Genetic variations in adiponectin levels and dietary patterns on metabolic health among children with normal weight versus obesity: the BCAMS study.

BACKGROUND/OBJECTIVES: Adiponectin represents an important link between adipose tissue dysfunction and cardiometabolic risk in obesity; however, there is a lack of data on the effects of adiponectin-related genetic variations and gene-diet interactions on metabolic disorders in children. We aimed to investigate possible interactions between adiponectin-related genetic variants and habitual dietary patterns on metabolic health among children with normal weight versus overweight/obesity, and whether these effects in childhood longitudinally contribute to metabolic risk at follow-up. SUBJECTS/METHODS: In total, 3,317 Chinese children aged 6-18 at baseline and 339 participants at 10-year follow-up from the Beijing Child and Adolescent Metabolic Syndrome study cohort were included. Baseline lifestyle factors, plasma adiponectin levels, and six adiponectin-related genetic variants resulting from GWAS in East Asians (loci in/near ADIPOQ, CDH13, WDR11FGF, CMIP, and PEPD) were assessed for their associations with the metabolic disorders. Being metabolically unhealthy was defined by exhibiting any metabolic syndrome component. RESULTS: Among the six loci, ADIPOQ rs6773957 (OR 1.26, 95% CI:1.07-1.47, P = 0.004) and adiponectin receptor CDH13 rs4783244 (0.82, 0.69-0.96, P = 0.017) were correlated with metabolic risks independent of lifestyle factors in normal-weight children, but the associations were less obvious in those with overweight/obesity. A significant interaction between rs6773957 and diet (Pinteraction = 0.004) for metabolic health was observed in normal-weight children. The adiponectin-decreasing allele of rs6773957 was associated with greater metabolic risks in individuals with unfavorable diet patterns (P < 0.001), but not in those with healthy patterns (P > 0.1). A similar interaction effect was observed using longitudinal data (Pinteraction = 0.029). CONCLUSIONS: These findings highlight a novel gene-diet interaction on the susceptibility to cardiometabolic disorders, which has a long-term impact from childhood onward, particularly in those with normal weight. Personalized dietary advice in these individuals may be recommended as an early possible therapeutic measure to improve metabolic health.

Porcine reproductive and respiratory syndrome virus 2 (PRRSV-2) genetic diversity and occurrence of wild type and vaccine-like strains in the United States swine industry.

Porcine reproductive and respiratory syndrome virus genotype 2 (PRRSV-2) genetic diversity in the U.S. was assessed using a database comprising 10 years' worth of sequence data obtained from swine production systems routine monitoring and outbreak investigations. A total of 26,831 ORF5 PRRSV-2 sequences from 34 production systems were included in this analysis. Within group mean genetic distance (i.e. mean proportion of nucleotide differences within ORF5) per year according to herd type was calculated for all PRRSV-2 sequences. The percent nucleotide difference between each sequence and the ORF5 sequences from four commercially available PRRSV-2 vaccines (Ingelvac PRRS MLV, Ingelvac PRRS ATP, Fostera PRRS, and Prevacent PRRS) within the same lineage over time was used to classify sequences in wild-type or vaccine-like. The mean ORF5 genetic distance fluctuated from 0.09 to 0.13, being generally smaller in years in which there was a relative higher frequency of dominant lineage. Vaccine-like sequences comprised about one fourth of sequences obtained through routine monitoring of PRRS. We found that lineage 5 sequences were mostly Ingelvac PRRS MLV-like. Lineage 8 sequences up to 2011 were 62.9% Ingelvac PRRS ATP-like while the remaining were wild-type viruses. From 2012 onwards, 51.9% of lineage 8 sequences were Ingelvac PRRS ATP-like, 45.0% were Fostera PRRS-like, and only 3.2% were wild-type. For lineage 1 sequences, 0.1% and 1.7% of the sequences were Prevacent PRRS-like in 2009-2018 and 2019, respectively. These results suggest that repeated introductions of vaccine-like viruses through use of modified live vaccines might decrease within-lineage viral diversity as vaccine-like strains become more prevalent. Overall, this compilation of private data from routine monitoring provides valuable information on PRRSV viral diversity.

Identification of Novel GCK and HNF4α Gene Variants in Japanese Pediatric Patients with Onset of Diabetes before 17 Years of Age.

BACKGROUND: Maturity-onset diabetes of the young (MODY) is commonly misdiagnosed as type 1 or type 2 diabetes. Common reasons for misdiagnosis are related to limitations in genetic testing. A precise molecular diagnosis is essential for the optimal treatment of patients and allows for early diagnosis of their asymptomatic family members. OBJECTIVE: The aim of this study was to identify rare monogenic variants of common MODY genes in Japanese pediatric patients. METHODS: We investigated 45 Japanese pediatric patients based on the following clinical criteria: development of diabetes before 17 years of age, a family history of diabetes, testing negative for glutamate decarboxylase-65 (GAD 65) antibodies and insulinoma-2-associated autoantibodies (IA-2A), no significant obesity, and evidence of endogenous insulin production. Genetic screening for MODY1 (HNF4α), MODY2 (GCK), MODY3 (HNF1α), and MODY5 (HNF1ß) was performed by direct sequencing followed by multiplex ligation amplification assays. RESULTS: We identified 22 missense variants (3 novel variants) in 27 patients (60.0%) in the GCK, HNF4α, and HNF1α genes. We also detected a whole exon deletion in the HNF1ß gene and an exon 5-6 aberration in the GCK gene, each in one proband (4.4%). There were a total of 29 variations (64.4%), giving a relative frequency of 53.3% (24/45) for GCK, 2.2% (1/45) for HNF4α, 6.7% (3/45) for HNF1α, and 2.2% (1/45) for HNF1ß genes. CONCLUSIONS: Clinicians should consider collecting and assessing detailed clinical information, especially regarding GCK gene variants, in young antibody-negative patients with diabetes. Correct molecular diagnosis of MODY better predicts the clinical course of diabetes and facilitates individualized management.

Assessment of morphological and molecular variability of some Solanum melongena L. cultivars and wild Solanum incanum L. in Saudi Arabia.

The genus Solanum exhibits a wide range of variability in morphology, flavor, and tolerance to biotic and abiotic stresses. Phenotypic and genetic variability using ISSR and RAPD markers of Solanum incanum distributed in Al-Baha province of the Kingdom of Saudi Arabia is assessed. Thirty samples are representing three different locations: Baljershy, Aqeeq, and Tohama, besides twenty-five samples representing five different commercial cultivars tested. Growth type, the number of leaves per plant, fruit size (phenotypic traits), crude protein, carbohydrates, digestive organic matter, and Mg, Ca, P were the principal contributors in the PCA. Molecular analysis showed that 114 ISSR and 80 RAPD alleles with a 100% polymorphism were recorded. The polymorphism information content (PIC) values ranged from 0.84 to 0.91 for ISSR and from 0.59 to 0.89 for RAPD data. Similarity values ranged from 0.16 to 1.00, with an average of 0.47 for ISSR and from 0.01 to 0.97, with an average of 0.36 for RAPD. It resulted in a positive and significant correlation between morphological, molecular, nutritional, and chemical analysis of fruits using Mantel analysis. UPGMA and PCA for morphological traits and molecular data discriminated commercial cultivars and wild relatives. Solanum incanum was more diverse than commercial varieties. This study revealed a wide genetic diversity among and within collected eggplant accessions and may use in breeding programs of eggplants. There is a need to increase the present eggplant collection to widen the genetic diversity of cultivated eggplant varieties in Saudi Arabia.

Association analysis and evaluation of genetic diversity in wheat genotypes using SSR markers.

A population of 105 wheat genotypes (including 94 hexaploid and 11 tetraploid genotypes) was used to determine genetic diversity. Samples were grown based on the randomized complete block design with three replications under salinity stress (120 mM NaCl (and control (10 mM NaCl (conditions. Morpho-physiological traits associated with tolerance of salinity at the seedling stage were recorded. The results of the analysis of variance showed that there were significant differences between genotypes in all studied traits, except K+/Na+ ratio. The amount of potassium content of leaves and roots in control was higher than salt stress conditions. Salinity significantly decreased all traits measured except Na+ concentration in root and shoot and leaf stomata conduction. A total of 12 SSR (simple sequence repeats) markers were assessed for the existence of polymorphism between genotypes. The highest Nei (Nei 1973) gene diversity was observed for gwm410 (0.72) and gpw2181 (0.71) markers, and PIC (polymorphic information content index) values ranged from 0.2 to 0.67. According to PIC, only six markers were informative during this study. These markers could be more efficient in displaying the genotypic differentiation of the near-wheat species as they showed the highest genetic diversity. Simple regression analysis showed that barc212 marker had the most significant relationship with root dry weight, leaf moisture and stomatal conductance (at 0.01 significant level). The gpw2181 marker showed a significant correlation with different traits under stress conditions. It was suggested that this marker could be used for marker-assisted selection to improve salt stress tolerance of wheat.

The influence of ApoE4 on the clinical outcomes and pathophysiology of degenerative cervical myelopathy.

Degenerative cervical myelopathy (DCM) is the most common cause of nontraumatic spinal cord injury in adults worldwide. Surgical decompression is generally effective in improving neurological outcomes and halting progression of myelopathic deterioration. However, a subset of patients experience suboptimal neurological outcomes. Given the emerging evidence that apolipoprotein E4 (ApoE4) allelic status influences neurodegenerative conditions, we examined whether the presence of the ApoE4 allele may account for the clinical heterogeneity of treatment outcomes in patients with DCM. Our results demonstrate that human ApoE4+ DCM patients have a significantly lower extent of improvement after decompression surgery. Functional analysis of our DCM mouse model in targeted-replacement mice expressing human ApoE4 revealed delayed gait recovery, forelimb grip strength, and hind limb mechanical sensitivity after decompression surgery, compared with their ApoE3 counterparts. This was accompanied by an exacerbated proinflammatory response resulting in higher concentrations of TNF-α, IL-6, CCL3, and CXCL9. At the site of injury, there was a significant decrease in gray matter area, an increase in the activation of microglia/macrophages, and increased astrogliosis after decompression surgery in the ApoE4 mice. Our study is the first to our knowledge to investigate the pathophysiological underpinnings of ApoE4 in DCM, which suggests a possible personalized medicine approach for the treatment of DCM in ApoE4 carriers.

Colombia's bioregions as a source of useful plants.

The aim of our study was to assess the importance of different Colombian bioregions in terms of the supply of useful plant species and the quality of the available distribution data. We assembled a dataset of georeferenced collection localities of all vascular plants of Colombia available from global and local online databases. We then assembled a list of species, subspecies and varieties of Colombia's useful plants and retrieved all point locality information associated with these taxa. We overlaid both datasets with a map of Colombia's bioregions to retrieve all species and useful species distribution records in each bioregion. To assess the reliability of our estimates of species numbers, we identified information gaps, in geographic and environmental space, by estimating their completeness and coverage. Our results confirmed that Colombia's third largest bioregion, the Andean moist forest followed by the Amazon, Pacific, Llanos and Caribbean moist forests contained the largest numbers of useful plant species. Medicinal use was the most common useful attribute across all bioregions, followed by Materials, Environmental uses, and Human Food. In all bioregions, except for the Andean páramo, the proportion of well-surveyed 10×10 km grid cells (with ≥ 25 observation records of useful plants) was below 50% of the total number of surveyed cells. Poor survey coverage was observed in the three dry bioregions: Caribbean deserts and xeric shrublands, and Llanos and Caribbean dry forests. This suggests that additional primary data is needed. We document knowledge gaps that will hinder the incorporation of useful plants into Colombia's stated plans for a bioeconomy and their sustainable management. In particular, future research should focus on the generation of additional primary data on the distribution of useful plants in the Amazon and Llanos (Orinoquia) regions where both survey completeness and coverage appeared to be less adequate compared with other regions.

Addiction-Associated Genetic Variants Implicate Brain Cell Type- and Region-Specific Cis-Regulatory Elements in Addiction Neurobiology.

Recent large genome-wide association studies have identified multiple confident risk loci linked to addiction-associated behavioral traits. Most genetic variants linked to addiction-associated traits lie in noncoding regions of the genome, likely disrupting cis-regulatory element (CRE) function. CREs tend to be highly cell type-specific and may contribute to the functional development of the neural circuits underlying addiction. Yet, a systematic approach for predicting the impact of risk variants on the CREs of specific cell populations is lacking. To dissect the cell types and brain regions underlying addiction-associated traits, we applied stratified linkage disequilibrium score regression to compare genome-wide association studies to genomic regions collected from human and mouse assays for open chromatin, which is associated with CRE activity. We found enrichment of addiction-associated variants in putative CREs marked by open chromatin in neuronal (NeuN+) nuclei collected from multiple prefrontal cortical areas and striatal regions known to play major roles in reward and addiction. To further dissect the cell type-specific basis of addiction-associated traits, we also identified enrichments in human orthologs of open chromatin regions of female and male mouse neuronal subtypes: cortical excitatory, D1, D2, and PV. Last, we developed machine learning models to predict mouse cell type-specific open chromatin, enabling us to further categorize human NeuN+ open chromatin regions into cortical excitatory or striatal D1 and D2 neurons and predict the functional impact of addiction-associated genetic variants. Our results suggest that different neuronal subtypes within the reward system play distinct roles in the variety of traits that contribute to addiction.SIGNIFICANCE STATEMENT We combine statistical genetic and machine learning techniques to find that the predisposition to for nicotine, alcohol, and cannabis use behaviors can be partially explained by genetic variants in conserved regulatory elements within specific brain regions and neuronal subtypes of the reward system. Our computational framework can flexibly integrate open chromatin data across species to screen for putative causal variants in a cell type- and tissue-specific manner for numerous complex traits.

Prospects for the natural distribution of crop wild-relatives with limited adaptability: The case of the wild pea Pisum fulvum.

Plant breeders and conservationist depend on knowledge about the genetic variation of their species of interest. Pisum fulvum, a wild relative of domesticated pea, has attracted attention as a genetic resource for crop improvement, yet little information about its diversity in the wild has been published hitherto. We sampled 15 populations of P. fulvum from Israeli natural habitats and conducted genotyping by sequencing to analyse their genetic diversity and adaptive state. We also attempted to evaluate the species past demography and the prospects of its future reaction to environmental changes. The results suggest that genetic diversity of P. fulvum is low to medium and is distributed between well diverged populations. Surprisingly, with 56 % in the total population the selfing rate was found to be significantly lower than expected from a species that is commonly assumed to be a predominant selfer. We found a strong genetic bottleneck during the last glacial period and only limited patterns of isolation by distance and environment, which explained 13 %-18 % of the genetic variation. Despite the weak signatures of genome-wide IBE, 1,354 markers were significantly correlated with environmental factors, 1,233 of which were located within known genes with a nonsynonymous to synonymous ratio of 0.382. Species distribution modelling depicted an ongoing fragmentation and decreased habitable area over the next 80 years under two different socio-economic pathways. Our results suggest that complex interactions of substantial drift and selection shaped the genome of P. fulvum. Climate changeis likely to cause further erosion of genetic diversity in P. fulvum. Systematic ex-situ conservation may be advisable to safeguard genetic variability for future utilization of this species.

Hyperlipidemia Influences the Accuracy of Glucometer-Measured Blood Glucose Concentrations in Genetically Diverse Mice.

BACKGROUND: Glucometers are widely used in animal research due to simplicity and ease of utilization, but their accuracy in blood glucose assessment for hyperlipidemic mice is unknown. METHODS: Here, we compared blood glucose levels measured by a glucometer with plasma glucose levels measured by a standard enzymatic assay for 325 genetically diverse F2 mice derived from LP and BALB/c (BALB) Apoe-/- mice. Non-fasting glucose levels were measured before initiation of a Western diet and after 11 weeks on the diet. RESULTS: On chow diet, lab-measured plasma glucose levels were 279.5 ± 42.6 mg/dl (mean ± SD), while blood glucose values measured by glucometer were 138.7 ± 16.6 mg/dl. The two measures had no correlation (R2 = 0.006, p = 0.167). On the Western diet, plasma glucose levels rose to 351.1 ± 121.6 mg/dl, while glucometer-measured blood glucose fell to 128.7 ± 27.9 mg/dl. The two measures showed a moderate correlation (R2 = 0.111, p = 3.1E-9). Lab-measured plasma glucose showed strong positive correlations with plasma triglyceride and non-high-density lipoprotein cholesterol levels, while glucometer-measured blood glucose showed an inverse correlation with non-high-density lipoprotein levels on the chow diet. CONCLUSIONS: Our results indicate that hyperlipidemia affects the accuracy of glucometers in measuring blood glucose levels of mice.

Characterization of the blood-brain barrier in genetically diverse laboratory mouse strains.

BACKGROUND: Genetic variation in a population has an influence on the manifestation of monogenic as well as multifactorial disorders, with the underlying genetic contribution dependent on several interacting variants. Common laboratory mouse strains used for modelling human disease lack the genetic variability of the human population. Therefore, outcomes of rodent studies show limited relevance to human disease. The functionality of brain vasculature is an important modifier of brain diseases. Importantly, the restrictive interface between blood and brain-the blood-brain barrier (BBB) serves as a major obstacle for the drug delivery into the central nervous system (CNS). Using genetically diverse mouse strains, we aimed to investigate the phenotypic and transcriptomic variation of the healthy BBB in different inbred mouse strains. METHODS: We investigated the heterogeneity of brain vasculature in recently wild-derived mouse strains (CAST/EiJ, WSB/EiJ, PWK/PhJ) and long-inbred mouse strains (129S1/SvImJ, A/J, C57BL/6J, DBA/2J, NOD/ShiLtJ) using different phenotypic arms. We used immunohistochemistry and confocal laser microscopy followed by quantitative image analysis to determine vascular density and pericyte coverage in two brain regions-cortex and hippocampus. Using a low molecular weight fluorescence tracer, sodium fluorescein and spectrophotometry analysis, we assessed BBB permeability in young and aged mice of selected strains. For further phenotypic characterization of endothelial cells in inbred mouse strains, we performed bulk RNA sequencing of sorted endothelial cells isolated from cortex and hippocampus. RESULTS: Cortical vessel density and pericyte coverage did not differ among the investigated strains, except in the cortex, where PWK/PhJ showed lower vessel density compared to NOD/ShiLtJ, and a higher pericyte coverage than DBA/2J. The vascular density in the hippocampus differed among analyzed strains but not the pericyte coverage. The staining patterns of endothelial arteriovenous zonation markers were similar in different strains. BBB permeability to a small fluorescent tracer, sodium fluorescein, was also similar in different strains, except in the hippocampus where the CAST/EiJ showed higher permeability than NOD/ShiLtJ. Transcriptomic analysis of endothelial cells revealed that sex of the animal was a major determinant of gene expression differences. In addition, the expression level of several genes implicated in endothelial function and BBB biology differed between wild-derived and long-inbred mouse strains. In aged mice of three investigated strains (DBA/2J, A/J, C57BL/6J) vascular density and pericyte coverage did not change-expect for DBA/2J, whereas vascular permeability to sodium fluorescein increased in all three strains. CONCLUSIONS: Our analysis shows that although there were no major differences in parenchymal vascular morphology and paracellular BBB permeability for small molecular weight tracer between investigated mouse strains or sexes, transcriptomic differences of brain endothelial cells point to variation in gene expression of the intact BBB. These baseline variances might be confounding factors in pathological conditions that may lead to a differential functional outcome dependent on the sex or genetic polymorphism.

Genomic Insights into Myasthenia Gravis Identify Distinct Immunological Mechanisms in Early and Late Onset Disease.

OBJECTIVE: The purpose of this study was to identify disease relevant genes and explore underlying immunological mechanisms that contribute to early and late onset forms of myasthenia gravis. METHODS: We used a novel genomic methodology to integrate genomewide association study (GWAS) findings in myasthenia gravis with cell-type specific information, such as gene expression patterns and promotor-enhancer interactions, in order to identify disease-relevant genes. Subsequently, we conducted additional genomic investigations, including an expression quantitative analysis of 313 healthy people to provide mechanistic insights. RESULTS: We identified several genes that were specifically linked to early onset myasthenia gravis including TNIP1, ORMDL3, GSDMB, and TRAF3. We showed that regulators of toll-like receptor 4 signaling were enriched among these early onset disease genes (fold enrichment = 3.85, p = 6.4 × 10-3 ). In contrast, T-cell regulators CD28 and CTLA4 were exclusively linked to late onset disease. We identified 2 causal genetic variants (rs231770 and rs231735; posterior probability = 0.98 and 0.91) near the CTLA4 gene. Subsequently, we demonstrated that these causal variants result in low expression of CTLA4 (rho = -0.66, p = 1.28 × 10-38 and rho = -0.52, p = 7.01 × 10-22 , for rs231735 and rs231770, respectively). INTERPRETATION: The disease-relevant genes identified in this study are a unique resource for many disciplines, including clinicians, scientists, and the pharmaceutical industry. The distinct immunological pathways linked to early and late onset myasthenia gravis carry important implications for drug repurposing opportunities and for future studies of drug development. ANN NEUROL 2021;90:455-463.

Evidence of cryptic diversity in freshwater Macrobrachium prawns from Indochinese riverine systems revealed by DNA barcode, species delimitation and phylogenetic approaches.

The diversity of Indochinese prawns in genus Macrobrachium is enormous due to the habitat diversification and broad tributary networks of two river basins: the Chao Phraya and the Mekong. Despite long-standing interest in SE-Asian decapod diversity, the subregional Macrobrachium fauna is still not yet comprehensively clarified in terms of taxonomic identification or genetic diversification. In this study, integrative taxonomic approaches including morphological examination, DNA barcoding, and molecular species delimitation were used to emphasize the broad scale systematics of Macrobrachium prawns in Indochina. Twenty-seven nominal species were successfully re-verified by traditional and molecular taxonomy. Barcode gap analysis supported broad overlapping of species boundaries. Taxonomic ambiguity of several deposited samples in the public database is related to inter- and intraspecific genetic divergence as indicated by BOLD discordance. Diagnostic nucleotide positions were found in six Macrobrachium species. Eighteen additional putative lineages are herein assigned using the consensus of species delimitation methods. Genetic divergence indicates the possible existence of cryptic species in four morphologically complex and wide-ranging species: M. lanchesteri, M. niphanae, M. sintangense, and some members of the M. pilimanus group. The geographical distribution of some species supports the connections and barriers attributed to paleo-historical events of SE-Asian rivers and land masses. Results of this study show explicitly the importance of freshwater ecosystems in Indochinese subregions, especially for the Mekong River Basin due to its high genetic diversity and species composition found throughout its tributaries.

Loss-of-function variants in Kv 11.1 cardiac channels as a biomarker for SUDEP.

OBJECTIVE: To compare the frequency and impact on the channel function of KCNH2 variants in SUDEP patients with epilepsy controls comprising patients older than 50 years, a group with low SUDEP risk, and establish loss-of-function KCNH2 variants as predictive biomarkers of SUDEP risk. METHODS: We searched for KCNH2 variants with a minor allele frequency of <5%. Functional analysis in Xenopus laevis oocytes was performed for all KCNH2 variants identified. RESULTS: KCNH2 variants were found in 11.1% (10/90) of SUDEP individuals compared to 6.0% (20/332) of epilepsy controls (p = 0.11). Loss-of-function KCNH2 variants, defined as causing >20% reduction in maximal amplitude, were observed in 8.9% (8/90) SUDEP patients compared to 3.3% (11/332) epilepsy controls suggesting about threefold enrichment (nominal p = 0.04). KCNH2 variants that did not change channel function occurred at a similar frequency in SUDEP (2.2%; 2/90) and epilepsy control (2.7%; 9/332) cohorts (p > 0.99). Rare KCNH2 variants (<1% allele frequency) associated with greater loss of function and an ~11-fold enrichment in the SUDEP cohort (nominal p = 0.03). In silico tools were unable to predict the impact of a variant on function highlighting the need for electrophysiological analysis. INTERPRETATION: These data show that loss-of-function KCNH2 variants are enriched in SUDEP patients when compared to an epilepsy population older than 50 years, suggesting that cardiac mechanisms contribute to SUDEP risk. We propose that genetic screening in combination with functional analysis can identify loss-of-function KCNH2 variants that could act as biomarkers of an individual's SUDEP risk.

Analysis of natural variation of the rice blast resistance gene Pike and identification of a novel allele Pikg.

Plant major resistance (R) genes are effective in detecting pathogen signal molecules and triggering robust defense responses. Investigating the natural variation in R genes will allow identification of the critical amino acid residues determining recognition specificity in R protein and the discovery of novel R alleles. The rice blast resistance gene Pike, comprising of two adjacent CC-NBS-LRR genes, namely, Pike-1 and Pike-2, confers broad-spectrum resistance to Magnaporthe oryzae. Here, we demonstrated that Pike-1 determined Pike-specific resistance through direct interaction with the pathogen signal molecule AvrPik. Analysis of natural variation in 79 Pike-1 variants in the Asian cultivated rice Oryza sativa and its wild relatives revealed that the CC and NBS regions, particularly the CC region of the Pike-1 protein were the most diversified. We also found that balancing selection had occurred in O. sativa and O. rufipogon to maintain the genetic diversity of the Pike-1 alleles. By analysis of amino acid sequences, we identified 40 Pike-1 variants in these rice germplasms. These variants were divided into three major groups that corresponded to their respective clades. A new Pike allele, designated Pikg, that differed from Pike by a single amino acid substitution (D229E) in the Pike-1 CC region of the Pike protein was identified from wild rice relatives. Pathogen assays of Pikg transgenic plants revealed a unique reaction pattern that was different from that of the previously identified Pike alleles, namely, Pik, Pikh, Pikm, Pikp, Piks and Pi1. These findings suggest that minor amino acid residues in Pike-1/Pikg-1 determine pathogen recognition specificity and plant resistance. As a new blast R gene derived from rice wild relatives, Pikg has potential applications in rice breeding.

Variation in the vulnerability of mice expressing human superoxide dismutase 1 to prion-like seeding: a study of the influence of primary amino acid sequence.

Misfolded forms of superoxide dismutase 1 (SOD1) with mutations associated with familial amyotrophic lateral sclerosis (fALS) exhibit prion characteristics, including the ability to act as seeds to accelerate motor neuron disease in mouse models. A key feature of infectious prion seeding is that the efficiency of transmission is governed by the primary sequence of prion protein (PrP). Isologous seeding, where the sequence of the PrP in the seed matches that of the host, is generally much more efficient than when there is a sequence mis-match. Here, we used paradigms in which mutant SOD1 seeding homogenates were injected intraspinally in newborn mice or into the sciatic nerve of adult mice, to assess the influence of SOD1 primary sequence on seeding efficiency. We observed a spectrum of seeding efficiencies depending upon both the SOD1 expressed by mice injected with seeds and the origin of the seed preparations. Mice expressing WT human SOD1 or the disease variant G37R were resistant to isologous seeding. Mice expressing G93A SOD1 were also largely resistant to isologous seeding, with limited success in one line of mice that express at low levels. By contrast, mice expressing human G85R-SOD1 were highly susceptible to isologous seeding but resistant to heterologous seeding by homogenates from paralyzed mice over-expressing mouse SOD1-G86R. In other seeding experiments with G85R SOD1:YFP mice, we observed that homogenates from paralyzed animals expressing the H46R or G37R variants of human SOD1 were less effective than seeds prepared from mice expressing the human G93A variant. These sequence mis-match effects were less pronounced when we used purified recombinant SOD1 that had been fibrilized in vitro as the seeding preparation. Collectively, our findings demonstrate diversity in the abilities of ALS variants of SOD1 to initiate or sustain prion-like propagation of misfolded conformations that produce motor neuron disease.

Intraspecific variation in immune gene expression and heritable symbiont density.

Host genetic variation plays an important role in the structure and function of heritable microbial communities. Recent studies have shown that insects use immune mechanisms to regulate heritable symbionts. Here we test the hypothesis that variation in symbiont density among hosts is linked to intraspecific differences in the immune response to harboring symbionts. We show that pea aphids (Acyrthosiphon pisum) harboring the bacterial endosymbiont Regiella insecticola (but not all other species of symbionts) downregulate expression of key immune genes. We then functionally link immune expression with symbiont density using RNAi. The pea aphid species complex is comprised of multiple reproductively-isolated host plant-adapted populations. These 'biotypes' have distinct patterns of symbiont infections: for example, aphids from the Trifolium biotype are strongly associated with Regiella. Using RNAseq, we compare patterns of gene expression in response to Regiella in aphid genotypes from multiple biotypes, and we show that Trifolium aphids experience no downregulation of immune gene expression while hosting Regiella and harbor symbionts at lower densities. Using F1 hybrids between two biotypes, we find that symbiont density and immune gene expression are both intermediate in hybrids. We propose that in this system, Regiella symbionts are suppressing aphid immune mechanisms to increase their density, but that some hosts have adapted to prevent immune suppression in order to control symbiont numbers. This work therefore suggests that antagonistic coevolution can play a role in host-microbe interactions even when symbionts are transmitted vertically and provide a clear benefit to their hosts. The specific immune mechanisms that we find are downregulated in the presence of Regiella have been previously shown to combat pathogens in aphids, and thus this work also highlights the immune system's complex dual role in interacting with both beneficial and harmful microbes.

Rescue of two trafficking-defective variants of the neuronal glycine transporter GlyT2 associated to hyperekplexia.

Hyperekplexia is a rare sensorimotor syndrome characterized by pathological startle reflex in response to unexpected trivial stimuli for which there is no specific treatment. Neonates suffer from hypertonia and are at high risk of sudden death due to apnea episodes. Mutations in the human SLC6A5 gene encoding the neuronal glycine transporter GlyT2 may disrupt the inhibitory glycinergic neurotransmission and cause a presynaptic form of the disease. The phenotype of missense mutations giving rise to protein misfolding but maintaining residual activity could be rescued by facilitating folding or intracellular trafficking. In this report, we characterized the trafficking properties of two mutants associated with hyperekplexia (A277T and Y707C, rat numbering). Transporter molecules were partially retained in the endoplasmic reticulum showing increased interaction with the endoplasmic reticulum chaperone calnexin. One transporter variant had export difficulties and increased ubiquitination levels, suggestive of enhanced endoplasmic reticulum-associated degradation. However, the two mutant transporters were amenable to correction by calnexin overexpression. Within the search for compounds capable of rescuing mutant phenotypes, we found that the arachidonic acid derivative N-arachidonoyl glycine can rescue the trafficking defects of the two variants in heterologous cells and rat brain cortical neurons. N-arachidonoyl glycine improves the endoplasmic reticulum output by reducing the interaction transporter/calnexin, increasing membrane expression and improving transport activity in a comparable way as the well-established chemical chaperone 4-phenyl-butyrate. This work identifies N-arachidonoyl glycine as a promising compound with potential for hyperekplexia therapy.