Molecular Phylogeny and Evolution of the Plant-Specific Seven-Transmembrane MLO Family

Abstract Homologues of barley Mlo encode the only family of seven-transmembrane (TM) proteins in plants. Their topology, subcellular localization, and sequence diversification are reminiscent of those of G-protein coupled receptors (GPCRs) from animals and fungi. We present a computational analysis of MLO family members based on 31 full-size and 3 partial sequences, which originate from several monocot species, the dicot Arabidopsis thaliana, and the moss Ceratodon purpureus. This enabled us to date the origin of the Mlo gene family back at least to the early stages of land plant evolution. The genomic organization of the corresponding genes supports a monophyletic origin of the Mlo gene family. Phylogenetic analysis revealed five clades, of which three contain both monocot and dicot members, while two indicate class-specific diversification. Analysis of the ratio of nonsynonymous-to-synonymous changes in coding sequences provided evidence for functional constraint on the evolution of the DNA sequences and purifying selection, which appears to be reduced in the first extracellular loop of 12 closely related orthologues. The 31 full-size sequences were examined for potential domain-specific intramolecular coevolution. This revealed evidence for concerted evolution of all three cytoplasmic domains with each other and the C-terminal cytoplasmic tail, suggesting interplay of all intracellular domains for MLO function.     

Fibronectin induces macrophage migration through a SFK-FAK/CSF-1R pathway

Integrins, following binding to proteins of the extracellular matrix (ECM) including collagen, laminin and fibronectin (FN), are able to transduce molecular signals inside the cells and to regulate several biological functions such as migration, proliferation and differentiation. Besides activation of adaptor molecules and kinases, integrins transactivate Receptor Tyrosine Kinases (RTK). In particular, adhesion to the ECM may promote RTK activation in the absence of growth factors. The Colony-Stimulating Factor-1 Receptor (CSF-1R) is a RTK that supports the survival, proliferation, and motility of monocytes/macrophages, which are essential components of innate immunity and cancer development. Macrophage interaction with FN is recognized as an important aspect of host defense and wound repair. The aim of the present study was to investigate on a possible cross-talk between FN-elicited signals and CSF-1R in macrophages. FN induced migration in BAC1.2F5 and J774 murine macrophage cell lines and in human primary macrophages. Adhesion to FN determined phosphorylation of the Focal Adhesion Kinase (FAK) and Src Family Kinases (SFK) and activation of the SFK/FAK complex, as witnessed by paxillin phosphorylation. SFK activity was necessary for FAK activation and macrophage migration. Moreover, FN-induced migration was dependent on FAK in either murine macrophage cell lines or human primary macrophages. FN also induced FAK-dependent/ligand-independent CSF-1R phosphorylation, as well as the interaction between CSF-1R and β1. CSF-1R activity was necessary for FN-induced macrophage migration. Indeed, genetic or pharmacological inhibition of CSF-1R prevented FN-induced macrophage migration. Our results identified a new SFK-FAK/CSF-1R signaling pathway that mediates FN-induced migration of macrophages.     

Assessment of DNA damages in lymphocytes of agricultural workers exposed to pesticides by comet assay in a cross-sectional study

Purpose: To assess the predictive power of the comet assay in the context of occupational exposure to pesticides.

Materials and methods: The recruited subjects completed a structured questionnaire and gave a blood sample. Exposure to pesticides was measured by means of an algorithm based on Dosemeci’s work (Agricultural Health Study). Approximately 50 images were analyzed for each sample via fluorescence microscopy. The extent of DNA damage was estimated by tail moment (TM) and is the product of tail DNA (%) and tail Length.

Results: Crude significant risks (odds ratios, ORs) for values higher than the 75th percentile of TM were observed among the exposed subjects (score?>?1). The frequency of some confounding factors (sex, age and smoking) was significantly higher among the exposed workers. A significant dose–effect relationship was observed between TM and exposure score. Significant high-risk estimates (ORs), adjusted by the studied confounding factors, among exposure to pesticides and TM, % tail DNA and tail length were confirmed using unconditional logistic regression models.

Conclusions: The adjusted associations (ORs) between the comet parameters and exposure to pesticides were significant. The sensitivity of the comet test was low (41%), the specificity (89%) and the predictive positive value (0.77) were found acceptable.     

Molecular phylogeny and evolution of the plant-specific seven-transmembrane MLO family

Homologues of barley Mlo encode the only family of seven-transmembrane (TM) proteins in plants. Their topology, subcellular localization, and sequence diversification are reminiscent of those of G-protein coupled receptors (GPCRs) from animals and fungi. We present a computational analysis of MLO family members based on 31 full-size and 3 partial sequences, which originate from several monocot species, the dicot Arabidopsis thaliana, and the moss Ceratodon purpureus. This enabled us to date the origin of the Mlo gene family back at least to the early stages of land plant evolution. The genomic organization of the corresponding genes supports a monophyletic origin of the Mlo gene family. Phylogenetic analysis revealed five clades, of which three contain both monocot and dicot members, while two indicate class-specific diversification. Analysis of the ratio of nonsynonymous-to-synonymous changes in coding sequences provided evidence for functional constraint on the evolution of the DNA sequences and purifying selection, which appears to be reduced in the first extracellular loop of 12 closely related orthologues. The 31 full-size sequences were examined for potential domain-specific intramolecular coevolution. This revealed evidence for concerted evolution of all three cytoplasmic domains with each other and the C-terminal cytoplasmic tail, suggesting interplay of all intracellular domains for MLO function.     

Association of the quinate : NAD+ oxidoreductase with one dehydroquinate hydro-lyase isoenzyme in corn seedlings

A quinate : NAD⁺ oxidoreductase has been purified from cornseedlings. This enzyme co-migrates with a dehydroquinate hydro-lyaseisoenzyme whatever separation technic is used. This is strongevidence that the two activities are associated in an enzymecomplex or in a bifunctional enzyme, in addition to the previouslycharacterized association of the shikimate : NADP⁺ oxidoreductaseand another dehydroquinate hydro-lyase isoenzyme. The purifiedquinate : NAD⁺ oxidoreductase has a poor affinity for quinicacid and is only active in the presence of NAD⁺. The associateddehydroquinate hydro-lyase isoenzyme is strongly activated invitro by shikimic acid in a pH-dependent process. The possible role of this new association is discussed in thelight of previous results from alicyclic metabolism studiesin plants and microorganisms. (Received July 18, 1980; )     

A calcium-binding protein is a regulatory subunit of quinate:NAD+ oxidoreductase from dark-grown carrot cells

On transfer of carrot cell-suspensions from light to dark conditions, quinate:NAD+ oxidoreductase binds, post-translationally, an additional subunit. As a consequence, the oligomeric enzyme becomes activatable by Ca++ and behaves as a Ca++ binding protein. The additional subunit has been shown to be the Ca++ binding moiety of the molecule that protects the phosphorylated amino acids from dephosphorylation. It is suggested that the regulatory subunit may be a new class of calciprotein.     

A Functional Topographic Map for Spinal Sensorimotor Reflexes

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Mechanical Allodynia Circuitry in the Dorsal Horn Is Defined by the Nature of the Injury

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Clobetasol promotes neuromuscular plasticity in mice after motoneuronal loss via sonic hedgehog signaling,immunomodulation and metabolic rebalancing

Motoneuronal loss is the main feature of amyotrophic lateral sclerosis, although pathogenesis is extremely complex involving both neural and muscle cells. In order to translationally engage the sonic hedgehog pathway, which is a promising target for neural regeneration, recent studies have reported on the neuroprotective effects of clobetasol, an FDA-approved glucocorticoid, able to activate this pathway via smoothened. Herein we sought to examine functional, cellular, and metabolic effects of clobetasol in a neurotoxic mouse model of spinal motoneuronal loss. We found that clobetasol reduces muscle denervation and motor impairments in part by restoring sonic hedgehog signaling and supporting spinal plasticity. These effects were coupled with reduced pro-inflammatory microglia and reactive astrogliosis, reduced muscle atrophy, and support of mitochondrial integrity and metabolism. Our results suggest that clobetasol stimulates a series of compensatory processes and therefore represents a translational approach for intractable denervating and neurodegenerative disorders.Subject terms: Biochemistry, Diseases of the nervous system, Glial biology, Physiology     

Association of the quinate : NAD+ oxidoreductase with one dehydroquinate hydro-lyase isoenzyme in corn seedlings

A quinate : NAD⁺ oxidoreductase has been purified from cornseedlings. This enzyme co-migrates with a dehydroquinate hydro-lyaseisoenzyme whatever separation technic is used. This is strongevidence that the two activities are associated in an enzymecomplex or in a bifunctional enzyme, in addition to the previouslycharacterized association of the shikimate : NADP⁺ oxidoreductaseand another dehydroquinate hydro-lyase isoenzyme. The purifiedquinate : NAD⁺ oxidoreductase has a poor affinity for quinicacid and is only active in the presence of NAD⁺. The associateddehydroquinate hydro-lyase isoenzyme is strongly activated invitro by shikimic acid in a pH-dependent process. The possible role of this new association is discussed in thelight of previous results from alicyclic metabolism studiesin plants and microorganisms. (Received July 18, 1980; )