Pleiotropic and Novel Phenotypes in The \u3cem\u3eDrosophila\u3c/em\u3e Gut Caused by Mutation of \u3cem\u3eDrop-Dead\u3c/em\u3e

Normal gut function is vital for animal survival, and deviations from such function can contribute to malnutrition, inflammation, increased susceptibility to pathogens, diabetes, neurodegenerative diseases, and cancer. In the fruit fly Drosophila melanogaster, mutation of the gene drop-dead (drd) results in defective gut function, as measured by enlargement of the crop and reduced food movement through the gut, and drd mutation also causes the unrelated phenotypes of neurodegeneration, early adult lethality and female sterility. In the current work, adult drd mutant flies are also shown to lack the peritrophic matrix (PM), an extracellular barrier that lines the lumen of the midgut and is found in many insects including flies, mosquitos and termites. The use of a drd-gal4 construct to drive a GFP reporter in late pupae and adults revealed drd expression in the anterior cardia, which is the site of PM synthesis in Drosophila. Moreover, the ability of drd knockdown or rescue with several gal4 drivers to recapitulate or rescue the gut phenotypes (lack of a PM, reduced defecation, and reduced adult survival 10–40 days post-eclosion) was correlated to the level of expression of each driver in the anterior cardia. Surprisingly, however, knocking down drd expression only in adult flies, which has previously been shown not to affect survival, eliminated the PM without reducing defecation rate. These results demonstrate that drd mutant flies have a novel phenotype, the absence of a PM, which is functionally separable from the previously described gut dysfunction observed in these flies. As the first mutant Drosophila strain reported to lack a PM, drd mutants will be a useful tool for studying the synthesis of this structure

Key inflammatory pathway activations in the MCI stage of Alzheimer's disease

OBJECTIVE: To determine the key inflammatory pathways that are activated in the peripheral and CNS compartments at the mild cognitive impairment (MCI) stage of Alzheimer's disease (AD). METHODS: A cross-sectional study of patients with clinical and biomarker characteristics consistent with MCI-AD in a discovery cohort, with replication in the Alzheimer's Disease Neuroimaging Initiative (ADNI) cohort. Inflammatory analytes were measured in the CSF and plasma with the same validated multiplex analyte platform in both cohorts and correlated with AD biomarkers (CSF Aβ42, total tau (t-tau), phosphorylated tau (p-tau) to identify key inflammatory pathway activations. The pathways were additionally validated by evaluating genes related to all analytes in coexpression networks of brain tissue transcriptome from an autopsy confirmed AD cohort to interrogate if the same pathway activations were conserved in the brain tissue gene modules. RESULTS: Analytes of the tumor necrosis factor (TNF) signaling pathway (KEGG ID:4668) in the CSF and plasma best correlated with CSF t-tau and p-tau levels, and analytes of the complement and coagulation pathway (KEGG ID:4610) best correlated with CSF Aβ42 levels. The top inflammatory signaling pathways of significance were conserved in the peripheral and the CNS compartments. They were also confirmed to be enriched in AD brain transcriptome gene clusters. INTERPRETATION: A cell-protective rather than a proinflammatory analyte profile predominates in the CSF in relation to neurodegeneration markers among MCI-AD patients. Analytes from the TNF signaling and the complement and coagulation pathways are relevant in evaluating disease severity at the MCI stage of AD

Fabrication and Selective Functionalization of Amine-Reactive Polymer Multilayers on Topographically Patterned Microwell Cell Culture Arrays

We report an approach to the fabrication and selective functionalization of amine-reactive polymer multilayers on the surfaces of 3-D polyurethane-based microwell cell culture arrays. Reactive layer-by-layer assembly of multilayers using branched polyethyleneimine (BPEI) and the azlactone- functionalized polymer poly(2-vinyl-4,4′-dimethylazlactone) (PVDMA) yielded film-coated microwell arrays that could be chemically functionalized postfabrication by treatment with different amine-functionalized macromolecules or small molecule primary amines. Treatment of film-coated arrays with the small molecule amine d-glucamine resulted in microwell surfaces that resisted the adhesion and proliferation of mammalian fibroblast cells in vitro. These and other experiments demonstrated that it was possible to functionalize different structural features of these arrays in a spatially resolved manner to create dual-functionalized substrates (e.g., to create arrays having either (i) azlactone-functionalized wells, with regions between the wells functionalized with glucamine or (ii) substrates with spatially resolved regions of two different cationic polymers). In particular, spatial control over glucamine functionalization yielded 3-D substrates that could be used to confine cell attachment and growth to microwells for periods of up to 28 days and support the 3-D culture of arrays of cuboidal cell clusters. These approaches to dual functionalization could prove useful for the long-term culture and maintenance of cell types for which the presentation of specific and chemically well-defined 3-D culture environments is required for control over cell growth, differentiation, and other important behaviors. More generally, our approach provides methods for the straightforward chemical functionalization of otherwise unreactive topographically patterned substrates that could prove to be useful in a range of other fundamental and applied contexts. © 2011 American Chemical Society

Key Inflammatory Pathway Activations in the MCI Stage of Alzheimer’s disease

Objective: To determine the key inflammatory pathways that are activated in the peripheral and CNS compartments at the mild cognitive impairment (MCI) stage of Alzheimer’s disease (AD). Methods: A cross-sectional study of patients with clinical and biomarker characteristics consistent with MCI-AD in a discovery cohort, with replication in the Alzheimer’s Disease Neuroimaging Initiative (ADNI) cohort. Inflammatory analytes were measured in the CSF and plasma with the same validated multiplex analyte platform in both cohorts and correlated with AD biomarkers (CSF Aβ42, total tau (t-tau), phosphorylated tau (p-tau) to identify key inflammatory pathway activations. The pathways were additionally validated by evaluating genes related to all analytes in coexpression networks of brain tissue transcriptome from an autopsy confirmed AD cohort to interrogate if the same pathway activations were conserved in the brain tissue gene modules. Results: Analytes of the tumor necrosis factor (TNF) signaling pathway (KEGG ID:4668) in the CSF and plasma best correlated with CSF t-tau and p-tau levels, and analytes of the complement and coagulation pathway (KEGG ID:4610) best correlated with CSF Aβ42 levels. The top inflammatory signaling pathways of significance were conserved in the peripheral and the CNS compartments. They were also confirmed to be enriched in AD brain transcriptome gene clusters. Interpretation: A cell-protective rather than a proinflammatory analyte profile predominates in the CSF in relation to neurodegeneration markers among MCI-AD patients. Analytes from the TNF signaling and the complement and coagulation pathways are relevant in evaluating disease severity at the MCI stage of AD

Evolutionary relationships in Panicoid grasses based on plastome phylogenomics (Panicoideae; Poaceae)

Background: Panicoideae are the second largest subfamily in Poaceae (grass family), with 212 genera and approximately 3316 species. Previous studies have begun to reveal relationships within the subfamily, but largely lack resolution and/or robust support for certain tribal and subtribal groups. This study aims to resolve these relationships, as well as characterize a putative mitochondrial insert in one linage. Results: 35 newly sequenced Panicoideae plastomes were combined in a phylogenomic study with 37 other species: 15 Panicoideae and 22 from outgroups. A robust Panicoideae topology largely congruent with previous studies was obtained, but with some incongruences with previously reported subtribal relationships. A mitochondrial DNA (mtDNA) to plastid DNA (ptDNA) transfer was discovered in the Paspalum lineage. Conclusions: The phylogenomic analysis returned a topology that largely supports previous studies. Five previously recognized subtribes appear on the topology to be non-monophyletic. Additionally, evidence for mtDNA to ptDNA transfer was identified in both Paspalum fimbriatum and P. dilatatum, and suggests a single rare event that took place in a common progenitor. Finally, the framework from this study can guide larger whole plastome sampling to discern the relationships in Cyperochloeae, Steyermarkochloeae, Gynerieae, and other incertae sedis taxa that are weakly supported or unresolved.Fil: Burke, Sean V.. Northern Illinois University; Estados UnidosFil: Wysocki, William P.. Northern Illinois University; Estados UnidosFil: Zuloaga, Fernando Omar. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Botánica Darwinion. Academia Nacional de Ciencias Exactas, Físicas y Naturales. Instituto de Botánica Darwinion; ArgentinaFil: Craine, Joseph M.. Jonah Ventures; Estados UnidosFil: Pires, J. Chris. University of Missouri; Estados UnidosFil: Edger, Patrick P.. Michigan State University; Estados UnidosFil: Mayfield Jones, Dustin. Donald Danforth Plant Science Center; Estados UnidosFil: Clark, Lynn G.. Iowa State University; Estados UnidosFil: Kelchner, Scot A.. University of Idaho; Estados UnidosFil: Duvall, Melvin R.. Northern Illinois University; Estados Unido

A 250 plastome phylogeny of the grass family (Poaceae): topological support under different data partitions

The systematics of grasses has advanced through applications of plastome phylogenomics, although studies have been largely limited to subfamilies or other subgroups of Poaceae. Here we present a plastome phylogenomic analysis of 250 complete plastomes (179 genera) sampled from 44 of the 52 tribes of Poaceae. Plastome sequences were determined from high throughput sequencing libraries and the assemblies represent over 28.7 Mbases of sequence data. Phylogenetic signal was characterized in 14 partitions, including (1) complete plastomes; (2) protein coding regions; (3) noncoding regions; and (4) three loci commonly used in single and multi-gene studies of grasses. Each of the four main partitions was further refined, alternatively including or excluding positively selected codons and also the gaps introduced by the alignment. All 76 protein coding plastome loci were found to be predominantly under purifying selection, but specific codons were found to be under positive selection in 65 loci. The loci that have been widely used in multi-gene phylogenetic studies had among the highest proportions of positively selected codons, suggesting caution in the interpretation of these earlier results. Plastome phylogenomic analyses confirmed the backbone topology for Poaceae with maximum bootstrap support (BP). Among the 14 analyses, 82 clades out of 309 resolved were maximally supported in all trees. Analyses of newly sequenced plastomes were in agreement with current classifications. Five of seven partitions in which alignment gaps were removed retrieved Panicoideae as sister to the remaining PACMAD subfamilies. Alternative topologies were recovered in trees from partitions that included alignment gaps. This suggests that ambiguities in aligning these uncertain regions might introduce a false signal. Resolution of these and other critical branch points in the phylogeny of Poaceae will help to better understand the selective forces that drove the radiation of the BOP and PACMAD clades comprising more than 99.9% of grass diversity

PhosphoregDB: The tissue and sub-cellular distribution of mammalian protein kinases and phosphatases

BACKGROUND: Protein kinases and protein phosphatases are the fundamental components of phosphorylation dependent protein regulatory systems. We have created a database for the protein kinase-like and phosphatase-like loci of mouse that integrates protein sequence, interaction, classification and pathway information with the results of a systematic screen of their sub-cellular localization and tissue specific expression data mined from the GNF tissue atlas of mouse. RESULTS: The database lets users query where a specific kinase or phosphatase is expressed at both the tissue and sub-cellular levels. Similarly the interface allows the user to query by tissue, pathway or sub-cellular localization, to reveal which components are co-expressed or co-localized. A review of their expression reveals 30% of these components are detected in all tissues tested while 70% show some level of tissue restriction. Hierarchical clustering of the expression data reveals that expression of these genes can be used to separate the samples into tissues of related lineage, including 3 larger clusters of nervous tissue, developing embryo and cells of the immune system. By overlaying the expression, sub-cellular localization and classification data we examine correlations between class, specificity and tissue restriction and show that tyrosine kinases are more generally expressed in fewer tissues than serine/threonine kinases. CONCLUSION: Together these data demonstrate that cell type specific systems exist to regulate protein phosphorylation and that for accurate modelling and for determination of enzyme substrate relationships the co-location of components needs to be considered

Pac13 is a small, monomeric dehydratase that mediates the formation of the 3′-deoxy nucleoside of pacidamycins

This work was supported by the EPSRC council (Grant number 1398501), Wellcome Trust (Investigator Award) and GlaxoSmithKline.The uridyl peptide antibiotics (UPAs), of which pacidamycin is a member, have a clinically unexploited mode of action and an unusual assembly. Perhaps the most striking feature of these molecules is the biosynthetically unique 3′-deoxyuridine that they share. This moiety is generated by an unusual, small and monomeric dehydratase, Pac13, which catalyses the dehydration of uridine-5’-aldehyde. Here we report the structural characterisation of Pac13 with a series of ligands, and gain insight into the enzyme’s mechanism demonstrating that H42 is critical to the enzyme’s activity and that the reaction is likely to proceed via an E1cB mechanism. The resemblance of the 3′-deoxy pacidamycin moiety with the synthetic anti-retrovirals, presents a potential opportunity for the utilisation of Pac13 in the biocatalytic generation of antiviral compounds.Publisher PDFPeer reviewe