Linkage QTL Mapping and Genome-Wide Association Study on Resistance in Chickpea to Pythium ultimum

The soilborne oomycete plant pathogen Pythium ultimum causes seed rot and pre-emergence damping-off of chickpea (Cicer arietinum L.). The pathogen has been controlled for several decades using the fungicide metalaxyl as seed treatment but has re-emerged as a severe problem with the detection of metalaxyl-resistant isolates of the pathogen from infested fields in the United States Pacific Northwest. The objective of this study was to identify genetic markers and candidate genes associated with resistance to P. ultimum in an interspecific recombinant inbred line population (CRIL-7) derived from a cross between C. reticulatum (PI 599072) x C. arietinum (FLIP 84-92C) and conduct genome-wide association studies (GWAS) for disease resistance using a chickpea diversity panel consisting of 184 accessions. CRIL-7 was examined using 1029 SNP markers spanning eight linkage groups. A major QTL, “qpsd4-1,” was detected on LG 4 that explained 41.8% of phenotypic variance, and a minor QTL, “qpsd8-1,” was detected on LG8 that explained 4.5% of phenotypic variance. Seven candidate genes were also detected using composite interval mapping including several genes previously associated with disease resistance in other crop species. A total of 302,902 single nucleotide polymorphic (SNP) markers were used to determine population structure and kinship of the diversity panel. Marker–trait associations were established by employing different combinations of principal components (PC) and kinships (K) in the FarmCPU model. Genome-wide association studies detected 11 significant SNPs and seven candidate genes associated with disease resistance. SNP Ca4_1765418, detected by GWAS on chromosome 4, was located within QTL qpsd4-1 that was revealed in the interspecific CRIL-7 population. The present study provides tools to enable MAS for resistance to P. ultimum and identified genomic domains and candidate genes involved in the resistance of chickpea to soilborne diseases

Structure of the Shroom-Rho Kinase Complex Reveals a Binding Interface with Monomeric Shroom That Regulates Cell Morphology and Stimulates Kinase Activity

Shroom-mediated remodeling of the actomyosin cytoskeleton is a critical driver of cellular shape and tissue morphology that underlies the development of many tissues including the neural tube, eye, intestines, and vasculature. Shroom uses a conserved SD2 domain to direct the subcellular localization of Rho-associated kinase (Rock), which in turn drives changes in the cytoskeleton and cellular morphology through its ability to phosphorylate and activate non-muscle myosin II. Here, we present the structure of the human Shroom-Rock binding module, revealing an unexpected stoichiometry for Shroom in which two Shroom SD2 domains bind independent surfaces on Rock. Mutation of interfacial residues impaired Shroom-Rock binding in vitro and resulted in altered remodeling of the cytoskeleton and loss of Shroom-mediated changes in cellular morphology. Additionally, we provide the first direct evidence that Shroom can function as a Rock activator. These data provide molecular insight into the Shroom-Rock interface and demonstrate that Shroom directly participates in regulating cytoskeletal dynamics, adding to its known role in Rock localization

Schiedea haakoaensis, a new facultatively autogamous species of Schiedea sect. Mononeura (Caryophyllaceae) from the Hawaiian Islands

In 2016 during a survey for potential fencing of the Ha‘akoa unit on windward Mauna Kea, Hawai‘i Island (Hawaiian Islands) a single plant of the genus Schiedea was discovered. No species of the genus had ever been known to occur in this area, and only three species of Schiedea were known previously from Hawai‘i Island. Two are vining species and the third is a coastal subshrub. The single plant obviously represented an interesting find, and because the plant was vegetative another visit was scheduled to collect a flowering specimen, but by then the plant had died. Soil taken from the site with seeds in the soil produced two plants, one of which flowered in cultivation in 2021. A study of this individual indicated it was a member of Schiedea sect. Mononeura, characterized by erect to ascending habit, quadrangular stems, seeds not persistent on the placenta and readily dispersing from the dehisced capsule, and flowers facultatively autogamous. With the discovery of this new species there are 35 species in this Hawaiian endemic genus

Structural basis for the RING catalyzed synthesis of K63 linked ubiquitin chains

This work was supported by grants from Cancer Research UK (C434/A13067), the Wellcome Trust (098391/Z/12/Z) and Biotechnology and Biological Sciences Research Council (BB/J016004/1).The RING E3 ligase catalysed formation of lysine 63 linked ubiquitin chains by the Ube2V2–Ubc13 E2 complex is required for many important biological processes. Here we report the structure of the RING domain dimer of rat RNF4 in complex with a human Ubc13~Ub conjugate and Ube2V2. The structure has captured Ube2V2 bound to the acceptor (priming) ubiquitin with Lys63 in a position that could lead to attack on the linkage between the donor (second) ubiquitin and Ubc13 that is held in the active “folded back” conformation by the RING domain of RNF4. The interfaces identified in the structure were verified by in vitro ubiquitination assays of site directed mutants. This represents the first view of the synthesis of Lys63 linked ubiquitin chains in which both substrate ubiquitin and ubiquitin-loaded E2 are juxtaposed to allow E3 ligase mediated catalysis.PostprintPeer reviewe

Measurement of Directional Wave Spectra Using Aircraft Laser Altimeters

A remote sensing method to measure directional oceanic surface waves by three laser altimeters on the NOAA LongEZ aircraft is investigated. To examine feasibility and sensitivity of the wavelet analysis method to various waves, aircraft motions, and aircraft flight directions relative to wave propagation directions, idealized surface waves are simulated from various idealized aircraft flights. In addition, the wavelet analysis method is also applied to two cases from field measurements, and the results are compared with traditional wave spectra from buoys. Since the wavelet analysis method relies on the “wave slopes” measured through phase differences between the time series of the laser distances between the aircraft and sea surface at spatially separated locations, the resolved directional wavenumber and wave propagation direction are not affected by aircraft motions if the resolved frequencies of the aircraft motion and the wave are not the same. However, the encounter wave frequency, which is directly resolved using the laser measurement from the moving aircraft, is affected by the Doppler shift due to aircraft motion relative to wave propagations. The wavelet analysis method could fail if the aircraft flies in the direction such that the aircraft speed along the wave propagation direction is the same as the wave phase speed (i.e., the aircraft flies along wave crests or troughs) or if two waves with different wavelengths and phase speed have the same encountered wavelength from the aircraft. In addition, the data noise due to laser measurement uncertainty or natural isotropic surface elevation perturbations can also affect the relative phase difference between the laser distance measurements, which in turn affects the accuracy of the resolved wavenumber and wave propagation direction. The smallest waves measured by the lasers depend on laser sampling rate and horizontal distances between the lasers (for the LongEZ this is 2 m). The resolved wave direction and wavenumber at the peak wave from the two field experiments compared well with on-site buoy observations. Overall, the study demonstrates that three spatially separated laser altimeters on moving platforms can be utilized to resolve two-dimensional wave spectra

Triosephosphate isomerase I170V alters catalytic site, enhances stability and induces pathology in a Drosophila model of TPI deficiency ☆

Triosephosphate isomerase (TPI) is a glycolytic enzyme which homodimerizes for full catalytic activity. Mutations of the TPI gene elicit a disease known as TPI Deficiency, a glycolytic enzymopathy noted for its unique severity of neurological symptoms. Evidence suggests that TPI Deficiency pathogenesis may be due to conformational changes of the protein, likely affecting dimerization and protein stability. In this report, we genetically and physically characterize a human disease-associated TPI mutation caused by an I170V substitution. Human TPI I170V elicits behavioral abnormalities in Drosophila. An examination of hTPI I170V enzyme kinetics revealed this substitution reduced catalytic turnover, while assessments of thermal stability demonstrated an increase in enzyme stability. The crystal structure of the homodimeric I170V mutant reveals changes in the geometry of critical residues within the catalytic pocket. Collectively these data reveal new observations of the structural and kinetic determinants of TPI Deficiency pathology, providing new insights into disease pathogenesis

Mutant Versions of the S. cerevisiae Transcription Elongation Factor Spt16 Define Regions of Spt16 That Functionally Interact with Histone H3

In eukaryotic cells, the highly conserved FACT (FAcilitates Chromatin Transcription) complex plays important roles in several chromatin-based processes including transcription initiation and elongation. During transcription elongation, the FACT complex interacts directly with nucleosomes to facilitate histone removal upon RNA polymerase II (Pol II) passage and assists in the reconstitution of nucleosomes following Pol II passage. Although the contribution of the FACT complex to the process of transcription elongation has been well established, the mechanisms that govern interactions between FACT and chromatin still remain to be fully elucidated. Using the budding yeast Saccharomyces cerevisiae as a model system, we provide evidence that the middle domain of the FACT subunit Spt16 – the Spt16-M domain – is involved in functional interactions with histone H3. Our results show that the Spt16-M domain plays a role in the prevention of cryptic intragenic transcription during transcription elongation and also suggest that the Spt16-M domain has a function in regulating dissociation of Spt16 from chromatin at the end of the transcription process. We also provide evidence for a role for the extreme carboxy terminus of Spt16 in functional interactions with histone H3. Taken together, our studies point to previously undescribed roles for the Spt16 M-domain and extreme carboxy terminus in regulating interactions between Spt16 and chromatin during the process of transcription elongation

FACT Prevents the Accumulation of Free Histones Evicted from Transcribed Chromatin and a Subsequent Cell Cycle Delay in G1

The FACT complex participates in chromatin assembly and disassembly during transcription elongation. The yeast mutants affected in the SPT16 gene, which encodes one of the FACT subunits, alter the expression of G1 cyclins and exhibit defects in the G1/S transition. Here we show that the dysfunction of chromatin reassembly factors, like FACT or Spt6, down-regulates the expression of the gene encoding the cyclin that modulates the G1 length (CLN3) in START by specifically triggering the repression of its promoter. The G1 delay undergone by spt16 mutants is not mediated by the DNA–damage checkpoint, although the mutation of RAD53, which is otherwise involved in histone degradation, enhances the cell-cycle defects of spt16-197. We reveal how FACT dysfunction triggers an accumulation of free histones evicted from transcribed chromatin. This accumulation is enhanced in a rad53 background and leads to a delay in G1. Consistently, we show that the overexpression of histones in wild-type cells down-regulates CLN3 in START and causes a delay in G1. Our work shows that chromatin reassembly factors are essential players in controlling the free histones potentially released from transcribed chromatin and describes a new cell cycle phenomenon that allows cells to respond to excess histones before starting DNA replication

What Was the Set of Ubiquitin and Ubiquitin-Like Conjugating Enzymes in the Eukaryote Common Ancestor?

Ubiquitin (Ub)-conjugating enzymes (E2) are key enzymes in ubiquitination or Ub-like modifications of proteins. We searched for all proteins belonging to the E2 enzyme super-family in seven species (Homo sapiens, Mus musculus, Drosophila melanogaster, Caenorhabditis elegans, Schizosaccharomyces pombe, Saccharomyces cerevisiae, and Arabidopsis thaliana) to identify families and to reconstruct each family’s phylogeny. Our phylogenetic analysis of 207 genes led us to define 17 E2 families, with 37 E2 genes, in the human genome. The subdivision of E2 into four classes did not correspond to the phylogenetic tree. The sequence signature HPN (histidine–proline–asparagine), followed by a tryptophan residue at 16 (up to 29) amino acids, was highly conserved. When present, the active cysteine was found 7 to 8 amino acids from the C-terminal end of HPN. The secondary structures were characterized by a canonical alpha/beta fold. Only family 10 deviated from the common organization because the proteins were devoid of enzymatic activity. Family 7 had an insertion between beta strands 1 and 2; families 3, 5 and 14 had an insertion between the active cysteine and the conserved tryptophan. The three-dimensional data of these proteins highlight a strong structural conservation of the core domain. Our analysis shows that the primitive eukaryote ancestor possessed a diversified set of E2 enzymes, thus emphasizing the importance of the Ub pathway. This comprehensive overview of E2 enzymes emphasizes the diversity and evolution of this superfamily and helps clarify the nomenclature and true orthologies. A better understanding of the functions of these enzymes is necessary to decipher several human diseases

Accurate Inference of Subtle Population Structure (and Other Genetic Discontinuities) Using Principal Coordinates

Accurate inference of genetic discontinuities between populations is an essential component of intraspecific biodiversity and evolution studies, as well as associative genetics. The most widely-used methods to infer population structure are model-based, Bayesian MCMC procedures that minimize Hardy-Weinberg and linkage disequilibrium within subpopulations. These methods are useful, but suffer from large computational requirements and a dependence on modeling assumptions that may not be met in real data sets. Here we describe the development of a new approach, PCO-MC, which couples principal coordinate analysis to a clustering procedure for the inference of population structure from multilocus genotype data.PCO-MC uses data from all principal coordinate axes simultaneously to calculate a multidimensional "density landscape", from which the number of subpopulations, and the membership within subpopulations, is determined using a valley-seeking algorithm. Using extensive simulations, we show that this approach outperforms a Bayesian MCMC procedure when many loci (e.g. 100) are sampled, but that the Bayesian procedure is marginally superior with few loci (e.g. 10). When presented with sufficient data, PCO-MC accurately delineated subpopulations with population F(st) values as low as 0.03 (G'(st)>0.2), whereas the limit of resolution of the Bayesian approach was F(st) = 0.05 (G'(st)>0.35).We draw a distinction between population structure inference for describing biodiversity as opposed to Type I error control in associative genetics. We suggest that discrete assignments, like those produced by PCO-MC, are appropriate for circumscribing units of biodiversity whereas expression of population structure as a continuous variable is more useful for case-control correction in structured association studies