Redox-engineering enhances maize thermotolerance and grain yield in the field

Contains fulltext : 252904.pdf (Publisher’s version ) (Open Access)08 juli 202

Tomato class II glutaredoxin mutants generated via multiplex CRISPR/Cas9 genome editing technology are susceptible to multiple abiotic stresses

Doctor of PhilosophyDepartment of Horticulture and Natural ResourcesSunghun ParkGene silencing technologies such as clustered regulatory short palindromic repeats (CRISPR)/Cas9 and RNA interference (RNAi) created a revolution in genome engineering. They are highly site-specific, simple, fast, and cost-effective. Since their discovery, gene silencing technologies have extensively been implemented in various organisms including humans, animals, plants, and microbes. They have been used for both basic science studies such as gene functional analysis and applied science such as medicine and crop improvement. In this work, we used multiplex CRISPR/Cas9 system to knock out all four members of the class II glutaredoxin (GRX) gene family including S14, S15, S16 and S17 in tomato and RNAi technology to express mouse complement 3 (C3) and complement factor 7 (CF7) small interfering RNAs (siRNAs) in lettuce. Reactive oxygen species (ROS) are induced under abiotic and biotic stresses and also as byproducts of aerobic metabolism, and their overproduction causes oxidative damage to macromolecules such as proteins, lipids, carbohydrates, and nucleic acids. GRXs are small ubiquitous proteins that are known to be involved in cellular redox homeostasis by reducing disulfate bonds and scavenging ROS. To investigate the functions of each member of class II GRX gene family in tomato’s response to abiotic stresses, single and multiple knockout lines for class II GRXs were obtained using multiplex CRISPR/Cas9 system. Mutant lines and wild-type plants were subjected to heat, drought, chilling, cadmium (Cd) toxicity, and short photoperiod stresses. Phenotyping data showed that members of the class II GRX gene family are critical for tomato’s growth, development, and survival under several abiotic stresses. Our findings propose novel functions for members of class II SlGRXs. RNAi technology can be utilized to target disease-related proteins. However, the application of siRNAs is challenging predominantly due to the difficult delivery and instability of siRNA into the host system. Recent findings of bioactivity and bioavailability of plants’ miRNAs through animals’ digestive systems led to the newly introduced field of dietary siRNAs. Animal’s siRNA can be expressed in plant tissues and delivered as dietary siRNA. Here, an expression vector made based on a rice miRNA backbone, Osa-MIR528, was utilized to construct two plant expression vectors containing siRNAs silencing mouse C3 and CF7 proteins. Both C3 and CF7 proteins are involved in blood clotting which could lead to cardiovascular dysfunction. Expression of both primary and mature C3 and CF7 siRNAs in lettuce was validated by semi quantitative real-time PCR and end-point PCR, respectively, and was confirmed via Sanger sequencing. Established amiRNA system in lettuce through this work will have further applications. As an edible leafy plant with high biomass, lettuce can be used as a valuable host to produce various diseases targeting siRNAs

Redox potential tuning in bio-relevant heterocycles via (anti)aromaticity modulated H-bonding (AMHB)

Hydrogen bonds are arguably the most important non-covalent interactions in chemistry and biology, and their strength and directionality have been elegantly exploited in the rational design of complex structures. We recently noted that the variable responses of cyclic π-systems upon H-bond formation reciprocally lead to modulations of the H-bonds’ strengths, a phenomenon that we dubbed (anti)aromaticity-modulated hydrogen bonding (AMHB) [J. Am. Chem. Soc. 2016, 138, 3427–3432]. Species that switch from aromatic to antiaromatic or vice versa upon changing π-electron counts should be oppositely stabilized by the AMHB effects, so their redox potentials should be significantly “tuned” by H-bond formation. Herein, using quantum chemical simulations, we explore the effects of these H-bond induced π-electron polarizations on the redox potentials of (anti)aromatic heterocycles. The systems chosen for this study have embedded amide groups and amidine moieties capable of forming two-point H-bonds in their cyclic π-systems. Thus, as the 4-electron and 6-electron π-systems in redox-capable monocycles (e.g., quinones) can be differentially stabilized, their redox potentials can be modulated by H-bond formation by as much as 6 kcal/mol (258 mV for one electron transfer). In fused rings, the connectivity patterns are as important as the π-electron counts. Extending these ideas to flavin, a biologically relevant case, we find that H-bonding patterns like those found in its crystals can vary its redox potential by up to 1.3 kcal/mol.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

I need closure: Is hyperbaric oxygen therapy better than skin substitute therapy for chronic wounds?

Individuals with diabetes mellitus deal with chronic wounds that are associated with poor health outcomes and frequently result in immobility, infections, and amputations. For these patients, there is a significant amount of literature on the use of oxygen therapy and synthetic skin substitute therapy in promoting chronic wound healing. However, there is limited published evidence comparing the effectiveness of the two therapies. Our goal is to summarize existing evidence of how hyperbaric oxygen therapy and substitute skin therapy affect wound healing in 9 months. We use the PRISMA model to guide our review of literature found in the Nursing & Allied Health Database using hyperbaric oxygen therapy, skin substitute therapy, chronic wounds, and clinical studies to retrieve relevant studies. We narrowed our review of literature to 22 studies based on criteria that required no affiliations between therapies, manufacturers, or brands. We also reduced inclusion of literature that rated high with selection, performance, and reporting bias. With scholarship from different countries around the world dealing with patient populations with chronic wounds, our review yielded mixed results. Few studies reported hyperbaric oxygen therapy as a primary therapy for treating chronic wounds while others indicated no significant differences between the two. We recommend that more research and development on hyperbaric oxygen therapy and skin substitute therapy is necessary to determine the best options and clinical outcomes for a growing population of patients needing our best in medicine and treatment

Expression analyses of salinity stress- associated ESTs in Aeluropus littoralis

Salinity is among the most important abiotic stresses affecting crop production throughout the earth. Halophyte plants can sustain high salinity levels, therefore elucidating molecular mechanisms underlying their salinity resistance is beneficial for crop improvement. Aeluropus littoralis, a halophyte weed, is a great genetic resource for this purpose. Isolated expressed sequence taq (EST) sequences from A. littoralis under salinity stress, have given us the chance to find and analyze transcripts of genes involved in response to salinity. Transcriptome analyses indicated the expression levels of mRNAs corresponding to 10 of sequences were increased under treatments. All mRNAs were significantly induced under salt treatment with the highest peaks observed at different hours of treatments. Moreover, the full-length cDNA of vacuolar H+-pyrophosphatase (VP) was isolated utilizing 3′ and 5′ rapid amplification of cDNA ends polymerase chain reaction (RACE-PCR) and characterized (GenBank accession number of KT253223.1). The extracted full-length of VP was 2732 bp, which contained ORF of 2292 bp encoding 763 amino acids.</p

AMHB: (Anti)aromaticity-Modulated Hydrogen Bonding

This <i>in silico</i> survey shows that changes in the (anti)­aromatic character of π-conjugated heterocycles can be used to fine-tune their hydrogen (H-)­bond strengths. Upon H-bonding dimerization, the π-electrons of these rings can be polarized to reinforce or disrupt their (anti)­aromatic π-conjugated circuits (πCCs) and stabilize or destabilize the resulting H-bonded complexes. H-bonding interactions that enhance aromaticity or relieve antiaromaticity are fortified, whereas those that intensify antiaromaticity or disrupt aromaticity are weakened, relative to analogues lacking full π-circuits. Computed dissected nucleus-independent chemical shifts, NICS(1)_<i>zz</i>, reveal a uniform pattern and document changes in the magnetic (anti)­aromatic character of the heterocycles considered. Recognition of this (anti)­aromaticity-modulated H-bonding (AMHB) phenomenon offers insights into a range of fields from organocatalysis and self-assembly to pharmaceutical chemistry and molecular biology

Ectopic Expression of a Heterologous Glutaredoxin Enhances Drought Tolerance and Grain Yield in Field Grown Maize

Drought stress is a major constraint in global maize production, causing almost 30–90% of the yield loss depending upon growth stage and the degree and duration of the stress. Here, we report that ectopic expression of Arabidopsis glutaredoxin S17 (AtGRXS17) in field grown maize conferred tolerance to drought stress during the reproductive stage, which is the most drought sensitive stage for seed set and, consequently, grain yield. AtGRXS17-expressing maize lines displayed higher seed set in the field, resulting in 2-fold and 1.5-fold increase in yield in comparison to the non-transgenic plants when challenged with drought stress at the tasseling and silking/pollination stages, respectively. AtGRXS17-expressing lines showed higher relative water content, higher chlorophyll content, and less hydrogen peroxide accumulation than wild-type (WT) control plants under drought conditions. AtGRXS17-expressing lines also exhibited at least 2-fold more pollen germination than WT plants under drought stress. Compared to the transgenic maize, WT controls accumulated higher amount of proline, indicating that WT plants were more stressed over the same period. The results present a robust and simple strategy for meeting rising yield demands in maize under water limiting conditions

Expression of a monothiol glutaredoxin, AtGRXS17, in tomato (Solanum lycopersicum) enhances drought tolerance

Abiotic stresses are a major factor limiting crop growth and productivity. The Arabidopsis thaliana glutaredoxin S17 (AtGRXS17) gene has conserved functions in plant tolerance to heat and chilling stress in Arabidopsis and, when expressed ectopically, in tomato. Here, we report that ectopic expression of AtGRXS17 in tomato also enhanced tolerance to drought and oxidative stress. AtGRXS17-expressing tomato plants contained twice the shoot water content compared to wild-type plants under water limiting conditions. This enhanced drought tolerance correlated with a higher maximal photosynthetic efficiency of photosystem II (Fv/Fm). Ectopic AtGRXS17-expression was concomitant with the expression of Solanum lycopersicum catalase 1 (SlCAT1) and mitigated defects in the growth of primary roots in response to methyl viologen exposure. In addition, AtGRXS17 expression was found to prolong elevated expression levels of the Solanum lycopersicum ABA-responsive element binding protein 1 (SlAREB1) during drought stress. The findings demonstrate that expression of AtGRXS17 can simultaneously improve the tolerance of tomato, and possibly other agriculturally important crops, to drought, heat, and chilling stresses