Predictions of Arsenic in Domestic Well Water Sourced from Alluvial Aquifers of the Western Great Basin, USA.

Chronic exposure to high levels of arsenic in drinking water can have wide-ranging health effects and is a global health concern. The domestic well population of the western Great Basin (WGB) is at increased risk of exposure to arsenic due to the hydrologic, geologic, and climatic setting of the region. A logistic regression (LR) model was developed to predict the probability of elevated arsenic (≥5 µg/L) in alluvial aquifers and assess the potential geologic hazard level posed to domestic well populations. Alluvial aquifers are susceptible to arsenic contamination, which is a concern because they are the primary source of water for domestic well users of the WGB. The probability of elevated arsenic at a domestic well is strongly influenced by tectonic and geothermal variables, including the total Quaternary fault length in the hydrographic basin and the distance between the sampled well and a geothermal system. The model had an overall accuracy of 81%, sensitivity of 92%, and specificity of 55%. Results show a >50% probability of elevated arsenic in untreated well water for approximately 49 thousand (64%) alluvial-aquifer domestic well users in northern Nevada, northeastern California, and western Utah.

The road not taken: Evolution of tetrodotoxin resistance in the Sierra garter snake (Thamnophis couchii) by a path less travelled.

The repeated evolution of tetrodotoxin (TTX) resistance provides a model for testing hypotheses about the mechanisms of convergent evolution. This poison is broadly employed as a potent antipredator defence, blocking voltage-gated sodium channels (Nav ) in muscles and nerves, paralysing and sometimes killing predators. Resistance in taxa bearing this neurotoxin and a few predators appears to come from convergent replacements in specific Nav residues that interact with TTX. This stereotyped genetic response suggests molecular and phenotypic evolution may be constrained and predictable. Here, we investigate the extent of mechanistic convergence in garter snakes (Thamnophis) that prey on TTX-bearing newts (Taricha) by examining the physiological and genetic basis of TTX resistance in the Sierra garter snake (Th. couchii). We characterize variation in this predatory adaptation across populations at several biological scales: whole-animal TTX resistance; skeletal muscle resistance; functional genetic variation in three Nav encoding loci; and levels of gene expression for one of these loci. We found Th. couchii possess extensive geographical variation in resistance at the whole-animal and skeletal muscle levels. As in other Thamnophis, resistance at both levels is highly correlated, suggesting convergence across the biological levels linking organism to organ. However, Th. couchii shows no functional variation in Nav loci among populations or difference in candidate gene expression. Local variation in TTX resistance in Th. couchii cannot be explained by the same relationship between genotype and phenotype seen in other taxa. Thus, historical contingencies may lead different species of Thamnophis down alternative routes to local adaptation.

Drought tolerance of the grapevine, Vitis champinii cv. Ramsey, is associated with higher photosynthesis and greater transcriptomic responsiveness of abscisic acid biosynthesis and signaling.

BACKGROUND: Grapevine is an economically important crop for which yield and berry quality is strongly affected by climate change. Large variations in drought tolerance exist across Vitis species. Some of these species are used as rootstock to enhance abiotic and biotic stress tolerance. In this study, we investigated the physiological and transcriptomic responses to water deficit of four different genotypes that differ in drought tolerance: Ramsey (Vitis champinii), Riparia Gloire (Vitis riparia), Cabernet Sauvignon (Vitis vinifera), and SC2 (Vitis vinifera x Vitis girdiana). RESULTS: Ramsey was particularly more drought tolerant than the other three genotypes. Ramsey maintained a higher stomatal conductance and photosynthesis at equivalent levels of moderate water deficit. We identified specific and common transcriptomic responses shared among the four different Vitis species using RNA sequencing analysis. A weighted gene co-expression analysis identified a water deficit core gene set with the ABA biosynthesis and signaling genes, NCED3, RD29B and ABI1 as potential hub genes. The transcript abundance of many abscisic acid metabolism and signaling genes was strongly increased by water deficit along with genes associated with lipid metabolism, galactinol synthases and MIP family proteins. This response occurred at smaller water deficits in Ramsey and with higher transcript abundance than the other genotypes. A number of aquaporin genes displayed differential and unique responses to water deficit in Ramsey leaves. Genes involved in cysteine biosynthesis and metabolism were constitutively higher in the roots of Ramsey; thus, linking the gene expression of a known factor that influences ABA biosynthesis to this genotype's increased NCED3 transcript abundance. CONCLUSION: The drought tolerant Ramsey maintained higher photosynthesis at equivalent water deficit than the three other grapevine genotypes. Ramsey was more responsive to water deficit; its transcriptome responded at smaller water deficits, whereas the other genotypes did not respond until more severe water deficits were reached. There was a common core gene network responding to water deficit for all genotypes that included ABA metabolism and signaling. The gene clusters and sub-networks identified in this work represent interesting gene lists to explore and to better understand drought tolerance molecular mechanisms.

Particulate matter and emergency visits for asthma: a time-series study of their association in the presence and absence of wildfire smoke in Reno, Nevada, 2013-2018.

BACKGROUND: Health risks due to particulate matter (PM) from wildfires may differ from risk due to PM from other sources. In places frequently subjected to wildfire smoke, such as Reno, Nevada, it is critical to determine whether wildfire PM poses unique risks. Our goal was to quantify the difference in the association of adverse asthma events with PM on days when wildfire smoke was present versus days when wildfire smoke was not present. METHODS: We obtained counts of visits for asthma at emergency departments and urgent care centers from a large regional healthcare system in Reno for the years 2013-2018. We also obtained dates when wildfire smoke was present from the Washoe County Health District Air Quality Management Division. We then examined whether the presence of wildfire smoke modified the association of PM2.5, PM10-2.5, and PM10 with asthma visits using generalized additive models. We improved on previous studies by excluding wildfire-smoke days where the PM concentration exceeded the maximum PM concentration on other days, thus accounting for possible nonlinearity in the association between PM concentration and asthma visits. RESULTS: Air quality was affected by wildfire smoke on 188 days between 2013 and 2018. We found that the presence of wildfire smoke increased the association of a 5 µg/m3 increase in daily and three-day averages of PM2.5 with asthma visits by 6.1% (95% confidence interval (CI): 2.1-10.3%) and 6.8% (CI: 1.2-12.7%), respectively. Similarly, the presence of wildfire smoke increased the association of a 5 µg/m3 increase in daily and three-day averages of PM10 with asthma visits by 5.5% (CI: 2.5-8.6%) and 7.2% (CI: 2.6-12.0%), respectively. We did not observe any significant increases in association for PM10-2.5 or for seven-day averages of PM2.5 and PM10. CONCLUSIONS: Since we found significantly stronger associations of PM2.5 and PM10 with asthma visits when wildfire smoke was present, our results suggest that wildfire PM is more hazardous than non-wildfire PM for patients with asthma.

Single-nucleotide polymorphisms in a cohort of significantly obese women without cardiometabolic diseases.

BACKGROUND/OBJECTIVES: Obesity is an important risk factor for the development of diseases such as diabetes mellitus, hypertension, and dyslipidemia; however, a small number of individuals with long-standing obesity do not present with these cardiometabolic diseases. Such individuals are referred to as metabolically healthy obese (MHO) and potentially represent a subgroup of the general population with a protective genetic predisposition to obesity-related diseases. We hypothesized that individuals who were metabolically healthy, but significantly obese (BMI ≥ 35 kg/m2) would represent a highly homogenous subgroup, with which to investigate potential genetic associations to obesity. We further hypothesized that such a cohort may lend itself well to investigate potential genotypes that are protective with respect to the development of cardiometabolic disease. SUBJECTS/METHODS: In the present study, we implemented this novel selection strategy by screening 892 individuals diagnosed as Class 2 or Class 3 obese and identified 38 who presented no manifestations of cardiometabolic disease. We then assessed these subjects for single-nucleotide polymorphisms (SNPs) that associated with this phenotype. RESULTS: Our analysis identified 89 SNPs that reach statistical significance (p < 1 × 10-5), some of which are associated with genes of biological pathways that influences dietary behavior; others are associated with genes previously linked to obesity and cardiometabolic disease as well as neuroimmune disease. This study, to the best of our knowledge, represents the first genetic screening of a cardiometabolically healthy, but significantly obese population.

A comparison of heat-stress transcriptome changes between wild-type Arabidopsis pollen and a heat-sensitive mutant harboring a knockout of cyclic nucleotide-gated cation channel 16 (cngc16).

BACKGROUND: In flowering plants, the male gametophyte (pollen) is one of the most vulnerable cells to temperature stress. In Arabidopsis thaliana, a pollen-specific Cyclic Nucleotide-Gated cation Channel 16 (cngc16), is required for plant reproduction under temperature-stress conditions. Plants harboring a cncg16 knockout are nearly sterile under conditions of hot days and cold nights. To understand the underlying cause, RNA-Seq was used to compare the pollen transcriptomes of wild type (WT) and cngc16 under normal and heat stress (HS) conditions. RESULTS: Here we show that a heat-stress response (HSR) in WT pollen resulted in 2102 statistically significant transcriptome changes (≥ 2-fold changes with adjusted p-value ≤0.01), representing approximately 15% of 14,226 quantified transcripts. Of these changes, 89 corresponded to transcription factors, with 27 showing a preferential expression in pollen over seedling tissues. In contrast to WT, cngc16 pollen showed 1.9-fold more HS-dependent changes (3936 total, with 2776 differences between WT and cngc16). In a quantitative direct comparison between WT and cngc16 transcriptomes, the number of statistically significant differences increased from 21 pre-existing differences under normal conditions to 192 differences under HS. Of the 20 HS-dependent changes in WT that were most different in cngc16, half corresponded to genes encoding proteins predicted to impact cell wall features or membrane dynamics. CONCLUSIONS: Results here define an extensive HS-dependent reprogramming of approximately 15% of the WT pollen transcriptome, and identify at least 27 transcription factor changes that could provide unique contributions to a pollen HSR. The number of statistically significant transcriptome differences between WT and cngc16 increased by more than 9-fold under HS, with most of the largest magnitude changes having the potential to specifically impact cell walls or membrane dynamics, and thereby potentiate cngc16 pollen to be hypersensitive to HS. However, HS-hypersensitivity could also be caused by the extensive number of differences throughout the transcriptome having a cumulative effect on multiple cellular pathways required for tip growth and fertilization. Regardless, results here support a model in which a functional HS-dependent reprogramming of the pollen transcriptome requires a specific calcium-permeable Cyclic Nucleotide-Gated cation Channel, CNGC16.

Evaluation of four clinical laboratory parameters for the diagnosis of myalgic encephalomyelitis.

BACKGROUND: Myalgic encephalomyelitis (ME) is a complex and debilitating disease that often initially presents with flu-like symptoms, accompanied by incapacitating fatigue. Currently, there are no objective biomarkers or laboratory tests that can be used to unequivocally diagnosis ME; therefore, a diagnosis is made when a patient meets series of a costly and subjective inclusion and exclusion criteria. The purpose of the present study was to evaluate the utility of four clinical parameters in diagnosing ME. METHODS: In the present study, we utilized logistic regression and classification and regression tree analysis to conduct a retrospective investigation of four clinical laboratory in 140 ME cases and 140 healthy controls. RESULTS: Correlations between the covariates ranged between [- 0.26, 0.61]. The best model included the serum levels of the soluble form of CD14 (sCD14), serum levels of prostaglandin E2 (PGE2), and serum levels of interleukin 8, with coefficients 0.002, 0.249, and 0.005, respectively, and p-values of 3 × 10-7, 1 × 10-5, and 3 × 10-3, respectively. CONCLUSIONS: Our findings show that these parameters may help physicians in their diagnosis of ME and may additionally shed light on the pathophysiology of this disease.

Cardiac mitochondrial metabolism may contribute to differences in thermal tolerance of red- and white-blooded Antarctic notothenioid fishes.

Studies in temperate fishes provide evidence that cardiac mitochondrial function and the capacity to fuel cardiac work contribute to thermal tolerance. Here, we tested the hypothesis that decreased cardiac aerobic metabolic capacity contributes to the lower thermal tolerance of the haemoglobinless Antarctic icefish, Chaenocephalus aceratus, compared with that of the red-blooded Antarctic species, Notothenia coriiceps. Maximal activities of citrate synthase (CS) and lactate dehydrogenase (LDH), respiration rates of isolated mitochondria, adenylate levels and changes in mitochondrial protein expression were quantified from hearts of animals held at ambient temperature or exposed to their critical thermal maximum (CTmax). Compared with C. aceratus, activity of CS, ATP concentration and energy charge were higher in hearts of N. coriiceps at ambient temperature and CTmax While state 3 mitochondrial respiration rates were not impaired by exposure to CTmax in either species, state 4 rates, indicative of proton leakage, increased following exposure to CTmax in C. aceratus but not N. coriiceps The interactive effect of temperature and species resulted in an increase in antioxidants and aerobic metabolic enzymes in N. coriiceps but not in C. aceratus Together, our results support the hypothesis that the lower aerobic metabolic capacity of C. aceratus hearts contributes to its low thermal tolerance.

The common transcriptional subnetworks of the grape berry skin in the late stages of ripening.

BACKGROUND: Wine grapes are important economically in many countries around the world. Defining the optimum time for grape harvest is a major challenge to the grower and winemaker. Berry skins are an important source of flavor, color and other quality traits in the ripening stage. Senescent-like processes such as chloroplast disorganization and cell death characterize the late ripening stage. RESULTS: To better understand the molecular and physiological processes involved in the late stages of berry ripening, RNA-seq analysis of the skins of seven wine grape cultivars (Cabernet Franc, Cabernet Sauvignon, Merlot, Pinot Noir, Chardonnay, Sauvignon Blanc and Semillon) was performed. RNA-seq analysis identified approximately 2000 common differentially expressed genes for all seven cultivars across four different berry sugar levels (20 to 26 °Brix). Network analyses, both a posteriori (standard) and a priori (gene co-expression network analysis), were used to elucidate transcriptional subnetworks and hub genes associated with traits in the berry skins of the late stages of berry ripening. These independent approaches revealed genes involved in photosynthesis, catabolism, and nucleotide metabolism. The transcript abundance of most photosynthetic genes declined with increasing sugar levels in the berries. The transcript abundance of other processes increased such as nucleic acid metabolism, chromosome organization and lipid catabolism. Weighted gene co-expression network analysis (WGCNA) identified 64 gene modules that were organized into 12 subnetworks of three modules or more and six higher order gene subnetworks. Some gene subnetworks were highly correlated with sugar levels and some subnetworks were highly enriched in the chloroplast and nucleus. The petal R package was utilized independently to construct a true small-world and scale-free complex gene co-expression network model. A subnetwork of 216 genes with the highest connectivity was elucidated, consistent with the module results from WGCNA. Hub genes in these subnetworks were identified including numerous members of the core circadian clock, RNA splicing, proteolysis and chromosome organization. An integrated model was constructed linking light sensing with alternative splicing, chromosome remodeling and the circadian clock. CONCLUSIONS: A common set of differentially expressed genes and gene subnetworks from seven different cultivars were examined in the skin of the late stages of grapevine berry ripening. A densely connected gene subnetwork was elucidated involving a complex interaction of berry senescent processes (autophagy), catabolism, the circadian clock, RNA splicing, proteolysis and epigenetic regulation. Hypotheses were induced from these data sets involving sugar accumulation, light, autophagy, epigenetic regulation, and fruit development. This work provides a better understanding of berry development and the transcriptional processes involved in the late stages of ripening.

Sporobolus stapfianus: Insights into desiccation tolerance in the resurrection grasses from linking transcriptomics to metabolomics.

BACKGROUND: Understanding the response of resurrection angiosperms to dehydration and rehydration is critical for deciphering the mechanisms of how plants cope with the rigors of water loss from their vegetative tissues. We have focused our studies on the C4 resurrection grass, Sporobolus stapfianus Gandoger, as a member of a group of important forage grasses. METHODS: We have combined non-targeted metabolomics with transcriptomics, via a NimbleGen array platform, to develop an understanding of how gene expression and metabolite profiles can be linked to generate a more detailed mechanistic appreciation of the cellular response to both desiccation and rehydration. RESULTS: The rehydration transcriptome and metabolome are primarily geared towards the rapid return of photosynthesis, energy metabolism, protein turnover, and protein synthesis during the rehydration phase. However, there are some metabolites associated with ROS protection that remain elevated during rehydration, most notably the tocopherols. The analysis of the dehydration transcriptome reveals a strong concordance between transcript abundance and the associated metabolite abundance reported earlier, but only in responses that are directly related to cellular protection during dehydration: carbohydrate metabolism and redox homeostasis. The transcriptome response also provides strong support for the involvement of cellular protection processes as exemplified by the increases in the abundance of transcripts encoding late embryogenesis abundant (LEA) proteins, anti-oxidant enzymes, early light-induced proteins (ELIP) proteins, and cell-wall modification enzymes. There is little concordance between transcript and metabolite abundance for processes such as amino acid metabolism that do not appear to contribute directly to cellular protection, but are nonetheless important for the desiccation tolerant phenotype of S. stapfianus. CONCLUSIONS: The transcriptomes of both dehydration and rehydration offer insight into the complexity of the regulation of responses to these processes that involve complex signaling pathways and associated transcription factors. ABA appears to be important in the control of gene expression in both the latter stages of the dehydration and the early stages of rehydration. These findings add to the growing body of information detailing how plants tolerate and survive the severe cellular perturbations of dehydration, desiccation, and rehydration.

Transcriptomic network analyses of leaf dehydration responses identify highly connected ABA and ethylene signaling hubs in three grapevine species differing in drought tolerance.

BACKGROUND: Grapevine is a major food crop that is affected by global climate change. Consistent with field studies, dehydration assays of grapevine leaves can reveal valuable information of the plant's response at physiological, transcript, and protein levels. There are well-known differences in grapevine rootstocks responses to dehydration. We used time-series transcriptomic approaches combined with network analyses to elucidate and identify important physiological processes and network hubs that responded to dehydration in three different grapevine species differing in their drought tolerance. RESULTS: Transcriptomic analyses of the leaves of Cabernet Sauvignon, Riparia Gloire, and Ramsey were evaluated at different times during a 24-h controlled dehydration. Analysis of variance (ANOVA) revealed that approximately 11,000 transcripts changed significantly with respect to the genotype x treatment interaction term and approximately 6000 transcripts changed significantly according to the genotype x treatment x time interaction term indicating massive differential changes in gene expression over time. Standard analyses determined substantial effects on the transcript abundance of genes involved in the metabolism and signaling of two known plant stress hormones, abscisic acid (ABA) and ethylene. ABA and ethylene signaling maps were constructed and revealed specific changes in transcript abundance that were associated with the known drought tolerance of the genotypes including genes such as VviABI5, VviABF2, VviACS2, and VviWRKY22. Weighted-gene coexpression network analysis (WGCNA) confirmed these results. In particular, WGCNA identified 30 different modules, some of which had highly enriched gene ontology (GO) categories for photosynthesis, phenylpropanoid metabolism, ABA and ethylene signaling. The ABA signaling transcription factors, VviABI5 and VviABF2, were highly connected hubs in two modules, one being enriched in gaseous transport and the other in ethylene signaling. VviABI5 was distinctly correlated with an early response and high expression for the drought tolerant Ramsey and with little response from the drought sensitive Riparia Gloire. These ABA signaling transcription factors were highly connected to VviSnRK1 and other gene hubs associated with sugar, ethylene and ABA signaling. CONCLUSION: A leaf dehydration assay provided transcriptomic evidence for differential leaf responses to dehydration between genotypes differing in their drought tolerance. WGCNA proved to be a powerful network analysis approach; it identified 30 distinct modules (networks) with highly enriched GO categories and enabled the identification of gene hubs in these modules. Some of these genes were highly connected hubs in both the ABA and ethylene signaling pathways, supporting the hypothesis that there is substantial crosstalk between the two hormone pathways. This study identifies solid gene candidates for future investigations of drought tolerance in grapevine.

Abscisic acid transcriptomic signaling varies with grapevine organ.

BACKGROUND: Abscisic acid (ABA) regulates various developmental processes and stress responses over both short (i.e. hours or days) and longer (i.e. months or seasons) time frames. To elucidate the transcriptional regulation of early responses of grapevine (Vitis vinifera) responding to ABA, different organs of grape (berries, shoot tips, leaves, roots and cell cultures) were treated with 10 µM (S)-(+)-ABA for 2 h. NimbleGen whole genome microarrays of Vitis vinifera were used to determine the effects of ABA on organ-specific mRNA expression patterns. RESULTS: Transcriptomic analysis revealed 839 genes whose transcript abundances varied significantly in a specific organ in response to ABA treatment. No single gene exhibited the same changes in transcript abundance across all organs in response to ABA. The biochemical pathways affected by ABA were identified using the Cytoscape program with the BiNGO plug-in software. The results indicated that these 839 genes were involved in several biological processes such as flavonoid metabolism, response to reactive oxygen species, response to light, and response to temperature stimulus. ABA affected ion and water transporters, particularly in the root. The protein amino acid phosphorylation process was significantly overrepresented in shoot tips and roots treated with ABA. ABA affected mRNA abundance of genes (CYP707As, UGTs, and PP2Cs) associated with ABA degradation, conjugation, and the ABA signaling pathway. ABA also significantly affected the expression of several transcription factors (e.g. AP2/ERF, MYC/MYB, and bZIP/AREB). The greatest number of significantly differentially expressed genes was observed in the roots followed by cell cultures, leaves, berries, and shoot tips, respectively. Each organ had a unique set of gene responses to ABA. CONCLUSIONS: This study examined the short-term effects of ABA on different organs of grapevine. The responses of each organ were unique indicating that ABA signaling varies with the organ. Understanding the ABA responses in an organ-specific manner is crucial to fully understand hormone action and plant responses to water deficit.

Temporal-spatial interaction between reactive oxygen species and abscisic acid regulates rapid systemic acclimation in plants.

Being sessile organisms, plants evolved sophisticated acclimation mechanisms to cope with abiotic challenges in their environment. These are activated at the initial site of exposure to stress, as well as in systemic tissues that have not been subjected to stress (termed systemic acquired acclimation [SAA]). Although SAA is thought to play a key role in plant survival during stress, little is known about the signaling mechanisms underlying it. Here, we report that SAA in plants requires at least two different signals: an autopropagating wave of reactive oxygen species (ROS) that rapidly spreads from the initial site of exposure to the entire plant and a stress-specific signal that conveys abiotic stress specificity. We further demonstrate that SAA is stress specific and that a temporal-spatial interaction between ROS and abscisic acid regulates rapid SAA to heat stress in plants. In addition, we demonstrate that the rapid ROS signal is associated with the propagation of electric signals in Arabidopsis thaliana. Our findings unravel some of the basic signaling mechanisms underlying SAA in plants and reveal that signaling events and transcriptome and metabolome reprogramming of systemic tissues in response to abiotic stress occur at a much faster rate than previously envisioned.

Five omic technologies are concordant in differentiating the biochemical characteristics of the berries of five grapevine (Vitis vinifera L.) cultivars.

BACKGROUND: Grape cultivars and wines are distinguishable by their color, flavor and aroma profiles. Omic analyses (transcripts, proteins and metabolites) are powerful tools for assessing biochemical differences in biological systems. RESULTS: Berry skins of red- (Cabernet Sauvignon, Merlot, Pinot Noir) and white-skinned (Chardonnay, Semillon) wine grapes were harvested near optimum maturity (°Brix-to-titratable acidity ratio) from the same experimental vineyard. The cultivars were exposed to a mild, seasonal water-deficit treatment from fruit set until harvest in 2011. Identical sample aliquots were analyzed for transcripts by grapevine whole-genome oligonucleotide microarray and RNAseq technologies, proteins by nano-liquid chromatography-mass spectroscopy, and metabolites by gas chromatography-mass spectroscopy and liquid chromatography-mass spectroscopy. Principal components analysis of each of five Omic technologies showed similar results across cultivars in all Omic datasets. Comparison of the processed data of genes mapped in RNAseq and microarray data revealed a strong Pearson's correlation (0.80). The exclusion of probesets associated with genes with potential for cross-hybridization on the microarray improved the correlation to 0.93. The overall concordance of protein with transcript data was low with a Pearson's correlation of 0.27 and 0.24 for the RNAseq and microarray data, respectively. Integration of metabolite with protein and transcript data produced an expected model of phenylpropanoid biosynthesis, which distinguished red from white grapes, yet provided detail of individual cultivar differences. The mild water deficit treatment did not significantly alter the abundance of proteins or metabolites measured in the five cultivars, but did have a small effect on gene expression. CONCLUSIONS: The five Omic technologies were consistent in distinguishing cultivar variation. There was high concordance between transcriptomic technologies, but generally protein abundance did not correlate well with transcript abundance. The integration of multiple high-throughput Omic datasets revealed complex biochemical variation amongst five cultivars of an ancient and economically important crop species.

A roadmap for research on crassulacean acid metabolism (CAM) to enhance sustainable food and bioenergy production in a hotter, drier world.

Crassulacean acid metabolism (CAM) is a specialized mode of photosynthesis that features nocturnal CO2 uptake, facilitates increased water-use efficiency (WUE), and enables CAM plants to inhabit water-limited environments such as semi-arid deserts or seasonally dry forests. Human population growth and global climate change now present challenges for agricultural production systems to increase food, feed, forage, fiber, and fuel production. One approach to meet these challenges is to increase reliance on CAM crops, such as Agave and Opuntia, for biomass production on semi-arid, abandoned, marginal, or degraded agricultural lands. Major research efforts are now underway to assess the productivity of CAM crop species and to harness the WUE of CAM by engineering this pathway into existing food, feed, and bioenergy crops. An improved understanding of CAM has potential for high returns on research investment. To exploit the potential of CAM crops and CAM bioengineering, it will be necessary to elucidate the evolution, genomic features, and regulatory mechanisms of CAM. Field trials and predictive models will be required to assess the productivity of CAM crops, while new synthetic biology approaches need to be developed for CAM engineering. Infrastructure will be needed for CAM model systems, field trials, mutant collections, and data management.

Transcriptomic analysis of the late stages of grapevine (Vitis vinifera cv. Cabernet Sauvignon) berry ripening reveals significant induction of ethylene signaling and flavor pathways in the skin.

BACKGROUND: Grapevine berry, a nonclimacteric fruit, has three developmental stages; the last one is when berry color and sugar increase. Flavors derived from terpenoid and fatty acid metabolism develop at the very end of this ripening stage. The transcriptomic response of pulp and skin of Cabernet Sauvignon berries in the late stages of ripening between 22 and 37 °Brix was assessed using whole-genome micorarrays. RESULTS: The transcript abundance of approximately 18,000 genes changed with °Brix and tissue type. There were a large number of changes in many gene ontology (GO) categories involving metabolism, signaling and abiotic stress. GO categories reflecting tissue differences were overrepresented in photosynthesis, isoprenoid metabolism and pigment biosynthesis. Detailed analysis of the interaction of the skin and pulp with °Brix revealed that there were statistically significantly higher abundances of transcripts changing with °Brix in the skin that were involved in ethylene signaling, isoprenoid and fatty acid metabolism. Many transcripts were peaking around known optimal fruit stages for flavor production. The transcript abundance of approximately two-thirds of the AP2/ERF superfamily of transcription factors changed during these developmental stages. The transcript abundance of a unique clade of ERF6-type transcription factors had the largest changes in the skin and clustered with genes involved in ethylene, senescence, and fruit flavor production including ACC oxidase, terpene synthases, and lipoxygenases. The transcript abundance of important transcription factors involved in fruit ripening was also higher in the skin. CONCLUSIONS: A detailed analysis of the transcriptome dynamics during late stages of ripening of grapevine berries revealed that these berries went through massive transcriptional changes in gene ontology categories involving chemical signaling and metabolism in both the pulp and skin, particularly in the skin. Changes in the transcript abundance of genes involved in the ethylene signaling pathway of this nonclimacteric fruit were statistically significant in the late stages of ripening when the production of transcripts for important flavor and aroma compounds were at their highest. Ethylene transcription factors known to play a role in leaf senescence also appear to play a role in fruit senescence. Ethylene may play a bigger role than previously thought in this non-climacteric fruit.

EphB2 isolates a human marrow stromal cell subpopulation with enhanced ability to contribute to the resident intestinal cellular pool.

To identify human bone marrow stromal cell (BMSC) subsets with enhanced ability to engraft/contribute to the resident intestinal cellular pool, we transplanted clonally derived BMSCs into fetal sheep. Analysis at 75 d post-transplantation showed 2 of the 6 clones engrafting the intestine at 4- to 5-fold higher levels (5.03±0.089 and 5.04±0.15%, respectively) than the other clones (P<0.01), correlating with the percentage of donor-derived Musashi-1(+) (12.01-14.17 vs. 1.2-3.8%; P<0.01) or leucine-rich repeat-containing G-protein coupled receptor 5 (Lgr5)(+) cells within the intestinal stem cell (ISC) region. Phenotypic and transcriptome analysis determined that the clones with enhanced intestinal contribution expressed high levels of Ephrin type B receptor 2 (EphB2). Intestinal explants demonstrated proliferation of the engrafted cells and ability to generate crypt-like structures in vitro still expressing EphB2. Additional transplants based on BMSC EphB2 expression demonstrated that, at 7 d post-transplant, the EphB2(high) BMSCs engrafted in the ISC region at levels of 2.1 ± 0.2%, while control EphB2(low) BMSCs engrafted at 0.3 ± 0.1% (P<0.01). Therefore we identified a marker for isolating and culturing an expandable subpopulation of BMSCs with enhanced intestinal homing and contribution to the ISC region.

Screening Familial Risk for Hereditary Breast and Ovarian Cancer.

Importance: Most patients with pathogenic or likely pathogenic (P/LP) variants for breast cancer have not undergone genetic testing. Objective: To identify patients meeting family history criteria for genetic testing in the electronic health record (EHR). Design, Setting, and Participants: This study included both cross-sectional (observation date, February 1, 2024) and retrospective cohort (observation period, January 1, 2018, to February 1, 2024) analyses. Participants included patients aged 18 to 79 years enrolled in Renown Health, a large health system in Northern Nevada. Genotype was known for 38 003 patients enrolled in Healthy Nevada Project (HNP), a population genomics study. Exposure: An EHR indicating that a patient is positive for criteria according to the Seven-Question Family History Questionnaire (hereafter, FHS7 positive) assessing familial risk for hereditary breast and ovarian cancer (HBOC). Main Outcomes and Measures: The primary outcomes were the presence of P/LP variants in the ATM, BRCA1, BRCA2, CHEK2, or PALB2 genes (cross-sectional analysis) or a diagnosis of cancer (cohort analysis). Age-adjusted cancer incidence rates per 100 000 patients per year were calculated using the 2020 US population as the standard. Hazard ratios (HRs) for cancer attributable to FHS7-positive status were estimated using cause-specific hazard models. Results: Among 835 727 patients, 423 393 (50.7%) were female and 29 913 (3.6%) were FHS7 positive. Among those who were FHS7 positive, 24 535 (82.0%) had no evidence of prior genetic testing for HBOC in their EHR. Being FHS7 positive was associated with increased prevalence of P/LP variants in BRCA1/BRCA2 (odds ratio [OR], 3.34; 95% CI, 2.48-4.47), CHEK2 (OR, 1.62; 95% CI, 1.05-2.43), and PALB2 (OR, 2.84; 95% CI, 1.23-6.16) among HNP female individuals, and in BRCA1/BRCA2 (OR, 3.35; 95% CI, 1.93-5.56) among HNP male individuals. Being FHS7 positive was also associated with significantly increased risk of cancer among 131 622 non-HNP female individuals (HR, 1.44; 95% CI, 1.22-1.70) but not among 114 982 non-HNP male individuals (HR, 1.11; 95% CI, 0.87-1.42). Among 1527 HNP survey respondents, 352 of 383 EHR-FHS7 positive patients (91.9%) were survey-FHS7 positive, but only 352 of 883 survey-FHS7 positive patients (39.9%) were EHR-FHS7 positive. Of the 29 913 FHS7-positive patients, 19 764 (66.1%) were identified only after parsing free-text family history comments. Socioeconomic differences were also observed between EHR-FHS7-negative and EHR-FHS7-positive patients, suggesting disparities in recording family history. Conclusions and Relevance: In this cross-sectional study, EHR-derived FHS7 identified thousands of patients with familial risk for breast cancer, indicating a substantial gap in genetic testing. However, limitations in EHR family history data suggested that other identification methods, such as direct-to-patient questionnaires, are required to fully address this gap.

The human uterine smooth muscle S-nitrosoproteome fingerprint in pregnancy, labor, and preterm labor.

Molecular mechanisms involved in uterine quiescence during gestation and those responsible for induction of labor at term are incompletely known. More than 10% of babies born worldwide are premature and 1,000,000 die annually. Preterm labor results in preterm delivery in 50% of cases in the United States explaining 75% of fetal morbidity and mortality. There is no Food and Drug Administration-approved treatment to prevent preterm delivery. Nitric oxide-mediated relaxation of human uterine smooth muscle is independent of global elevation of cGMP following activation of soluble guanylyl cyclase. S-nitrosation is a likely mechanism to explain cGMP-independent relaxation to nitric oxide and may reveal S-nitrosated proteins as new therapeutic targets for the treatment of preterm labor. Employing S-nitrosoglutathione as an nitric oxide donor, we identified 110 proteins that are S-nitrosated in 1 or more states of human pregnancy. Using area under the curve of extracted ion chromatograms as well as normalized spectral counts to quantify relative expression levels for 62 of these proteins, we show that 26 proteins demonstrate statistically significant S-nitrosation differences in myometrium from spontaneously laboring preterm patients compared with nonlaboring patients. We identified proteins that were up-S-nitrosated as well as proteins that were down-S-nitrosated in preterm laboring tissues. Identification and relative quantification of the S-nitrosoproteome provide a fingerprint of proteins that can form the basis of hypothesis-directed efforts to understand the regulation of uterine contraction-relaxation and the development of new treatment for preterm labor.

ERCP and laparoscopic cholecystectomy in a combined (one-step) procedure: a random comparison to the standard (two-step) procedure.

BACKGROUND: Current treatment of complicated calculous biliary disease typically involves a two-step procedure consisting of preoperative endoscopic retrograde cholangiopancreatography (ERCP) followed by laparoscopic cholecystectomy. Alternatively, laparoscopic cholecystectomy with intraoperative cholangiogram (IOC) and intraoperative common bile duct exploration or ERCP at a later date may be performed. This study compared the benefits of the traditional two-step procedure to the novel one-step procedure for the management of calculous biliary disease. METHODS: A retrospective review of 20 patients was conducted comparing one-step to two-step procedures for the management of choledocholithiasis. We define the one-step procedure to be a laparoscopic cholecystectomy with IOC to confirm the presence or absence of stones. Intraoperative ERCP with stone extraction was conducted if necessary as part of the one-step procedure. RESULTS: A statistically significant difference existed between hospital charges for one-step ($58,145.30, SD $17,963.09) and two-step ($78,895.53, SD $21,954.78) procedures (p = 0.033). Other parameters (length of stay, preoperative days) trended toward significance; however, statistical significance was not achieved. CONCLUSIONS: There appears to be a significant cost reduction with implementation of the one-step treatment of calculous biliary disease. Further research with a larger study population is necessary to determine the additional benefits of this procedure and to help augment the surgical endoscopists' armamentarium.