polypeptides block proximity-induced activation of certain procaspases; and ( c) certain cellular inhibitor of apoptosis proteins (cIAPs) can bind to and inhibit active caspases. Once activated, caspases cleave a variety of intracellular polypeptides, including major structural elements of the cytoplasm and nucleus, components of the DNA repair machinery, and a number of protein kinases. Collectively, these scissions disrupt survival pathways and disassemble important architectural components of the cell, contributing to the stereotypic morphological and biochemical changes that characterize apoptotic cell death.

Mammalian caspases: structure ,a ctivation ,s ubstrates, and functions during apoptosis

Besides improved mineral nutrition, plants colonised by arbuscular mycorrhizal (AM) fungi often display increased biomass and higher tolerance to biotic and abiotic stresses. Notwithstanding the global importance of wheat as an agricultural crop, its response to AM symbiosis has been poorly investigated. We focused on the role of an AM fungus on mineral nutrition of wheat, and on its potential protective effect against Xanthomonas translucens. To address these issues, phenotypical, molecular and metabolomic approaches were combined. Morphological observations highlighted that AM wheat plants displayed an increased biomass and grain yield, as well as a reduction in lesion area following pathogen infection. To elucidate the molecular mechanisms underlying the mycorrhizal phenotype, we investigated changes of transcripts and proteins in roots and leaves during the double (wheat-AM fungus) and tripartite (wheat-AM fungus-pathogen) interaction. Transcriptomic and proteomic profiling identified the main pathways involved in enhancing plant biomass, mineral nutrition and in promoting the bio-protective effect against the leaf pathogen. Mineral and amino acid contents in roots, leaves and seeds, and protein oxidation profiles in leaves, supported the omics data, providing new insight into the mechanisms exerted by AM symbiosis to confer stronger productivity and enhanced resistance to X. translucens in wheat.

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Omics approaches revealed how arbuscular mycorrhizal symbiosis enhances yield and resistance to leaf pathogen in wheat

Climate change gives rise to numerous environmental stresses, including soil salinity. Salinity/salt stress is the second biggest abiotic factor affecting agricultural productivity worldwide by damaging numerous physiological, biochemical, and molecular processes. In particular, salinity affects plant growth, development, and productivity. Salinity responses include modulation of ion homeostasis, antioxidant defense system induction, and biosynthesis of numerous phytohormones and osmoprotectants to protect plants from osmotic stress by decreasing ion toxicity and augmented reactive oxygen species scavenging. As most crop plants are sensitive to salinity, improving salt tolerance is crucial in sustaining global agricultural productivity. In response to salinity, plants trigger stress-related genes, proteins, and the accumulation of metabolites to cope with the adverse consequence of salinity. Therefore, this review presents an overview of salinity stress in crop plants. We highlight advances in modern biotechnological tools, such as omics (genomics, transcriptomics, proteomics, and metabolomics) approaches and different genome editing tools (ZFN, TALEN, and CRISPR/Cas system) for improving salinity tolerance in plants and accomplish the goal of "zero hunger," a worldwide sustainable development goal proposed by the FAO.

/pdf/smart-reprograming-of-plants-against-salinity-stress-using-1ob0lsyv.pdf

Smart reprograming of plants against salinity stress using modern biotechnological tools.

The ATP-binding cassette (ABC) transporters belong to a large protein family predominantly present in diverse species. ABC transporters are driven by ATP hydrolysis and can act as exporters as well as importers. These proteins are localized in the membranes of chloroplasts, mitochondria, peroxisomes and vacuoles. ABC proteins are involved in regulating diverse biological processes in plants, such as growth, development, uptake of nutrients, tolerance to biotic and abiotic stresses, tolerance to metal toxicity, stomatal closure, shape and size of grains, protection of pollens, transport of phytohormones, etc. In mitochondria and chloroplast, the iron metabolism and its transport across the membrane are mediated by ABC transporters. Tonoplast-localized ABC transporters are involved in internal detoxification of metal ion; thus protecting against the DNA impairment and maintaining cell growth. ABC transporters are involved in the transport of secondary metabolites inside the cells. Microorganisms also engage a large number of ABC transporters to import and expel substrates decisive for their pathogenesis. ABC transporters also suppress the seed embryonic growth until favorable conditions come. This review aims at giving insights on ABC transporters, their evolution, structure, functions and roles in different biological processes for helping the terrestrial plants to survive under adverse environmental conditions. These specialized plant membrane transporters ensure a sustainable economic yield and high-quality products, especially under unfavorable conditions of growth. These transporters can be suitably manipulated to develop 'Plants for the Future'.

Role of ATP-binding cassette transporters in maintaining plant homeostasis under abiotic and biotic stresses

(+)-Valencene and (+)-nootkatone are high value-added sesquiterpenoids found in grapefruit. The synthesis of (+)-nootkatone by chemical oxidation from (+)-valencene cannot meet the increasing demand in natural aromatics markets. Development of a viable bioprocess using microorganisms is attractive. According to the yields of β-nootkatol and (+)-nootkatone by strains harboring different expression cassettes in the resting cell assay, premnaspirodiene oxygenase from Hyoscyamus muticus (HPO), cytochrome P450 reductase from Arabidopsis thaliana (AtCPR) and alcohol dehydrogenase (ADH1) from Saccharomyces cerevisiae were finally selected and overexpressed in CEN·PK2-1Ca, yielding β-nootkatol and (+)-nootkatone with 170.5 and 45.6 mg L−1 ethyl acetate, respectively. A combinational engineering strategy including promoter change, regulator ROX1 knockout, squalene pathway inhibition, and tHMGR overexpression was performed to achieve de novo (+)-valencene production. Subsequent culture investigations found that galactose as the induced carbon source and a lower temperature (25 °C) were beneficial to target accumulation. Also, replacing the inducible promoters (GAL1) of HPO and AtCPR with constitutive promoters (HXT7 and CYC1) dramatically increased the β-nootkatol accumulation from 108.2 to 327.8 mg L−1 ethyl acetate in resting-cell experiments using (+)-valencene as a substrate. Finally, the total terpenoid titer of the engineered strain of PK2-25 using glucose as a carbon source was improved to 157.8 mg L−1 cell culture, which was 56 times the initial value. We present a new candidate for production of (+)-valencene and its related sesquiterpenoids with attraction for industry.

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Stepwise engineering of Saccharomyces cerevisiae to produce (+)-valencene and its related sesquiterpenes

The primary goals of rice breeding programs are grain quality and yield potential improvement. With the high demand for rice varieties of premium cooking and eating quality, we developed low-amylose content breeding lines crossed with Samgwang and Milkyqueen through the marker-assisted backcross (MABc) breeding program. Trait markers of the SSIIIa gene referring to low-amylose content were identified through an SNP mapping activity, and the markers were applied to select favorable lines for a foreground selection. To rapidly recover the genetic background of Samgwang (recurrent parent genome, RPG), 386 genome-wide markers were used to select BC1F1 and BC2F1 individuals. Seven BC2F1 lines with targeted traits were selected, and the genetic background recovery range varied within 97.4-99.1% of RPG. The amylose content of the selected BC2F2 grains ranged from 12.4-16.8%. We demonstrated the MABc using a trait and genome-wide markers, allowing us to efficiently select lines of a target trait and reduce the breeding cycle effectively. In addition, the BC2F2 lines confirmed by molecular markers in this study can be utilized as parental lines for subsequent breeding programs of high-quality rice for cooking and eating.

https://www.mdpi.com/2223-7747/10/4/804/pdf

Breeding of High Cooking and Eating Quality in Rice by Marker-Assisted Backcrossing (MABc) Using KASP Markers.

Proteins encoded by U-box type ubiquitin ligase (PUB) genes in rice are known to play an important role in plant responses to abiotic and biotic stresses. Functional analysis has revealed a detailed molecular mechanism involving PUB proteins in relation to abiotic and biotic stresses. In this study, characteristics of 77 OsPUB genes in rice were identified. Systematic and comprehensive analyses of the OsPUB gene family were then performed, including analysis of conserved domains, phylogenetic relationships, gene structure, chromosome location, cis-acting elements, and expression patterns. Through transcriptome analysis, we confirmed that 16 OsPUB genes show similar expression patterns in drought stress and blast infection response pathways. Numerous cis-acting elements were found in promoter sequences of 16 OsPUB genes, indicating that the OsPUB genes might be involved in complex regulatory networks to control hormones, stress responses, and cellular development. We performed qRT-PCR on 16 OsPUB genes under drought stress and blast infection to further identify the reliability of transcriptome and cis-element analysis data. It was confirmed that the expression pattern was similar to RNA-sequencing analysis results. The transcription of OsPUB under various stress conditions indicates that the PUB gene might have various functions in the responses of rice to abiotic and biotic stresses. Taken together, these results indicate that the genome-wide analysis of OsPUB genes can provide a solid basis for the functional analysis of U-box E3 ubiquitin ligase genes. The molecular information of the U-box E3 ubiquitin ligase gene family in rice, including gene expression patterns and cis-acting regulatory elements, could be useful for future crop breeding programs by genome editing.

Molecular and Functional Analysis of U-box E3 Ubiquitin Ligase Gene Family in Rice (Oryzasativa).

Cysteine protease activity comprises the majority of proteolytic activities in plants. They are involved in almost every facet of the plant’s development. Accumulating evidence indicates multiple roles of this protease type in response to biotic and abiotic stress. To understand the regulations and functions of cysteine protease in rice, its evolutionary and structural evidence was uncovered in this study. Using MEROPS, a peptidase database, the 74 rice cysteine proteases belonging to six families were queried. Each of these families represents distinct proteolytic enzyme; C1 is a papain-like protease, C2 is a calpain-2-type, C12 is an ubiquitinyl hydrolase-L1 enzyme, C13 is legumain, C14 is a caspase-1 type, and C15 is a pyroglutamyl peptidase 1 enzyme type. Evolutionary expansion attributed to gene duplication and diversification was particularly evident in C1 family which showed the highest number (n = 53) of members, most of which contained the highest number and most variable introns and motifs, whereas families C13, C14, and C15 had only a few members which all contained lesser number and variation of intron and motif. Out of 74 total cysteine protease gene members, 73 were globular proteins and 55 were predicted as stable proteins. Spatial expression assay of selected C1 members showed that LOC_Os01g73980 and LOC_Os05g01810 were highly expressed in the stem and leaves, while LOC_Os02g27030 was constitutively expressed in all tissues. The expression of LOC_Os01g73980 and LOC_Os05g01810 was also highly activated by salinity stress, while LOC_Os02g27030 was activated by both salinity and heat. LOC_Os05g01810 overexpression transgenic rice exhibited moderate tolerance to salinity stress, which provides interesting clues on biological functions of these genes in rice.

Genome-wide identification and molecular characterization of cysteine protease genes in rice

Brown rice is composed of rice bran, pericarp, seed coat, and aleurone layers, and the rice bran layer contains a large number of substances useful for the human body, such as dietary fiber, α-tocopherol, α-tocotrienol, and vitamins. However, more than 90% of these substances are removed when polished, and white rice has the disadvantage of losing food-related ingredients, such as umami-related amino acids, when compared to the unpolished group. In this study, we tried to develop new breeding lines with a thinner seed coat and aleurone layer to provide high eating quality with softer chewing characteristics and processability in rice grain. We detected an SNP for foreground selection for the backcross population by comparing genome sequences between Samgwang and Seolgaeng and developed high eating quality brown rice breeding lines by applying marker-assisted backcrossing (MABC) breeding programs to backcross populations between Samgwang and Seolgaeng using KASP markers. SNP markers for foreground selection were identified to improve eating and processability through SNP mapping of Samgwang and Seolgaeng with SSIIa as a target gene in this study. Line selection according to genotype of KASP markers was successful in BC1F1 and BC2F1 generations, with the recurrent parent genome recovery ratio ranging from 91.22% to 98.65%. In BC2F1 seeds of the selected lines, thickness of the aleurone layer was found to range from 13.82 to 21.67 μm, which is much thinner than the 30.91 μm of the wild type, suggesting that selection by MABc could be used as an additional breeding material for the development of highly processed rice varieties. These lines will be useful to develop new brown rice varieties with softer chewing characteristics and processability in rice grain.

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Marker-Assisted Backcrossing (MABc) to Improve Eating Quality with Thin Seed Coat and Aleurone Layer of Non-Glutinous Japonica Variety in Rice

In plants, the orange (Or) gene plays roles in regulating carotenoid biosynthesis and responses to environmental stress. The present study investigated whether the expression of rice Or (OsOr) gene could enhance rice tolerance to heat stress conditions. The OsOr gene was cloned and constructed with OsOr or OsOr-R115H (leading to Arg to His substitution at position 115 on the OsOr protein), and transformed into rice plants. The chlorophyll contents and proline contents of transgenic lines were significantly higher than those of non-transgenic (NT) plants under heat stress conditions. However, we found that the levels of electrolyte leakage and malondialdehyde in transgenic lines were significantly reduced compared to NT plants under heat stress conditions. In addition, the levels of expression of four genes related to reactive oxygen species (ROS) scavenging enzymes (OsAPX2, OsCATA, OsCATB, OsSOD-Cu/Zn) and five genes (OsLEA3, OsDREB2A, OsDREB1A, OsP5CS, SNAC1) responded to abiotic stress was showed significantly higher in the transgenic lines than NT plants under heat stress conditions. Therefore, OsOr-R115H could be exploited as a promising strategy for developing new rice cultivars with improved heat stress tolerance.

Me-Sun Kim

Papers

Breeding of High Cooking and Eating Quality in Rice by Marker-Assisted Backcrossing (MABc) Using KASP Markers.

Molecular and Functional Analysis of U-box E3 Ubiquitin Ligase Gene Family in Rice (Oryzasativa).

Genome-wide identification and molecular characterization of cysteine protease genes in rice

Marker-Assisted Backcrossing (MABc) to Improve Eating Quality with Thin Seed Coat and Aleurone Layer of Non-Glutinous Japonica Variety in Rice

Overexpression of Orange Gene (OsOr-R115H) Enhances Heat Tolerance and Defense-Related Gene Expression in Rice (Oryza sativa L.)