Physiological traits associated with heat tolerance in bread wheat (Triticum aestivum L.)

Field experiments for evaluating heat tolerance-related physiological traits were conducted for two consecutive years using a mapping population of recombinant inbred lines (RILs) from the cross RAJ4014/WH730. Chlorophyll content (Chl) and chlorophyll fluorescence (CFL) were recorded under timely sown (TS) and late sown (LS) conditions. Late sowing exposes the terminal stage of plants to high temperature stress. Pooled analysis showed that CFL and Chl differed significantly under TS and LS conditions. The mean value of CFL (Fv/Fm) and Chl under both timely and late sown conditions were used as physiological traits for association with markers. Regression analysis revealed significant association of microsatellite markers viz., Xpsp3094 and Xgwm131 with coefficients of determination (R²) values for CFL (Fv/Fm) and Chl as 12 and 8 %, respectively. The correlation between thousand grain weight (TGW) with Chl and CFL were 14 and 7 % and correlation between grain wt./spike with Chl and CFL were 15 and 8 %, respectively. The genotypes showing tolerance to terminal heat stress as manifested by low heat susceptibility index (HSI = 0.43) for thousand grain weight, were also found having very low Chl, HSI (−0.52). These results suggest that these physiological traits may be used as a secondary character for screening heat-tolerant genotypes.     

The use of stable carbon isotopes to decipher global change effects on soil organic carbon: present status,limitations, and future prospects

Major global change factors, including carbon dioxide (CO₂) fertilization, warming, change in precipitation, nitrogen deposition, and land-use change have the potential to significantly affect future stocks of soil organic carbon (SOC). These factors, individually or by interacting with each other, can also trigger positive or negative feedback to the processes affecting the rate of SOC formation or loss. Despite rapid progress in the understanding of carbon (C) cycling processes in the last few decades, much uncertainty remains in our ability to precisely forecast potential changes in SOC stocks in the rapidly changing future world. Stable C isotopes have been extensively used in natural observational studies as well as in laboratory and field experiments that manipulate CO₂ concentration, temperature, moisture, nitrogen fertilization, and vegetation type to understand the complex interactions and feedbacks that result from changing climate, plants and their herbivores, as well as soil microorganisms. Newly developed tools such as compound-specific isotope analysis, nano-SIMS (secondary ion mass spectroscopy), and stable isotope probing (SIP) permit isotope tracing in a specific ecosystem pool into specific C compounds and processes, thus providing in-depth insights into many processes affecting C biogeochemistry. The recent availability of affordable and reliable field-deployable optical isotope monitoring devices has provided researchers with a new set of tools for continuously tracking the ¹³C-CO₂ fluxes at the ecosystem level, enabling deeper insights into C biogeochemistry under changing environmental conditions. Despite these great strides, there is a scarcity of review studies that have comprehensively examined the use of C isotopes in studying SOC responses under global change factors. This review highlights recent progress in understanding the effect of major global change factors on SOC fluxes and stocks using selected examples covering scales from plant rhizospheres to geographic regions. Moreover, we discuss the strengths and limitations of current approaches and recent scientific advancements to highlight the new prospects evolving from the exceptional temporal and spatial resolution of stable isotope analysis in studying how global change affects SOC. Finally, we suggest that studies using stable C isotopes are well-poised to focus on identifying how dominant SOC cycling processes respond to environment-specific limiting factors and any thresholds and tipping points that define those relationships.

     

Active displacement of RecA filaments by UvrD translocase activity

The UvrD helicase has been implicated in the disassembly of RecA nucleoprotein filaments in vivo and in vitro. We demonstrate that UvrD utilizes an active mechanism to remove RecA from the DNA. Efficient RecA removal depends on the availability of DNA binding sites for UvrD and/or the accessibility of the RecA filament ends. The removal of RecA from DNA also requires ATP hydrolysis by the UvrD helicase but not by RecA protein. The RecA-removal activity of UvrD is slowed by RecA variants with enhanced DNA-binding properties. The ATPase rate of UvrD during RecA removal is much slower than the ATPase activity of UvrD when it is functioning either as a translocase or a helicase on DNA in the absence of RecA. Thus, in this context UvrD may operate in a specialized disassembly mode.     

Glycosylation of sputum mucins is altered in cystic fibrosis patients

Cystic fibrosis (CF) is characterized by chronic lung infection and inflammation, with periods of acute exacerbation causing severe and irreversible lung tissue damage. We used protein and glycosylation analysis of high-molecular mass proteins in saline-induced sputum from CF adults with and without an acute exacerbation, CF children with stable disease and preserved lung function, and healthy non-CF adult and child controls to identify potential biomarkers of lung condition. While the main high-molecular mass proteins in the sputum from all subjects were the mucins MUC5B and MUC5AC, these appeared degraded in CF adults with an exacerbation. The glycosylation of these mucins also showed reduced sulfation, increased sialylation, and reduced fucosylation in CF adults compared with controls. Despite improvements in pulmonary function after hospitalization, these differences remained. Two CF children showed glycoprotein profiles similar to those of CF adults with exacerbations and also presented with pulmonary flares shortly after sampling, while the remaining CF children had profiles indistinguishable from those of healthy non-CF controls. Sputum mucin glycosylation and degradation are therefore not inherently different in CF, and may also be useful predictive biomarkers of lung condition.     

Deciphering the genetic architecture and ethnographic distribution of IRD in three ethnic populations by whole genome sequence analysis

Patients with inherited retinal dystrophies (IRDs) were recruited from two understudied populations: Mexico and Pakistan as well as a third well-studied population of European Americans to define the genetic architecture of IRD by performing whole-genome sequencing (WGS). Whole-genome analysis was performed on 409 individuals from 108 unrelated pedigrees with IRDs. All patients underwent an ophthalmic evaluation to establish the retinal phenotype. Although the 108 pedigrees in this study had previously been examined for mutations in known IRD genes using a wide range of methodologies including targeted gene(s) or mutation(s) screening, linkage analysis and exome sequencing, the gene mutations responsible for IRD in these 108 pedigrees were not determined. WGS was performed on these pedigrees using Illumina X10 at a minimum of 30X depth. The sequence reads were mapped against hg19 followed by variant calling using GATK. The genome variants were annotated using SnpEff, PolyPhen2, and CADD score; the structural variants (SVs) were called using GenomeSTRiP and LUMPY. We identified potential causative sequence alterations in 61 pedigrees (57%), including 39 novel and 54 reported variants in IRD genes. For 57 of these pedigrees the observed genotype was consistent with the initial clinical diagnosis, the remaining 4 had the clinical diagnosis reclassified based on our findings. In seven pedigrees (12%) we observed atypical causal variants, i.e. unexpected genotype(s), including 4 pedigrees with causal variants in more than one IRD gene within all affected family members, one pedigree with intrafamilial genetic heterogeneity (different affected family members carrying causal variants in different IRD genes), one pedigree carrying a dominant causative variant present in pseudo-recessive form due to consanguinity and one pedigree with a de-novo variant in the affected family member. Combined atypical and large structural variants contributed to about 20% of cases. Among the novel mutations, 75% were detected in Mexican and 50% found in European American pedigrees and have not been reported in any other population while only 20% were detected in Pakistani pedigrees and were not previously reported. The remaining novel IRD causative variants were listed in gnomAD but were found to be very rare and population specific. Mutations in known IRD associated genes contributed to pathology in 63% Mexican, 60% Pakistani and 45% European American pedigrees analyzed. Overall, contribution of known IRD gene variants to disease pathology in these three populations was similar to that observed in other populations worldwide. This study revealed a spectrum of mutations contributing to IRD in three populations, identified a large proportion of novel potentially causative variants that are specific to the corresponding population or not reported in gnomAD and shed light on the genetic architecture of IRD in these diverse global populations.     

Pigment Diverse Mutants of Pseudomonas sp.: Inhibition of Fungal Growth and Stimulation of Growth of Cicer arietinum

A Pseudomonas strain MRS16 inhibited growth of different pathogenic fungi (Aspergillus sp., Fusarium oxysporum, Pythium aphanidermatum and Rhizoctonia solani) in vitro. Larger inhibition zones were obtained on nutrient agar and King's B media compared to potato dextrose agar and pigment production media. Mutants altered in production of fluorescent pigment were derived by nitrosoguanidine mutagenesis. The pigment overproducer mutant MRS16M-1 was more inhibitory whereas nonproducer mutant MRS16M-5 was less inhibitory than parent strain on nutrient agar medium. Addition of iron (100 µM FeCl₃) in the medium decreased inhibition of fungal growth, suggesting the involvement of siderophores and other antifungal secondary metabolites. Seed bacterization of two cultivars of chickpea (Cicer arietinum cvs. H8618 and C235) differing in susceptibility to wilt caused initial root and shoot stunting at 5 d of growth followed by proliferation of secondary root growth at 10 d. Coinoculation of chickpea with Pseudomonas strain MRS16 or mutants and Rhizobium sp. Cicer strain Ca181 enhanced nodulation, nitrogen fixation and plant dry mass as compared to single inoculation with Rhizobium strain under sterile conditions.     

Illuminating the Gateway of Gene Silencing: Perspective of RNA Interference Technology in Clinical Therapeutics

A novel laboratory revolution for disease therapy, the RNA interference (RNAi) technology, has adopted a new era of molecular research as the next generation "Gene-targeted prophylaxis." In this review, we have focused on the chief technological challenges associated with the efforts to develop RNAi-based therapeutics that may guide the biomedical researchers. Many non-curable maladies, like neurodegenerative diseases and cancers have effectively been cured using this technology. Rapid advances are still in progress for the development of RNAi-based technologies that will be having a major impact on medical research. We have highlighted the recent discoveries associated with the phenomenon of RNAi, expression of silencing molecules in mammals along with the vector systems used for disease therapeutics.     

Inhibitor of diamine oxidase in cotyledons of groundnut seedlings

The apparent absence of diamine oxidase in extracts of cotyledon of germinating groundnut seeds is due to the presence of a natural inhibitor. The inhibitor was associated with the lipid in the upper layer obtained on centrifugation of the cotyledon extract. It was non-dialysable, thermolabile and was inactivated by TCA and Triton X-100. The inhibitor was detected in the cotyledon extracts from early stages of seed development and was present up to 20 days after germination.     

Metformin: A Potential Therapeutic Agent for Recurrent Colon Cancer

Accumulating evidence suggests that metformin, a biguanide class of anti-diabetic drugs, possesses anti-cancer properties. However, most of the studies to evaluate therapeutic efficacy of metformin have been on primary cancer. No information is available whether metformin could be effectively used for recurrent cancer, specifically colorectal cancer (CRC) that affects up to 50% of patients treated by conventional chemotherapies. Although the reasons for recurrence are not fully understood, it is thought to be due to re-emergence of chemotherapy-resistant cancer stem/stem-like cells (CSCs/CSLCs). Therefore, development of non-toxic treatment strategies targeting CSCs would be of significant therapeutic benefit.In the current investigation, we have examined the effectiveness of metformin, in combination with 5-fluorouracil and oxaliplatin (FuOx), the mainstay of colon cancer therapeutics, on survival of chemo-resistant colon cancer cells that are highly enriched in CSCs/CSLCs. Our data show that metformin acts synergistically with FuOx to (a) induce cell death in chemo resistant (CR) HT-29 and HCT-116 colon cancer cells, (b) inhibit colonospheres formation and (c) enhance colonospheres disintegration. In vitro cell culture studies have further demonstrated that the combinatorial treatment inhibits migration of CR colon cancer cells. These changes were associated with increased miRNA 145 and reduction in miRNA 21. Wnt/β-catenin signaling pathway was also down-regulated indicating its pivotal role in regulating the growth of CR colon cancer cells. Data from SCID mice xenograft model of CR HCT-116 and CR HT-29 cells show that the combination of metformin and FuOX is highly effective in inhibiting the growth of colon tumors as evidenced by ∼50% inhibition in growth following 5 weeks of combination treatment, when compared with the vehicle treated controls. Our current data suggest that metformin together with conventional chemotherapy could be an effective treatment regimen for recurring colorectal cancer (CRC).     

Surgical Technique for the Implantation of Tissue Engineered Vascular Grafts and Subsequent In Vivo Monitoring

The development of Tissue Engineered Vessels (TEVs) is advanced by the ability to routinely and effectively implant TEVs (4-5 mm in diameter) into a large animal model. A step by-step protocol for inter-positional placement of the TEV and real-time digital assessment of the TEV and native carotid arteries is described here. In vivo monitoring is made possible by the implantation of flow probes, catheters and ultrasonic crystals (capable of recording dynamic diameter changes of implanted TEVs and native carotid arteries) at the time of surgery. Once implanted, researchers can calculate arterial blood flow patterns, invasive blood pressure and artery diameter yielding parameters such as pulse wave velocity, augmentation index, pulse pressures and compliance. Data acquisition is accomplished using a single computer program for analysis throughout the duration of the experiment. Such invaluable data provides insight into TEV matrix remodeling, its resemblance to native/sham controls and overall TEV performance in vivo.