Comprehensive genotyping of the USA national maize inbred seed bank

Background

Genotyping by sequencing, a new low-cost, high-throughput sequencing technology was used to genotype 2,815 maize inbred accessions, preserved mostly at the National Plant Germplasm System in the USA. The collection includes inbred lines from breeding programs all over the world.

Results

The method produced 681,257 single-nucleotide polymorphism (SNP) markers distributed across the entire genome, with the ability to detect rare alleles at high confidence levels. More than half of the SNPs in the collection are rare. Although most rare alleles have been incorporated into public temperate breeding programs, only a modest amount of the available diversity is present in the commercial germplasm. Analysis of genetic distances shows population stratification, including a small number of large clusters centered on key lines. Nevertheless, an average fixation index of 0.06 indicates moderate differentiation between the three major maize subpopulations. Linkage disequilibrium (LD) decays very rapidly, but the extent of LD is highly dependent on the particular group of germplasm and region of the genome. The utility of these data for performing genome-wide association studies was tested with two simply inherited traits and one complex trait. We identified trait associations at SNPs very close to known candidate genes for kernel color, sweet corn, and flowering time; however, results suggest that more SNPs are needed to better explore the genetic architecture of complex traits.

Conclusions

The genotypic information described here allows this publicly available panel to be exploited by researchers facing the challenges of sustainable agriculture through better knowledge of the nature of genetic diversity.     

Mu opioid receptor: a gateway to drug addiction

Mu opioid receptors mediate positive reinforcement following direct (morphine) or indirect (alcohol, cannabinoids, nicotine) activation, and our understanding of mu receptor function is central to the development of addiction therapies. Recent data obtained in native neurons confirm that mu receptor signaling and regulation are strongly agonist-dependent. Current functional mapping reveals morphine-activated neurons in the extended amygdala and early genomic approaches have identified novel mu receptor-associated proteins. A classification of about 30 genes either promoting or counteracting the addictive properties of morphine is proposed from the analysis of knockout mice data. The targeting of effectors or regulatory proteins, beyond the mu receptor itself, might provide valuable strategies to treat addictive disorders.     

Divergent strategies for controlling the nuclear membrane satisfy geometric constraints during nuclear division

Eukaryotes segregate chromosomes in "open" or "closed" mitosis, depending on whether their nuclear envelopes (NEs) break down or remain intact. Here we show that the control of the nuclear surface area may determine the choice between these two modes. The dividing nucleus does not expand its surface in the fission yeast Schizosaccharomyces japonicus, confining the mitotic spindle and causing it to?buckle. The NE ruptures in anaphase, releasing the compressive stress and allowing chromosome segregation.?Blocking the NE expansion in the related species Schizosaccharomyces pombe that undergoes closed mitosis induces spindle buckling and collapse in the absence of an intrinsic NE rupture mechanism. We propose that scaling considerations could have shaped the evolution of eukaryotic mitosis by necessitating either nuclear surface expansion or the NE breakdown.     

Polythiophenes inhibit prion propagation by stabilizing prion protein (PrP) aggregates

Luminescent conjugated polymers (LCPs) interact with ordered protein aggregates and sensitively detect amyloids of many different proteins, suggesting that they may possess antiprion properties. Here, we show that a variety of anionic, cationic, and zwitterionic LCPs reduced the infectivity of prion-containing brain homogenates and of prion-infected cerebellar organotypic cultured slices and decreased the amount of scrapie isoform of PrP(C) (PrP(Sc)) oligomers that could be captured in an avidity assay. Paradoxically, treatment enhanced the resistance of PrP(Sc) to proteolysis, triggered the compaction, and enhanced the resistance to proteolysis of recombinant mouse PrP(23-231) fibers. These results suggest that LCPs act as antiprion agents by transitioning PrP aggregates into structures with reduced frangibility. Moreover, ELISA on cerebellar organotypic cultured slices and in vitro conversion assays with mouse PrP(23-231) indicated that poly(thiophene-3-acetic acid) may additionally interfere with the generation of PrP(Sc) by stabilizing the conformation of PrP(C) or of a transition intermediate. Therefore, LCPs represent a novel class of antiprion agents whose mode of action appears to rely on hyperstabilization, rather than destabilization, of PrP(Sc) deposits.     

Herpes Virus MicroRNA Expression and Significance in Serous Ovarian Cancer

Serous ovarian cancer (SEOC) is the deadliest gynecologic malignancy. MicroRNAs (miRNAs) are a class of small noncoding RNAs which regulate gene expression and protein translation. MiRNAs are also encoded by viruses with the intent of regulating their own genes and those of the infected cells. This is the first study assessing viral miRNAs in SEOC. MiRNAs sequencing data from 487 SEOC patients were downloaded from the TCGA website and analyzed through in-house sequencing pipeline. To cross-validate TCGA analysis, we measured the expression of miR-H25 by quantitative immunofluorescence in an additional cohort of 161 SEOC patients. Gene, miRNA expression, and cytotoxicity assay were performed on multiple ovarian cancer cell lines transfected with miR-H25 and miR-BART7. Outcome analysis was performed using multivariate Cox and Kaplan-Meier method. Viral miRNAs are more expressed in SEOC than in normal tissues. Moreover, Herpetic viral miRNAs (miR-BART7 from EBV and miR-H25 from HSV-2) are significant and predictive biomarkers of outcome in multivariate Cox analysis. MiR-BART7 correlates with resistance to first line chemotherapy and early death, whereas miR-H25 appears to impart a protective effect and long term survival. Integrated analysis of gene and viral miRNAs expression suggests that miR-BART7 induces directly cisplatin-resistance, while miR-H25 alters RNA processing and affects the expression of noxious human miRNAs such as miR-143. This is the first investigation linking viral miRNA expression to ovarian cancer outcome. Viral miRNAs can be useful to develop biomarkers for early diagnosis and as a potential therapeutic tool to reduce SEOC lethality.     

Full engagement of liganded maltose‐binding protein stabilizes a semi‐open ATP‐binding cassette dimer in the maltose transporter

MalFGK₂ is an ATP‐binding cassette (ABC) transporter that mediates the uptake of maltose/maltodextrins into Escherichia coli. A periplasmic maltose‐binding protein (MBP) delivers maltose to the transmembrane subunits (MalFG) and stimulates the ATPase activity of the cytoplasmic nucleotide‐binding subunits (MalK dimer). This MBP‐stimulated ATPase activity is independent of maltose for purified transporter in detergent micelles. However, when the transporter is reconstituted in membrane bilayers, only the liganded form of MBP efficiently stimulates its activity. To investigate the mechanism of maltose stimulation, electron paramagnetic resonance spectroscopy was used to study the interactions between the transporter and MBP in nanodiscs and in detergent. We found that full engagement of both lobes of maltose‐bound MBP unto MalFGK₂ is facilitated by nucleotides and stabilizes a semi‐open MalK dimer. Maltose‐bound MBP promotes the transition to the semi‐open state of MalK when the transporter is in the membrane, whereas such regulation does not require maltose in detergent. We suggest that stabilization of the semi‐open MalK₂ conformation by maltose‐bound MBP is key to the coupling of maltose transport to ATP hydrolysis in vivo, because it facilitates the progression of the MalK dimer from the open to the semi‐open conformation, from which it can proceed to hydrolyze ATP.     

pIMP-PH114 Carrying bla IMP-4 in a Klebsiella pneumoniae Strain is Closely Related to Other Multidrug-Resistant IncA/C2 Plasmids

The IncA/C plasmids are broad host-range vehicles which have been associated with wide dissemination of CMY-2 among Enterobacteriaceae of human and animal origins. Acquired metallo-β-lactamases (MBLs) such as the IMP-type enzymes are increasingly reported in multidrug-resistant Gram-negative bacteria worldwide, particularly in Enterobacteriaceae. We described the complete sequence of the first IMP-4-encoding IncA/C₂ plasmid, pIMP-PH114 (151,885 bp), from a sequence type 1 Klebsiella pneumoniae strain that was recovered from a patient who was hospitalized in the Philippines. pIMP-PH114 consists of a backbone from the IncA/C₂ plasmids, with the insertion of a novel Tn21-like class 1 integron composite structure (containing the cassette array bla _IMP-4-qacG-aacA4-catB3, followed by a class C β-lactamase bla _DHA-1 and the mercury resistance operon, merRTPCADE) and a sul2-floR encoding region. Phylogenetic analysis of the IncA/C repA sequences showed that pIMP-PH114 formed a subgroup with other IncA/C plasmids involved in the international spread of CMY-2, TEM-24 and NDM-1. Identical bla _IMP-4 arrays have been described among different Enterobacteriaceae and Acinetobacter spp. in China, Singapore and Australia but the genetic context is different. The broad host range of IncA/C plasmids may have facilitated dissemination of the bla _IMP-4 arrays among different diverse groups of bacteria.     

Plasmin potentiates synaptic N-methyl-D-aspartate receptor function in hippocampal neurons through activation of protease-activated receptor-1

Protease-activated receptor-1 (PAR1) is activated by a number of serine proteases, including plasmin. Both PAR1 and plasminogen, the precursor of plasmin, are expressed in the central nervous system. In this study we examined the effects of plasmin in astrocyte and neuronal cultures as well as in hippocampal slices. We find that plasmin evokes an increase in both phosphoinositide hydrolysis (EC(50) 64 nm) and Fura-2/AM fluorescence (195 +/- 6.7% above base line, EC(50) 65 nm) in cortical cultured murine astrocytes. Plasmin also activates extracellular signal-regulated kinase (ERK1/2) within cultured astrocytes. The plasmin-induced rise in intracellular Ca(2+) concentration ([Ca(2+)](i)) and the increase in phospho-ERK1/2 levels were diminished in PAR1(-/-) astrocytes and were blocked by 1 microm BMS-200261, a selective PAR1 antagonist. However, plasmin had no detectable effect on ERK1/2 or [Ca(2+)](i) signaling in primary cultured hippocampal neurons or in CA1 pyramidal cells in hippocampal slices. Plasmin (100-200 nm) application potentiated the N-methyl-D-aspartate (NMDA) receptor-dependent component of miniature excitatory postsynaptic currents recorded from CA1 pyramidal neurons but had no effect on alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate- or gamma-aminobutyric acid receptor-mediated synaptic currents. Plasmin also increased NMDA-induced whole cell receptor currents recorded from CA1 pyramidal cells (2.5 +/- 0.3-fold potentiation over control). This effect was blocked by BMS-200261 (1 microm; 1.02 +/- 0.09-fold potentiation over control). These data suggest that plasmin may serve as an endogenous PAR1 activator that can increase [Ca(2+)](i) in astrocytes and potentiate NMDA receptor synaptic currents in CA1 pyramidal neurons.     

Controlling Meiotic Recombinational Repair – Specifying the Roles of ZMMs,Sgs1 and Mus81/Mms4 in Crossover Formation

Crossovers (COs) play a critical role in ensuring proper alignment and segregation of homologous chromosomes during meiosis. How the cell balances recombination between CO vs. noncrossover (NCO) outcomes is not completely understood. Further lacking is what constrains the extent of DNA repair such that multiple events do not arise from a single double-strand break (DSB). Here, by interpreting signatures that result from recombination genome-wide, we find that synaptonemal complex proteins promote crossing over in distinct ways. Our results suggest that Zip3 (RNF212) promotes biased cutting of the double Holliday-junction (dHJ) intermediate whereas surprisingly Msh4 does not. Moreover, detailed examination of conversion tracts in sgs1 and mms4-md mutants reveal distinct aberrant recombination events involving multiple chromatid invasions. In sgs1 mutants, these multiple invasions are generally multichromatid involving 3–4 chromatids; in mms4-md mutants the multiple invasions preferentially resolve into one or two chromatids. Our analysis suggests that Mus81/Mms4 (Eme1), rather than just being a minor resolvase for COs is crucial for both COs and NCOs in preventing chromosome entanglements by removing 3′- flaps to promote second-end capture. Together our results force a reevaluation of how key recombination enzymes collaborate to specify the outcome of meiotic DNA repair.