Enemy at the Gates: Interaction-Specific Stomatal Responses to Pathogenic Challenge

Stomata regulate gas exchange and their closure in response to pathogens may, in some cases, contribute to resistance. However, in the cereal mildew and rust systems, stomatal closure follows establishment of compatible infections. In incompatible systems, expression of major (R) gene controlled hypersensitive responses (HR), causes drastic, permanent stomatal dysfunction: stomata become locked open following powdery mildew attack and locked shut following rust attack. Thus, stomatal locking can be a hitherto unsuspected negative consequence of R gene resistance that carries a physiological cost affecting plant performance.Key Words: stomata, rust, mildew, hypersensitive response, stomatal lock-up     

The high-resolution NMR structure of the R21A Spc-SH3:P41 complex: Understanding the determinants of binding affinity by comparison with Abl-SH3

Background  

SH3 domains are small protein modules of 60–85 amino acids that bind to short proline-rich sequences with moderate-to-low affinity and specificity. Interactions with SH3 domains play a crucial role in regulation of many cellular processes (some are related to cancer and AIDS) and have thus been interesting targets in drug design. The decapeptide APSYSPPPPP (p41) binds with relatively high affinity to the SH3 domain of the Abl tyrosine kinase (Abl-SH3), while it has a 100 times lower affinity for the α-spectrin SH3 domain (Spc-SH3).     

A mutation in the HLA-B*2705-restricted NP383-391 epitope affects the human influenza A virus-specific cytotoxic T-lymphocyte response in vitro

Viruses can exploit a variety of strategies to evade immune surveillance by cytotoxic T lymphocytes (CTL), including the acquisition of mutations in or adjacent to CTL epitopes. Recently, an amino acid substitution (R384G) in an HLA-B^*2705-restricted CTL epitope in the influenza A virus nucleoprotein (nucleoprotein containing residues 383 to 391 [NP_383-391]; SRYWAIRTR, where R is the residue that was mutated) was associated with escape from CTL-mediated immunity. The effect of this mutation on the in vitro influenza A virus-specific CTL response was studied. To this end, two influenza A viruses, one with and one without the NP_383-391 epitope, were constructed by reverse genetics and designated influenza viruses A/NL/94-384R and A/NL/94-384G, respectively. The absence of the HLA-B^*2705-restricted CTL epitope in influenza virus A/NL/94-384G was confirmed by using ⁵¹Cr release assays with a T-cell clone specific for the NP_383-391 epitope. In addition, peripheral blood mononuclear cells (PBMC) stimulated with influenza virus A/NL/94-384G failed to recognize HLA-B^*2705-positive target cells pulsed with the original NP_383-391 peptide. The proportion of virus-specific CD8⁺ gamma interferon (IFN-γ)-positive T cells in in vitro-stimulated PBMC was determined by intracellular IFN-γ staining after restimulation with virus-infected autologous B-lymphoblastoid cell lines and C1R cell lines expressing only HLA-B^*2705. The proportion of virus-specific CD8⁺ T cells was lower in PBMC stimulated in vitro with influenza virus A/NL/94-384G obtained from several HLA-B^*2705-positive donors than in PBMC stimulated with influenza virus A/NL/94-384R. This finding indicated that amino acid variations in CTL epitopes can affect the virus-specific CTL response and that the NP_383-391 epitope is the most important HLA-B^*2705-restricted epitope in the nucleoprotein of influenza A viruses.     

Species delimitation of the liquorice tribe (Leguminosae: Glycyrrhizeae) based on phylogenomic and machine learning analyses

The liquorice tribe Glycyrrhizeae is a leguminous herbaceous group of plants comprised of the genera Glycyrrhiza and Glycyrrhizopsis. Some Glycyrrhiza taxa contain glycyrrhizin, a pharmacologically significant sweet substance that also has applications in crafting industrial materials. Here, we utilized an expanded taxon sampling of Glycyrrhizeae to reconstruct the phylogenetic relationships in the tribe based on genome skimming data, including whole chloroplast genomes, nuclear ribosomal DNA, and low-copy nuclear DNA. We also launched machine learning analysis (MLA) for one species pair with controversial taxonomic boundary. The integrated results indicated Glycyrrhizopsis should be split from Glycyrrhiza, while the former genus Meristotropis should be treated as part of Glycyrrhiza. Glycyrrhizopsis includes two species, Glycyrrhizopsis asymmetrica and Glycyrrhizopsis flavescens, and we recognize 13 species in Glycyrrhiza: Glycyrrhiza acanthocarpa, Glycyrrhiza astragalina, Glycyrrhiza bucharica, Glycyrrhiza echinata, Glycyrrhiza foetida, Glycyrrhiza glabra, Glycyrrhiza gontscharovii, Glycyrrhiza lepidota, Glycyrrhiza macedonica, Glycyrrhiza pallidiflora, Glycyrrhiza squamulosa, Glycyrrhiza triphylla, and Glycyrrhiza yunnanensis. We propose a broader G. glabra that includes former Glycyrrhiza aspera, G. glabra s.s., Glycyrrhiza inflata, and Glycyrrhiza uralensis, and represents the glycyrrhizin-contained medicinal group. Our ancestral state inferences show the ancestor of Glycyrrhiza lacked glycyrrhizin, and the presence of glycyrrhizin evolved twice within Glycyrrhiza during the last one million years. Our integrative phylogenomics-MLA study not only provides new insights into long-standing taxonomic controversies of Glycyrrhizeae, but also represents a useful approach for future taxonomic studies on other plant taxa.     

Rearrangements of integral membrane components during in vitro aging of sheep erythrocyte membranes

In vitro aged sheep erythrocytes and sheep erythrocyte ghosts spontaneously release vesicles that consist of long protrusions affixed to flattened headlike structures. The intramembranous particles seen on the protoplasmic face of freeze fracture electron micrographs of vesicle protrusions are arranged in paired particle rows. On the equivalent fracture face of headlike structures, the particle density is low; if particles are present, they are clustered along the rim of the flattened headlike structure and at the junction with the protrusion. The released vesicles are depleted of the intramembranous particles seen on the exoplasmic face of ghost but retain almost exclusively particles of the protoplasmic face. Correspondingly, the exoplasmic face of ghosts that have released vesicles reveals a 28 percent higher density of intramembranous particles than that of fresh ghosts. Purified vesicles are depleted of spectrin but retain integral membrane proteins, with one of an apparent mol wt of 160,000 accounting for nearly 50 percent of the total protein (Lutz, H.U.,R. Barber, and R.F. McGuire. 1976. J. Biol. Chem. 251:3500-3510). When vesicles are modified with the cleavable cross-linking reagent [(35)S]dithiobis (succinimidyl propionate)at 0 degrees C, the 160,000 mol wt protein is rapidly converted to disulfide-linked dimers and higher oligomers. Exposure of intact ghosts to the reagent in the same way fails to yield equivalent polymers. A comparison of the morphological and biochemical aspects of ghosts and vesicles suggest that a marked rearrangement of membrane proteins accompanies the supramolecular redistribution of intramembranous particles during spontaneous vesiculation. The results also suggest that the paired particles of the protoplasmic face of vesicle protrusions are arranged in paired helices and contain the 160,000 mol wt protein as dimers.     

The Role of Competence in the Cranio-Caudal Segregation of the Central Nervous System

Left to right thirds of Triturus presumptive prosencephalon show identical developmental potencies after implantation in a neutral Ambystoma environment. Such equipotential grafts were excised from stages between late-gastrula and mid-neurula and implanted into the neural plate of an Ambystoma host at different cranio-caudal levels. Their regional differentiation was independent of the age of the host, but dependent upon the age of the donor material; the older the latter the smaller the portion of the graft which was transformed into more posterior neural structures. Full transformation occurred in stage 11/12 grafts, while pure prosencephalic differentiation took place in stage 16 grafts, demonstrating that the period of competence of the neurectoderm for transformation extends from stage 11/12 up to stage 16. Irrespective of the level of implantation all grafts older than stage 11/12 and younger than stage 16 showed an uninterrupted cranially-oriented regional differentiation. The medio-lateral extension of the transformation process is primarily determined by the temporal loss of competence of the implanted neurectoderm. A comparison of grafts implanted at different cranio-caudal levels showed that transformation is more pronounced the more caudal the level of implantation, so that another factor(s) than competence must also play a role in the regional segregation of the CNS.     

Stoichiometry and compartmentation of NADH metabolism in Saccharomyces cerevisiae

In Saccharomyces cerevisiae, reduction of NAD(+) to NADH occurs in dissimilatory as well as in assimilatory reactions. This review discusses mechanisms for reoxidation of NADH in this yeast, with special emphasis on the metabolic compartmentation that occurs as a consequence of the impermeability of the mitochondrial inner membrane for NADH and NAD(+). At least five mechanisms of NADH reoxidation exist in S. cerevisiae. These are: (1) alcoholic fermentation; (2) glycerol production; (3) respiration of cytosolic NADH via external mitochondrial NADH dehydrogenases; (4) respiration of cytosolic NADH via the glycerol-3-phosphate shuttle; and (5) oxidation of intramitochondrial NADH via a mitochondrial 'internal' NADH dehydrogenase. Furthermore, in vivo evidence indicates that NADH redox equivalents can be shuttled across the mitochondrial inner membrane by an ethanol-acetaldehyde shuttle. Several other redox-shuttle mechanisms might occur in S. cerevisiae, including a malate-oxaloacetate shuttle, a malate-aspartate shuttle and a malate-pyruvate shuttle. Although key enzymes and transporters for these shuttles are present, there is as yet no consistent evidence for their in vivo activity. Activity of several other shuttles, including the malate-citrate and fatty acid shuttles, can be ruled out based on the absence of key enzymes or transporters. Quantitative physiological analysis of defined mutants has been important in identifying several parallel pathways for reoxidation of cytosolic and intramitochondrial NADH. The major challenge that lies ahead is to elucidate the physiological function of parallel pathways for NADH oxidation in wild-type cells, both under steady-state and transient-state conditions. This requires the development of techniques for accurate measurement of intracellular metabolite concentrations in separate metabolic compartments.     

The Association between Mycobacterium Tuberculosis Genotype and Drug Resistance in Peru

BackgroundThe comparison of Mycobacterium tuberculosis bacterial genotypes with phenotypic, demographic, geospatial and clinical data improves our understanding of how strain lineage influences the development of drug-resistance and the spread of tuberculosis.MethodsTo investigate the association of Mycobacterium tuberculosis bacterial genotype with drug-resistance. Drug susceptibility testing together with genotyping using both 15-loci MIRU-typing and spoligotyping, was performed on 2,139 culture positive isolates, each from a different patient in Lima, Peru. Demographic, geospatial and socio-economic data were collected using questionnaires, global positioning equipment and the latest national census.ResultsThe Latin American Mediterranean (LAM) clade (OR 2.4, p<0.001) was significantly associated with drug-resistance and alone accounted for more than half of all drug resistance in the region. Previously treated patients, prisoners and genetically clustered cases were also significantly associated with drug-resistance (OR''s 2.5, 2.4 and 1.8, p<0.001, p<0.05, p<0.001 respectively).ConclusionsTuberculosis disease caused by the LAM clade was more likely to be drug resistant independent of important clinical, genetic and socio-economic confounding factors. Explanations for this include; the preferential co-evolution of LAM strains in a Latin American population, a LAM strain bacterial genetic background that favors drug-resistance or the "founder effect" from pre-existing LAM strains disproportionately exposed to drugs.     

Spatial phylogenetics of the native woody plant species in Hainan,China

To better identify biodiversity hotspots for conservation on Hainan Island, a tropical island in southern China, we assessed spatial variation in phylogenetic diversity and species richness using 18,976 georeferenced specimen records and a newly reconstructed molecular phylogeny of 957 native woody plants. Within this framework, we delineated bioregions based on vegetation composition and mapped areas of neoendemism and paleoendemism to identify areas of priority for conservation. Our results reveal that the southwest of Hainan is the most important hot spot for endemism and plant diversity followed by the southeast area. The distribution of endemic species showed a scattered, rather than clustered, pattern on the island. Based on phylogenetic range‐weighted turnover metrics, we delineated three major vegetational zones in Hainan. These largely correspond to natural secondary growth and managed forests (e.g., rubber and timber forests) in central Hainan, old‐growth forests and natural secondary growth forest at the margins of Hainan, and nature reserves on the island (e.g., Jianfeng and Diaoluo National Nature Reserves). Our study helps to elucidate potential botanical conservation priorities for Hainan within an evolutionary, phylogenetic framework.