Phenotype to genotype using forward-genetic Mu-seq for identification and functional classification of maize mutants

In pursuing our long-term goals of identifying causal genes for mutant phenotypes in maize, we have developed a new, phenotype-to-genotype approach for transposon-based resources, and used this to identify candidate genes that co-segregate with visible kernel mutants. The strategy incorporates a redesigned Mu-seq protocol (sequence-based, transposon mapping) for high-throughput identification of individual plants carrying Mu insertions. Forward-genetic Mu-seq also involves a genetic pipeline for generating families that segregate for mutants of interest, and grid designs for concurrent analysis of genotypes in multiple families. Critically, this approach not only eliminates gene-specific PCR genotyping, but also profiles all Mu-insertions in hundreds of individuals simultaneously. Here, we employ this scalable approach to study 12 families that showed Mendelian segregation of visible seed mutants. These families were analyzed in parallel, and 7 showed clear co-segregation between the selected phenotype and a Mu insertion in a specific gene. Results were confirmed by PCR. Mutant genes that associated with kernel phenotypes include those encoding: a new allele of Whirly1 (a transcription factor with high affinity for organellar and single-stranded DNA), a predicted splicing factor with a KH domain, a small protein with unknown function, a putative mitochondrial transcription-termination factor, and three proteins with pentatricopeptide repeat domains (predicted mitochondrial). Identification of such associations allows mutants to be prioritized for subsequent research based on their functional annotations. Forward-genetic Mu-seq also allows a systematic dissection of mutant classes with similar phenotypes. In the present work, a high proportion of kernel phenotypes were associated with mutations affecting organellar gene transcription and processing, highlighting the importance and non-redundance of genes controlling these aspects of seed development

Identification of a Metabolic Disposal Route for the Oncometabolite S-(2-succino)cysteine in Bacillus subtilis

Cellular thiols such as cysteine spontaneously and readily react with the respiratory intermediate fumarate, resulting in the formation of stable S-(2-succino)-adducts. Fumarate-mediated succination of thiols increases in certain tumors and in response to glucotoxicity associated with diabetes. Therefore, S-(2-succino)-adducts such as S-(2-succino)cysteine (2SC) are considered oncometabolites and biomarkers for human disease. No disposal routes for S-(2-succino)-compounds have been reported prior to this study. Here, we show that Bacillus subtilis metabolizes 2SC to cysteine using a pathway encoded by the yxe operon. The first step is N-acetylation of 2SC followed by an oxygenation that we propose results in the release of oxaloacetate and N-acetylcysteine, which is deacetylated to give cysteine. Knockouts of the genes predicted to mediate each step in the pathway lose the ability to grow on 2SC as the sulfur source and accumulate the expected upstream metabolite(s). We further show that N-acetylation of 2SC relieves toxicity. This is the first demonstration of a metabolic disposal route for any S-(2-succino)-compound, paving the way toward the identification of corresponding pathways in other species

Divisions of labor in the thiamin biosynthetic pathway among organs of maize

The B vitamin thiamin is essential for central metabolism in all cellular organisms including plants. While plants synthesize thiamin de novo, organs vary widely in their capacities for thiamin synthesis. We use a transcriptomics approach to appraise the distribution of de novo synthesis and thiamin salvage pathways among organs of maize. We identify at least six developmental contexts in which metabolically active, non-photosynthetic organs exhibit low expression of one or both branches of the de novo thiamin biosynthetic pathway indicating a dependence on inter-cellular transport of thiamin and/or thiamin precursors. Neither the thiazole (THI4) nor pyrimidine (THIC) branches of the pathway are expressed in developing pollen implying a dependence on import of thiamin from surrounding floral and inflorescence organs. Consistent with that hypothesis, organs of the male inflorescence and flowers are shown to have high relative expression of the thiamin biosynthetic pathway and comparatively high thiamin contents. By contrast, divergent patterns of THIC and THI4 expression occur in the shoot apical meristem, embyro sac, embryo, endosperm, and root-tips suggesting that these sink organs acquire significant amounts of thiamin via salvage pathways. In the root and shoot meristems, expression of THIC in the absence of THI4 indicates a capacity for thiamin synthesis via salvage of thiazole, whereas the opposite pattern obtains in embryo and endosperm implying that seed storage organs are poised for pyrimidine salvage. Finally, stable isotope labeling experiments set an upper limit on the rate of de novo thiamin biosynthesis in maize leaf explants. Overall, the observed patterns of thiamin biosynthetic gene expression mirror the strategies for thiamin acquisition that have evolved in bacteria

Estrogen pathway polymorphisms in relation to primary open angle glaucoma: An analysis accounting for gender from the United States

Purpose Circulating estrogen levels are relevant in glaucoma phenotypic traits. We assessed the association between an estrogen metabolism single nucleotide polymorphism (SNP) panel in relation to primary open angle glaucoma (POAG), accounting for gender. Methods: We included 3,108 POAG cases and 3,430 controls of both genders from the Glaucoma Genes and Environment (GLAUGEN) study and the National Eye Institute Glaucoma Human Genetics Collaboration (NEIGHBOR) consortium genotyped on the Illumina 660W-Quad platform. We assessed the relation between the SNP panels representative of estrogen metabolism and POAG using pathway- and gene-based approaches with the Pathway Analysis by Randomization Incorporating Structure (PARIS) software. PARIS executes a permutation algorithm to assess statistical significance relative to the pathways and genes of comparable genetic architecture. These analyses were performed using the meta-analyzed results from the GLAUGEN and NEIGHBOR data sets. We evaluated POAG overall as well as two subtypes of POAG defined as intraocular pressure (IOP) ≥22 mmHg (high-pressure glaucoma [HPG]) or IOP 0.99). Among women, gene-based analyses revealed that the catechol-O-methyltransferase gene showed strong associations with HTG (permuted gene p≤0.001) and NPG (permuted gene p=0.01). Conclusions: The estrogen SNP pathway was associated with POAG among women

POPcorn: An Online Resource Providing Access to Distributed and Diverse Maize Project Data

The purpose of the online resource presented here, POPcorn (Project Portal for corn), is to enhance accessibility of maize genetic and genomic resources for plant biologists. Currently, many online locations are difficult to find, some are best searched independently, and individual project websites often degrade over time—sometimes disappearing entirely. The POPcorn site makes available (1) a centralized, web-accessible resource to search and browse descriptions of ongoing maize genomics projects, (2) a single, stand-alone tool that uses web Services and minimal data warehousing to search for sequence matches in online resources of diverse offsite projects, and (3) a set of tools that enables researchers to migrate their data to the long-term model organism database for maize genetic and genomic information: MaizeGDB. Examples demonstrating POPcorn's utility are provided herein

Search for the neutral Higgs bosons of the minimal supersymmetric standard model in pp collisions at root s=7 TeV with the ATLAS detector

A search for neutral Higgs bosons of the Minimal Supersymmetric Standard Model (MSSM) is reported. The analysis is based on a sample of proton-proton collisions at a centre-of-mass energy of 7TeV recorded with the ATLAS detector at the Large Hadron Collider. The data were recorded in 2011 and correspond to an integrated luminosity of 4.7 fb-1 to 4.8 fb-1. Higgs boson decays into oppositely-charged muon or τ lepton pairs are considered for final states requiring either the presence or absence of b-jets. No statistically significant excess over the expected background is observed and exclusion limits at the 95% confidence level are derived. The exclusion limits are for the production cross-section of a generic neutral Higgs boson, φ, as a function of the Higgs boson mass and for h/A/H production in the MSSM as a function of the parameters mA and tan β in the mhmax scenario for mA in the range of 90GeV to 500 GeV. Copyright CERN

CDKN2B-AS1 Genotype–Glaucoma Feature Correlations in Primary Open-Angle Glaucoma Patients From the United States

To assess the association between single nucleotide polymorphisms (SNPs) of the gene region containing cyclin dependent kinase inhibitor 2B antisense noncoding RNA (CDKN2B-AS1) and glaucoma features among primary open-angle glaucoma (POAG) patients

Hypothesis-independent pathway analysis implicates GABA and Acetyl-CoA metabolism in primary open-angle glaucoma and normal-pressure glaucoma

Primary open-angle glaucoma (POAG) is a leading cause of blindness worldwide. Using genome-wide association single-nucleotide polymorphism data from the Glaucoma Genes and Environment study and National Eye Institute Glaucoma Human Genetics Collaboration comprising 3,108 cases and 3,430 controls, we assessed biologic pathways as annotated in the KEGG database for association with risk of POAG. After correction for genic overlap among pathways, we found 4 pathways, butanoate metabolism (hsa00650), hematopoietic cell lineage (hsa04640), lysine degradation (hsa00310) and basal transcription factors (hsa03022) related to POAG with permuted p < 0.001. In addition, the human leukocyte antigen (HLA) gene family was significantly associated with POAG (p < 0.001). In the POAG subset with normal-pressure glaucoma (NPG), the butanoate metabolism pathway was also significantly associated (p < 0.001) as well as the MAPK and Hedgehog signaling pathways (hsa04010 and hsa04340), glycosaminoglycan biosynthesis-heparan sulfate pathway (hsa00534) and the phenylalanine, tyrosine and tryptophan biosynthesis pathway (hsa0400). The butanoate metabolism pathway overall, and specifically the aspects of the pathway that contribute to GABA and acetyl-CoA metabolism, was the only pathway significantly associated with both POAG and NPG. Collectively these results implicate GABA and acetyl-CoA metabolism in glaucoma pathogenesis, and suggest new potential therapeutic targets

Chromosome-level genome assembly of a regenerable maize inbred line A188.

BACKGROUND The maize inbred line A188 is an attractive model for elucidation of gene function and improvement due to its high embryogenic capacity and many contrasting traits to the first maize reference genome, B73, and other elite lines. The lack of a genome assembly of A188 limits its use as a model for functional studies. RESULTS Here, we present a chromosome-level genome assembly of A188 using long reads and optical maps. Comparison of A188 with B73 using both whole-genome alignments and read depths from sequencing reads identify approximately 1.1 Gb of syntenic sequences as well as extensive structural variation, including a 1.8-Mb duplication containing the Gametophyte factor1 locus for unilateral cross-incompatibility, and six inversions of 0.7 Mb or greater. Increased copy number of carotenoid cleavage dioxygenase 1 (ccd1) in A188 is associated with elevated expression during seed development. High ccd1 expression in seeds together with low expression of yellow endosperm 1 (y1) reduces carotenoid accumulation, accounting for the white seed phenotype of A188. Furthermore, transcriptome and epigenome analyses reveal enhanced expression of defense pathways and altered DNA methylation patterns of the embryonic callus. CONCLUSIONS The A188 genome assembly provides a high-resolution sequence for a complex genome species and a foundational resource for analyses of genome variation and gene function in maize. The genome, in comparison to B73, contains extensive intra-species structural variations and other genetic differences. Expression and network analyses identify discrete profiles for embryonic callus and other tissues

Common Variants at 9p21 and 8q22 Are Associated with Increased Susceptibility to Optic Nerve Degeneration in Glaucoma

Optic nerve degeneration caused by glaucoma is a leading cause of blindness worldwide. Patients affected by the normal-pressure form of glaucoma are more likely to harbor risk alleles for glaucoma-related optic nerve disease. We have performed a meta-analysis of two independent genome-wide association studies for primary open angle glaucoma (POAG) followed by a normal-pressure glaucoma (NPG, defined by intraocular pressure (IOP) less than 22 mmHg) subgroup analysis. The single-nucleotide polymorphisms that showed the most significant associations were tested for association with a second form of glaucoma, exfoliation-syndrome glaucoma. The overall meta-analysis of the GLAUGEN and NEIGHBOR dataset results (3,146 cases and 3,487 controls) identified significant associations between two loci and POAG: the CDKN2BAS region on 9p21 (rs2157719 [G], OR = 0.69 [95%CI 0.63–0.75], p = 1.86×10−18), and the SIX1/SIX6 region on chromosome 14q23 (rs10483727 [A], OR = 1.32 [95%CI 1.21–1.43], p = 3.87×10−11). In sub-group analysis two loci were significantly associated with NPG: 9p21 containing the CDKN2BAS gene (rs2157719 [G], OR = 0.58 [95% CI 0.50–0.67], p = 1.17×10−12) and a probable regulatory region on 8q22 (rs284489 [G], OR = 0.62 [95% CI 0.53–0.72], p = 8.88×10−10). Both NPG loci were also nominally associated with a second type of glaucoma, exfoliation syndrome glaucoma (rs2157719 [G], OR = 0.59 [95% CI 0.41–0.87], p = 0.004 and rs284489 [G], OR = 0.76 [95% CI 0.54–1.06], p = 0.021), suggesting that these loci might contribute more generally to optic nerve degeneration in glaucoma. Because both loci influence transforming growth factor beta (TGF-beta) signaling, we performed a genomic pathway analysis that showed an association between the TGF-beta pathway and NPG (permuted p = 0.009). These results suggest that neuro-protective therapies targeting TGF-beta signaling could be effective for multiple forms of glaucoma