Phylogenetic and Preliminary Phenotypic Analysis of Yeast PAQR Receptors: Potential Antifungal Targets

Proteins belonging to the Progestin and AdipoQ Receptor (PAQR) superfamily of membrane bound receptors are ubiquitously found in fungi. Nearly, all fungi possess two evolutionarily distinct paralogs of PAQR protein, which we have called the PQRA and PQRB subtypes. In the model fungus Saccharomyces cerevisiae, these subtypes are represented by the Izh2p and Izh3p proteins, respectively. S. cerevisiae also possesses two additional PQRA-type receptors called Izh1p and Izh4p that are restricted to other species within the “Saccharomyces complex”. Izh2p has been the subject of several recent investigations and is of particular interest because it regulates fungal growth in response to proteins produced by plants and, as such, represents a new paradigm for interspecies communication. We demonstrate that IZH2 and IZH3 gene dosage affects resistance to polyene antifungal drugs. Moreover, we provide additional evidence that Izh2p and Izh3p negatively regulate fungal filamentation. These data suggest that agonists of these receptors might make antifungal therapeutics, either by inhibiting fungal development or by sensitizing fungi to the toxic effects of current antifungal therapies. This is particularly relevant for pathogenic fungi such as Candida glabrata that are closely related to S. cerevisiae and contain the same complement of PAQR receptors

Analysis of the functional conservation of ethylene receptors between maize and Arabidopsis

Ethylene, a regulator of plant growth and development, is perceived by specific receptors that act as negative regulators of the ethylene response. Five ethylene receptors, i.e., ETR1, ERS1, EIN4, ETR2, and ERS2, are present in Arabidopsis and dominant negative mutants of each that confer ethylene insensitivity have been reported. In contrast, maize contains just two types of ethylene receptors: ZmERS1, encoded by ZmERS1a and ZmERS1b, and ZmETR2, encoded by ZmETR2a and ZmETR2b. In this study, we introduced a Cys to Tyr mutation in the transmembrane domain of ZmERS1b and ZmETR2b that is present in the etr1-1 dominant negative mutant and expressed each protein in Arabidopsis. Mutant Zmers1b and Zmetr2b receptors conferred ethylene insensitivity and Arabidopsis expressing Zmers1b or Zmetr2b were larger and exhibited a delay in leaf senescence characteristic of ethylene insensitive Arabidopsis mutants. Zmers1b and Zmetr2b were dominant and functioned equally well in a hemizygous or homozygous state. Expression of the Zmers1b N-terminal transmembrane domain was sufficient to exert dominance over endogenous Arabidopsis ethylene receptors whereas the Zmetr2b N-terminal domain failed to do so. Neither Zmers1b nor Zmetr2b functioned in the absence of subfamily 1 ethylene receptors, i.e., ETR1 and ERS1. These results suggest that Cys65 in maize ZmERS1b and ZmETR2b plays the same role that it does in Arabidopsis receptors. Moreover, the results demonstrate that the mutant maize ethylene receptors are functionally dependent on subfamily 1 ethylene receptors in Arabidopsis, indicating substantial functional conservation between maize and Arabidopsis ethylene receptors despite their sequence divergence

Fungal G-protein-coupled receptors::mediators of pathogenesis and targets for disease control

G-protein signalling pathways are involved in sensing the environment, enabling fungi to coordinate cell function, metabolism and development with their surroundings, thereby promoting their survival, propagation and virulence. G-protein-coupled receptors (GPCRs) are the largest class of cell surface receptors in fungi. Despite the apparent importance of GPCR signalling to fungal biology and virulence, relatively few GPCR–G-protein interactions, and even fewer receptor-binding ligands, have been identified. Approximately 40% of current pharmaceuticals target human GPCRs, due to their cell surface location and central role in cell signalling. Fungal GPCRs do not belong to any of the mammalian receptor classes, making them druggable targets for antifungal development. This Review Article evaluates developments in our understanding of fungal GPCR-mediated signalling, while substantiating the rationale for considering these receptors as potential antifungal targets. The need for insights into the structure–function relationship of receptor–ligand interactions is highlighted, which could facilitate the development of receptor-interfering compounds that could be used in disease control

Isolation And Partial Characterization Of An Anti-bothropic Complex From The Serum Of South American Didelphidae

An anti-bothropic fraction (ABF) with anti-Bothrops jararaca venom activity tested in mice was isolated from the serum of some South American Didelphidae (Didelphis marsupialis, Philander opossum and Lutreolina crassicaudata) by DEAE-Sephacel chromatography. ABF from D. marsupialis was shown to be 12 times more active in protection assays on a weight basis than the serum proteins. A similar fraction obtained from Metachirus nudicaudatum serum was shown to be inactive. An anti-bothropic complex (ABC) was isolated from D. marsupialis ABF. HPLC gel permeation chromatography of ABC from D. marsupialis indicated the presence of a main peak with mol. wt of 84,000. SDS-PAGE of this ABC showed the presence of two subunits of 48,000 and 43,000. The active ABF isolated from P. opossum and L. crassicaudata also showed the presence of these subunits by SDS-PAGE. Isolation of the 48,000 mol. wt D. marsupialis subunit by HPLC-hydrophobic interaction chromatography demonstrated that the 43,000 subunit was essential for the protective action of the complex. Both subunits from D. marsupialis, P. opossum and L. crassicaudata were Western-blotted and N-terminal sequenced. No N-terminal amino acid was found for the 43,000 subunit, whereas for the 48,000 subunit a high degree of homology was found:. D. marsupialis: H2N-L K A M D P T P P L W I K T E X P .;. L. crassicaudata: H2N-L K A M D P T P P L W I Q T E ...;. P. opossum: H2N-L K A M D T T P E .... No significant homology with known proteins was detected. © 1994.321012371249Andrews, Estimation of the molecular weights of proteins by Sephadex gel-filtration (1964) Biochem. J., 91, pp. 222-233Cerdas, Lomonte, Estudio de la capacidad ofiófaga y la resistencia de la zopilota (Clelia clelia Colubridae) de Costa Rica a los venenos de serpientes (1982) Toxicon, 20, pp. 936-939De Witt, Weström, Venom resistance in the hedgehog (Erinaceus europaeus) purification and identification of macroglobulin inhibitors as plasma antihemorrhagic factors (1987) Toxicon, 25, pp. 315-323Domont, Perales, Moussatché, Partial purification and physico-chemical characterization of a proteic complex from Philander opossum with protective action against Bothrops jararaca venom (1989) Brazilian-Sino Symposium on Chemistry and Pharmacology of Natural Products, , 10–14 December, Rio de Janeiro, BrazilDomont, Perales, Moussatché, Natural anti-snake venom proteins (1991) Toxicon, 29, pp. 1183-1194Fontana, (1781) Traité sur le venim de la vipére, p. 23. , FlorenceapudPhisalix, (1922) Animaux Venimeux et Venins, pp. 744-759. , Mason & Cie Editeurs, ParisFortes-Dias, Diniz, Kochva, Neutralization by homologous plasma of Crotalus durissus terrificus (South American rattlesnake) venom and crotoxin (1990) Ciênc. Cult., 42, pp. 501-506Garcia, Perez, The purification and characterization of an antihemorrhagic factor in woodrat (Neotoma micropus) serum (1984) Toxicon, 22, pp. 129-138Laemmli, Cleavage of structural proteins during the assembly of the head of Bacteriophage T-4 (1970) Nature, 227, pp. 608-609Landucci, Farah, Giglio, Domont, Marangoni, Protecting property of Didelphis albiventris serum against Bothrops jararaca snake venom (1990) Mem. Inst. Butantan, 52 (Supplement), p. 76Lowry, Rosebrough, Farr, Randall, Protein measurement with the Folin phenol reagent (1951) J. biol. Chem., 193, pp. 265-275Menchaca, Perez, The purification and characterization of an antihemorrhagic factor in opossum (Didelphis virginiana) serum (1981) Toxicon, 19, pp. 623-632Miranda, Couturier, Canziani, Seki, Vidal, Resistencia de Crotalus durissus terrificus y Bothrops neuwiedii a la neurotoxicidad de cantidades masivas de veneno crotálico (1982) Acta Physiol. Latinoamericana, 32, pp. 103-115Moussatché, Perales, Factors underlying the natural resistance of animals against snake venoms (1989) Memórias do Instituto Oswaldo Cruz, 84, pp. 391-394Moussatché, Yates, Leonardi, Borche, Experimentos sobre la resistencia del Didelphis venezolano (rabi-pelado) a los venenos de serpientes (1978) Acta Cient. Venezolana, 29, p. 55Moussatché, Yates, Leonardi, Borche, Mechanisms of resistance of the opossum to some snake venoms (1979) Toxicon, 17, p. 130Moussatché, Leonardi, Estudios de protección con sueros de mamiferos y reptiles a los venenos de serpientes Crotalidae (1982) Acta Cient. Venezolana, 33, p. 151Moussatché, Perales, Ferreira, Rocha, Villela, Domont, Resistencia de marsupiais brasileiros aos venenos de Bothrops jararaca e Crotalus durissus terrificus (1990) Vth Annual Meeting, Federation of Societies of Experimental Biology, MG, , BrazilOmori-Satoh, Antihemorrhagic factor as a proteinase inhibitor isolated from the serum of Trimeresurus flavoviridis (1977) Biochim. biophys. Acta, 495, pp. 93-98Omori-Satoh, Sadahiro, Ohsaka, Murata, Purification and characterization of an antihemorrhagic factor in the serum of Trimeresurus flavoviridis, a crotalidae (1972) Biochim. biophys. Acta, 285, pp. 414-426Ovadia, Purification and characterization of an antihemorrhagic factor from the serum of the snake Vipera palaestinae (1978) Toxicon, 16, pp. 661-672Ovadia, Kochva, Neutralization of Viperidae and Elapidae snake venoms by sera of different animals (1977) Toxicon, 15, pp. 541-547Ovadia, Moav, Kochva, Factors in the blood serum of Vipera palaestinae neutralizing fractions of its own venom (1975) Toxicon, 13, p. 113Ovadia, Kochva, Moav, The neutralization mechanism of Vipera palaestinae neurotoxin by a purified factor from homologous serum (1977) Biochim. biophys. Acta, 491, pp. 370-386Perales, Muñoz, Moussatché, Isolation and partial characterization of a protein fraction from the opossum (Didelphis marsupialis) serum, with protecting property against the Bothrops jararaca snake venom (1986) An. Acad. brasil. 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ISOLATION AND PARTIAL CHARACTERIZATION OF AN ANTI-BOTHROPIC COMPLEX FROM THE SERUM OF SOUTH-AMERICAN DIDELPHIDAE

An anti-bothropic fraction (ABF) with anti-Bothrops jararaca venom activity tested in mice was isolated from the serum of some South American Didelphidae (Didelphis marsupialis, Philander opossum and Lutreolina crassicaudata) by DEAE-Sephacel chromatography. ABF from D. marsupialis was shown to be 12 times more active in protection assays on a weight basis than the serum proteins. A similar fraction obtained from Metachirus nudicaudatum serum was shown to be inactive. An anti-bothropic complex (ABC) was isolated from D. marsupialis ABF. HPLC gel permeation chromatography of ABC from D. marsupialis indicated the presence of a main peak with mel. wt of 84,000. SDS-PAGE of this ABC showed the presence of two subunits of 48,000 and 43,000. The active ABF isolated from P. opossum and L. crassicaudata also showed the presence of these subunits by SDS-PAGE. Isolation of the 48,000 mol. wt D. marsupialis subunit by HPLC-hydrophobic interaction chromatography demonstrated that the 43,000 subunit was essential for the protective action of the complex. Both subunits from D. marsupialis, P. opossum and L. crassicaudata were Western-blotted and N-terminal sequenced. No N-terminal amino acid was found for the 43,000 subunit, whereas for the 48,000 subunit a high degree of homology was found: D. marsupialis: H2N-L K A M D P T P P L W I K T E X P . ; L. crassicaudata: H2N-L K A M D P T P P L W I Q T E . . . ; P. opossum: H2N-L K A M D T T P E . . . No significant homology with known proteins was detected.32101237124