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genetic analysis of the neurospora crassa rad14 homolog mus-43 and the rad10 homolog mus-44 reveals that they belong to the mus-38 pathway of two nucleotide excision repair systems.saccharomyces cerevisiae rad14 and rad10 proteins are essential for nucleotide excision repair (ner). rad14 is a uv-damaged dna binding protein and rad10 is a structure-specific endonuclease that functions in a complex with rad1. in this study, we identified and characterized the rad14 and rad10 homolog genes in neurospora crassa, which we named mus-43 and mus-44, respectively. disruption of mus-43 and mus-44 conferred sensitivity to uv and 4-nitroquinoline 1-oxide, but not to methyl methanesulf ...200818379129
lack of the gtpase rho-4 in neurospora crassa causes a reduction in numbers and aberrant stabilization of microtubules at hyphal tips.the multinucleate hyphae of the filamentous ascomycete fungus neurospora crassa grow by polarized hyphal tip extension. both the actin and microtubule cytoskeleton are required for maximum hyphal extension, in addition to other vital processes. previously, we have shown that the monomeric gtpase encoded by the n. crassa rho-4 locus is required for actin ring formation during the process of septation; rho-4 mutants lack septa. however, other phenotypic aspects of the rho-4 mutant, such as slow gr ...200818387834
the evolutionarily conserved family of cyanovirin-n homologs: structures and carbohydrate specificity.solution structures for three members of the recently discovered cyanovirin-n (cv-n) homolog family of lectins have been determined. cyanovirin-n homologs (cvnhs) from tuber borchii, ceratopteris richardii, and neurospora crassa, representing each of the three phylogenetic groups, were selected. all proteins exhibit the same fold, and the overall structures resemble that of the founding member of the family, cv-n, albeit with noteworthy differences in loop conformation and detailed local structu ...200818400178
identification and evolution of fungal mitochondrial tyrosyl-trna synthetases with group i intron splicing activity.the bifunctional neurospora crassa mitochondrial tyrosyl-trna synthetase (cyt-18 protein) both aminoacylates mitochondrial trna(tyr) and acts as a structure-stabilizing splicing cofactor for group i introns. previous studies showed that cyt-18 has distinct trna(tyr) and group i intron-binding sites, with the latter formed by three small "insertions" in the nucleotide-binding fold and other structural adaptations compared with nonsplicing bacterial tyrosyl-trna synthetases. here, analysis of geno ...200818413600
biological rhythms workshop ia: molecular basis of rhythms generation.current circadian models are based on genetic, biochemical, and structural data that, when combined, provide a comprehensive picture of the molecular basis for rhythms generation. these models describe three basic elements-input pathways, oscillator, and output pathways-to which each molecular component is assigned. the lines between these elements are often blurred because some proteins function in more than one element of the circadian system. the end result of these molecular oscillations is ...200718419259
circadian entrainment of neurospora crassa.the circadian clock evolved under entraining conditions, yet most circadian experiments and much circadian theory are built around free-running rhythms. the interpretation of entrainment experiments is certainly more complex than that of free-running rhythms due to the relationship between exogenous and endogenous cycles. here, we systematically describe entrainment in the simplest of the traditional eukaryotic model systems in circadian research, neurospora crassa. this fungus forms a mass of s ...200718419284
complexity of the neurospora crassa circadian clock system: multiple loops and oscillators.organisms from bacteria to humans use a circadian clock to control daily biochemical, physiological, and behavioral rhythms. we review evidence from neurospora crassa that suggests that the circadian clock is organized as a network of genes and proteins that form coupled evening- and morning-specific oscillatory loops that can function autonomously, respond differently to environmental inputs, and regulate phase-specific outputs. there is also evidence for coupled morning and evening oscillator ...200718419292
an f-actin-depleted zone is present at the hyphal tip of invasive hyphae of neurospora crassa.the distribution of filamentous actin (f-actin) in invasive and noninvasive hyphae of the ascomycete neurospora crassa was investigated. eighty six percent of noninvasive hyphae had f-actin in the tip region compared to only 9% of invasive hyphae. the remaining 91% of the invasive hyphae had no obvious tip high concentration of f-actin staining; instead they had an f-actin-depleted zone in this region, although some f-actin, possibly associated with the spitzenkörper, remained at the tip. the si ...200818421548
effects of oxygen limitation on xylose fermentation, intracellular metabolites, and key enzymes of neurospora crassa as3.1602.the effects of oxygen limitation on xylose fermentation of neurospora crassa as3.1602 were studied using batch cultures. the maximum yield of ethanol was 0.34 g/g at oxygen transfer rate (otr) of 8.4 mmol/l.h. the maximum yield of xylitol was 0.33 g/g at otr of 5.1 mmol/l.h. oxygen limitation greatly affected mycelia growth and xylitol and ethanol productions. the specific growth rate (micro) decreased 82% from 0.045 to 0.008 h(-1) when otr changed from 12.6 to 8.4 mmol/l.h. intracellular metabo ...200818425610
a systematic approach to identify stre-binding proteins of the gsn glycogen synthase gene promoter in neurospora crassa.the gene encoding glycogen synthase in neurospora crassa (gsn) is transcriptionally down-regulated when mycelium is exposed to a heat shock from 30 to 45 degrees c. the gsn promoter has one stress response element (stre) motif that is specifically bound by heat shock activated nuclear proteins. in this work, we used biochemical approaches together with mass spectrometric analysis to identify the proteins that bind to the stre motif and could participate in the gsn transcription regulation during ...200818425733
dynamin-like protein-dependent formation of woronin bodies in saccharomyces cerevisiae upon heterologous expression of a single protein.filamentous ascomycetes harbor woronin bodies and glyoxysomes, two types of microbodies, within one cell at the same time. the dominant protein of the neurospora crassa woronin body, hex1, forms a hexagonal core crystal via oligomerization and evidence has accumulated that woronin bodies bud off from glyoxysomes. we analyzed whether hex1 is sufficient to induce woronin body formation upon heterologous expression in saccharomyces cerevisiae, an organism devoid of this specialized organelle. in wi ...200818435762
dynamics of the preprotein translocation channel of the outer membrane of mitochondria.the protein translocase of the outer mitochondrial membrane (tom) serves as the main entry site for virtually all mitochondrial proteins. like many other protein translocases it also has an ion channel activity that can be used to study the dynamical properties of this supramolecular complex. we have purified tom core complex and tom40, the main pore forming subunit, from mitochondria of the filamentous fungus neurospora crassa and incorporated them into planar lipid bilayers. we then examined t ...200818456827
the genome sequence of the model ascomycete fungus podospora anserina.the dung-inhabiting ascomycete fungus podospora anserina is a model used to study various aspects of eukaryotic and fungal biology, such as ageing, prions and sexual development.200818460219
structural insights into biosynthesis of resorcinolic lipids by a type iii polyketide synthase in neurospora crassa.microbial type iii polyketide synthases (pkss) have revealed remarkable mechanistic as well as functional versatility. recently, a type iii pks homolog from azotobacter has been implicated in the biosynthesis of resorcinolic lipids, thus adding a new functional significance to this class of proteins. here, we report the structural and mutational investigations of a novel type iii pks protein from neurospora crassa involved in the biosynthesis of resorcinolic metabolites by utilizing long chain f ...200818462950
increased resistance of complex i mutants to phytosphingosine-induced programmed cell death.we have studied the effects of phytosphingosine (phs) on cells of the filamentous fungus neurospora crassa. highly reduced viability, impairment of asexual spore germination, dna condensation and fragmentation, and production of reactive oxygen species were observed in conidia treated with the drug, suggesting that phs induces an apoptosis-like death in this fungus. interestingly, we found that complex i mutants are more resistant to phs treatment than the wild type strain. this effect appears t ...200818474589
sexual and vegetative compatibility genes in the aspergilli.gene flow within populations can occur by sexual and/or parasexual means. analyses of experimental and in silico work are presented relevant to possible gene flow within the aspergilli. first, the discovery of mating-type (mat) genes within certain species of aspergillus is described. the implications for self-fertility, sexuality in supposedly asexual species and possible uses as phylogenetic markers are discussed. second, the results of data mining for heterokaryon incompatibility (het) and pr ...200718490952
[stress factor-induced changes in the activity of antioxidant protective mechanisms in the wild type strain of neurospora crassa and in its photoreceptor complex mutants].the effect of stress factors (changes in oxygen content, temperature, and illumination) on superoxide dismutase (sod) and catalase activity, as well as on the content of thiol and disulfide groups in low-molecular-weight compounds and proteins of neurospora crassa mycelium was studied in the wild type strain and white collar-1 (wc-1) and white collar-2 (wc-2) mutants. environmental stress factors induced the activation of both sod and catalase, as well as an increase in the thiol level in the wi ...200818522316
genetic characterization of morphologically variant strains of paracoccidioides brasiliensis.molecular characterization of paracoccidioides brasiliensis variant strains that had been preserved under mineral oil for decades was carried out by random amplified polymorphic dna analysis (rapd). on p. brasiliensis variants in the transitional phase and strains with typical morphology, rapd produced reproducible polymorphic amplification products that differentiated them. a dendrogram based on the generated rapd patterns placed the 14 p. brasiliensis strains into five groups with similarity c ...200818545858
light activation of the lov protein vivid generates a rapidly exchanging dimer.the fungal photoreceptor vivid (vvd) plays an important role in the adaptation of blue-light responses in neurospora crassa. vvd, an fad-binding lov (light, oxygen, voltage) protein, couples light-induced cysteinyl adduct formation at the flavin ring to conformational changes in the n-terminal cap (ncap) of the vvd pas domain. size-exclusion chromatography (sec), equilibrium ultracentrifugation, and static and dynamic light scattering show that these conformational changes generate a rapidly exc ...200818553928
mode of action and properties of xylanase and beta-xylosidase from neurospora crassa.extracellular beta-xylosidase (1,4-beta-d-xylan xylohydrolase, ec 3.2.1.37) from culture filtrates of neurospora crassa was purified to homogeneity by preparative isoelectric focusing followed by gel electrophoresis. the molecular weight of the purified xylosidase was 83,000 d and the k(m) on p-nitrophenyl-beta-d-xyloside was 0.047mm. the homogeneous xylanase (1,4-beta-d-xylan xylanohydrolase, ec 3.2.1.8) and beta-xylosidase showed differences in their mode of action towards xylooligosaccharides ...198618555300
the nuclear dbf2-related kinase cot1 and the mitogen-activated protein kinases mak1 and mak2 genetically interact to regulate filamentous growth, hyphal fusion and sexual development in neurospora crassa.ndr kinases, such as neurospora crassa cot1, are important for cell differentiation and polar morphogenesis, yet their input signals as well as their integration into a cellular signaling context are still elusive. here, we identify the cot-1 suppressor gul-4 as mak-2 and show that mutants of the gul-4/mak-2 mitogen-activated protein (map) kinase pathway suppress cot-1 phenotypes along with a concomitant reduction in protein kinase a (pka) activity. furthermore, mak-2 pathway defects are partial ...200818562669
osrecq1, a qde-3 homologue in rice, is required for rna silencing induced by particle bombardment for inverted repeat dna, but not for double-stranded rna.based on the nucleotide sequence of qde-3 in neurospora crassa, which is involved in rna silencing, rice (oryza sativa) mutant lines disrupted by the insertion of the rice retrotransposon tos17 were selected. homozygous individuals from the m(1) and m(2) generations were screened and used for further analyses. the expression of the gene was not detected in leaves or calli of the mutant lines, in contrast to the wild type (wt). induction of rna silencing by particle bombardment was performed to i ...200818564381
dna-binding specificity of the cys3 transcription factor of neurospora crassa defined by binding-site selection.the cys3 transcription factor is a basic region-leucine zipper (bzip) dna-binding protein that is essential for the expression of a coordinately regulated group of genes involved in the acquisition and utilization of sulfur in neurospora crassa. an approach of using binding-site selection from random-sequence oligonucleotides was used to define cys3-binding specificity. the derived consensus-binding site of atggcgccat defines a symmetrical sequence (half-site a t g/t g/a c/t) that resembles that ...200818565773
nadph oxidases nox-1 and nox-2 require the regulatory subunit nor-1 to control cell differentiation and growth in neurospora crassa.we have proposed that reactive oxygen species (ros) play essential roles in cell differentiation. enzymes belonging to the nadph oxidase (nox) family produce superoxide in a regulated manner. we have identified three distinct nox subfamilies in the fungal kingdom and have shown that noxa is required for sexual cell differentiation in aspergillus nidulans. here we show that neurospora crassa nox-1 elimination results in complete female sterility, decreased asexual development, and reduction of hy ...200818567788
multiple gene disruptions by marker recycling with highly efficient gene-targeting background (deltaligd) in aspergillus oryzae.previously we reported that double disruption of the proteinase genes (tppa and pepe) improved heterologous protein production by aspergillus oryzae (jin et al. appl microbiol biotechnol 76:1059-1068, 2007). since a. oryzae has 134 protease genes, the number of auxotrophy in a single host is limited for multiple disruptions of many protease genes. in order to rapidly perform multiple gene disruptions in a. oryzae, we generated the marker recycling system in highly efficient gene-targeting backgr ...200818574559
a cytosine methyltransferase homologue is essential for sexual development in aspergillus nidulans.the genome defense processes rip (repeat-induced point mutation) in the filamentous fungus neurospora crassa, and mip (methylation induced premeiotically) in the fungus ascobolus immersus depend on proteins with dna methyltransferase (dmt) domains. nevertheless, these proteins, rid and masc1, respectively, have not been demonstrated to have dmt activity. we discovered a close homologue in aspergillus nidulans, a fungus thought to have no methylation and no genome defense system comparable to rip ...200818575630
the production and properties of a new xylose reductase from fungus. neurospora crassa.neurospora crassa xi was found to ferment xylose and glucose simultaneously. xylose was the appropriate inducer for the production of xylose reductase that had two isoenzymes designated as ei and eii. both ei and eii, which were purified by affinity chromatography, had nadph-dependent xylose reductase activities. eii also had nadh-dependent activity, and ei is the only xylose reductase found so far without any nadh-dependent activity. ei and eii had mws of 30 kda and 27 kda, and pis of 5.6 and 5 ...199818576008
induced pelletized growth of neurospora crassa for tyrosinase biosynthesis in airlift fermenters. 198718576515
neurospora crassa heat shock factor 1 is an essential gene; a second heat shock factor-like gene, hsf2, is required for asexual spore formation.appropriate responses of organisms to heat stress are essential for their survival. in eukaryotes, adaptation to high temperatures is mediated by heat shock transcription factors (hsfs). hsfs regulate the expression of heat shock proteins, which function as molecular chaperones assisting in protein folding and stability. in many model organisms a great deal is known about the products of hsf genes. an important exception is the filamentous fungus and model eukaryote neurospora crassa. here we sh ...200818586951
genome mining of cyanide-degrading nitrilases from filamentous fungi.a variety of fungal species are known to degrade cyanide through the action of cyanide hydratases, a specialized subset of nitrilases which hydrolyze cyanide to formamide. in this paper, we report on two previously unknown and uncharacterized cyanide hydratases from neurospora crassa and aspergillus nidulans. recombinant forms of four cyanide hydratases from n. crassa, a. nidulans, gibberella zeae, and gloeocercospora sorghi were prepared after their genes were cloned with n-terminal hexahistidi ...200818587571
differential potency of drosomycin to neurospora crassa and its mutant: implications for evolutionary relationship between defensins from insects and plants.drosomycin, the first inducible antifungal peptide isolated from drosophila, belongs to the superfamily of csalphabeta-type defensins. in the present study we report a modified approach for high-level expression of drosomycin, which allows us to evaluate its differential potency on the filamentous fungus neurospora crassa wt (wild type) and n. crassa mut16, a specific resistance mutant strain to plant defensins, by using different approaches. the results presented here show for the first time th ...200818651922
an endophytic neurospora sp. from nothapodytes foetida producing camptothecin.the medicinal plant, nothapodytes foetida contains a number of important alkaloids like camptothecin (an anticancer drug molecule) but its concentration is less to meet the existing demand of this important molecule, so in an effort for accessible availability of camptothecin. an endophyte (designated zp5se) was isolated from the seed of nothapodytes foetida and was examined as potential source of anticancer drug lead compound i.e. camptothecin, when grown in sabouraud liquid culture media under ...201318669267
the cys3-cys4 loop of the hydrophobin eas is not required for rodlet formation and surface activity.class i hydrophobins are fungal proteins that self-assemble into robust amphipathic rodlet monolayers on the surface of aerial structures such as spores and fruiting bodies. these layers share many structural characteristics with amyloid fibrils and belong to the growing family of functional amyloid-like materials produced by microorganisms. although the three-dimensional structure of the soluble monomeric form of a class i hydrophobin has been determined, little is known about the molecular str ...200818674544
dissecting colony development of neurospora crassa using mrna profiling and comparative genomics approaches.colony development, which includes hyphal extension, branching, anastomosis, and asexual sporulation, is a fundamental aspect of the life cycle of filamentous fungi; genetic mechanisms underlying these phenomena are poorly understood. we conducted transcriptional profiling during colony development of the model filamentous fungus neurospora crassa, using 70-mer oligonucleotide microarrays. relative mrna expression levels were determined for six sections of defined age excised from a 27-h-old n. ...200818676954
direct interaction between dna methyltransferase dim-2 and hp1 is required for dna methylation in neurospora crassa.dna methylation is involved in gene silencing and genomic stability in mammals, plants, and fungi. genetics studies of neurospora crassa have revealed that a dna methyltransferase (dim-2), a histone h3k9 methyltransferase (dim-5), and heterochromatin protein 1 (hp1) are required for dna methylation. we explored the interrelationships of these components of the methylation machinery. a yeast two-hybrid screen revealed that hp1 interacts with dim-2. we confirmed the interaction in vivo and demonst ...200818678653
the nonoxidative conversion of nitroethane to ethylnitronate in neurospora crassa 2-nitropropane dioxygenase is catalyzed by histidine 196.the deprotonation of nitroethane catalyzed by neurospora crassa 2-nitropropane dioxygenase was investigated by measuring the formation and release of ethylnitronate formed in turnover as a function of ph and through mutagenesis studies. progress curves for the enzymatic reaction obtained by following the increase in absorbance at 228 nm over time were visibly nonlinear, requiring a logarithmic approximation of the initial reaction rates for the determination of the kinetic parameters of the enzy ...200818690716
evolutionary history of the ancient cutinase family in five filamentous ascomycetes reveals differential gene duplications and losses and in magnaporthe grisea shows evidence of sub- and neo-functionalization.* the cuticle is the first barrier for fungi that parasitize plants systematically or opportunistically. here, the evolutionary history is reported of the multimembered cutinase families of the plant pathogenic ascomycetes magnaporthe grisea, fusarium graminearum and botrytis cinerea and the saprotrophic ascomycetes aspergillus nidulans and neurospora crassa. * molecular taxonomy of all fungal cutinases demonstrates a clear division into two ancient subfamilies. no evidence was found for lateral ...200818713314
the rho1-specific gtpase-activating protein lrg1 regulates polar tip growth in parallel to ndr kinase signaling in neurospora.regulation of rho gtpase signaling is critical for cell shape determination and polarity. here, we investigated the role of lrg1, a novel member of the gtpase-activating proteins (gaps) of neurospora crassa. lrg1 is essential for apical tip extension and to restrict excessive branch formation in subapical regions of the hypha and is involved in determining the size of the hyphal compartments. lrg1 localizes to hyphal tips and sites of septation via its three lim domains. the accumulation of lrg1 ...200818716060
a connection between mapk pathways and circadian clocks.circadian clocks and mitogen-activated protein kinase (mapk) signaling pathways are fundamental features of eukaryotic cells. both pathways provide mechanisms for cells to respond to environmental stimuli, and links between them are known. we recently reported that the circadian clock in neurospora crassa regulates daily rhythms in accumulation of phosphorylated, and thus active, os-2 mapk, a relative of mammalian p38 mapk, when cells are grown in constant conditions. in the absence of acute str ...200818728391
toward predicting self-splicing and protein-facilitated splicing of group i introns.in the current era of massive discoveries of noncoding rnas within genomes, being able to infer a function from a nucleotide sequence is of paramount interest. although studies of individual group i introns have identified self-splicing and nonself-splicing examples, there is no overall understanding of the prevalence of self-splicing or the factors that determine it among the >2300 group i introns sequenced to date. here, the self-splicing activities of 12 group i introns from various organisms ...200818768647
systems biology of the clock in neurospora crassa.a model-driven discovery process, computing life, is used to identify an ensemble of genetic networks that describe the biological clock. a clock mechanism involving the genes white-collar-1 and white-collar-2 (wc-1 and wc-2) that encode a transcriptional activator (as well as a blue-light receptor) and an oscillator frequency (frq) that encodes a cyclin that deactivates the activator is used to guide this discovery process through three cycles of microarray experiments. central to this discover ...200818769678
mitop2: an integrative tool for the analysis of the mitochondrial proteome.mitochondria are crucial for normal cell metabolism and maintenance. mitochondrial dysfunction has been implicated in a spectrum of human diseases, ranging from rare monogenic to common multifactorial disorders. important for the understanding of organelle function is the assignment of its constituents, and although over 1,500 proteins are predicted to be involved in mammalian mitochondrial function, so far only about 900 are assigned to mitochondria with reasonable certainty. continuing efforts ...200818780189
genetic analysis of chk1 and chk2 homologues revealed a unique cross talk between atm and atr pathways in neurospora crassa.dna damage checkpoint is an important mechanism for organisms to maintain genome integrity. in neurospora crassa, mus-9 and mus-21 are homologues of atr and atm, respectively, which are pivotal factors of dna damage checkpoint in mammals. a n. crassa clock gene prd-4 has been identified as a chk2 homologue, but its role in dna damage response had not been elucidated. in this study, we identified another chk2 homologue and one chk1 homologue from the n. crassa genome database. as disruption of th ...200818790091
plasticity of a key metabolic pathway in fungi.beta oxidation is the principal metabolic pathway for fatty acid degradation. the pathway is virtually universally present throughout eukaryotes yet displays different forms in enzyme architecture, substrate specificity, and subcellular location. in this review, we examine beta oxidation across the fungal kingdom by conducting a large-scale in silico screen and localization prediction for all relevant enzymes in >50 species. the survey reveals that fungi exhibit an astounding diversity of beta o ...200918795352
the information highways of a biotechnological workhorse--signal transduction in hypocrea jecorina.the ascomycete hypocrea jecorina (anamorph trichoderma reesei) is one of the most prolific producers of biomass-degrading enzymes and frequently termed an industrial workhorse. to compete for nutrients in its habitat despite its shortcoming in certain degradative enzymes, efficient perception and interpretation of environmental signals is indispensable. a better understanding of these signals as well as their transmission machinery can provide sources for improvement of biotechnological processe ...200818803869
novel apocarotenoid intermediates in neurospora crassa mutants imply a new biosynthetic reaction sequence leading to neurosporaxanthin formation.neurosporaxanthin, beta-apo-4'-carotenoic acid (c35), represents the end-product of the carotenoid pathway in neurospora crassa. it is supposed to be synthesized in three steps catalyzed by sequential al-2, cao-2 and ylo-1 activities: (i) cyclization of 3,4-didehydrolycopene (c40); (ii) cleavage of torulene into beta-apo-4'-carotenal (c35); and finally (iii) oxidation of beta-apo-4'-carotenal. however, analyses of the ylo-1 mutant revealed the accumulation of intermediates other than beta-apo-4' ...200818812228
the growth of the isolated hyphal apices of neurospora crassa under carbon starvation. 201618841815
sulphur metabolism and cellulase gene expression are connected processes in the filamentous fungus hypocrea jecorina (anamorph trichoderma reesei).sulphur compounds like cysteine, methionine and s-adenosylmethionine are essential for the viability of most cells. thus many organisms have developed a complex regulatory circuit that governs the expression of enzymes involved in sulphur assimilation and metabolism. in the filamentous fungus hypocrea jecorina (anamorph trichoderma reesei) little is known about the participants in this circuit.200818842142
mitogen-activated protein kinase cascade required for regulation of development and secondary metabolism in neurospora crassa.mitogen-activated protein kinase (mapk) signaling cascades are composed of mapk kinase kinases (mapkkks), mapk kinases (mapkks), and mapks. in this study, we characterize components of a mapk cascade in neurospora crassa (mik-1, mapkkk; mek-1, mapkk; and mak-1, mapk) homologous to that controlling cell wall integrity in saccharomyces cerevisiae. growth of basal hyphae is significantly reduced in mik-1, mek-1, and mak-1 deletion mutants on solid medium. all three mutants formed short aerial hypha ...200818849472
synchronization of the fungal and the plant circadian clock by light.circadian clocks are endogenous time keeping devices that provide temporal control of physiology in accordance with predicted daily changes in the environment. photoentrainment is the process that synchronizes circadian clocks-and thereby clock-controlled gene expression and physiology-to the environmental day/night cycles. light is primarily detected by specialized photoreceptors that are coupled--directly or through other signaling components--to the rhythm-generating oscillator. as a conseque ...200818850603
rapid snp discovery and genetic mapping using sequenced rad markers.single nucleotide polymorphism (snp) discovery and genotyping are essential to genetic mapping. there remains a need for a simple, inexpensive platform that allows high-density snp discovery and genotyping in large populations. here we describe the sequencing of restriction-site associated dna (rad) tags, which identified more than 13,000 snps, and mapped three traits in two model organisms, using less than half the capacity of one illumina sequencing run. we demonstrated that different marker d ...200818852878
mglig4, a homolog of neurospora crassa mus-53 (dna ligase iv), is involved in, but not essential for, non-homologous end-joining events in magnaporthe grisea.in many eukaryotic organisms, the non-homologous end-joining (nhej) system is a major pathway for the repair of dna double-strand breaks (dsbs). dna ligase iv is a component of the nhej system and is strictly required for the nhej system in saccharomyces cerevisiae and in neurospora crassa. to investigate the functions of dna ligase iv in magnaporthe grisea, we generated deletion mutants of mglig4, which encodes a homolog of n. crassa dna ligase iv. mutants (mglig4) showed no defects in asexual ...200818854221
the induction of biochemical mutations in neurospora crassa by nitrogen mustard. 194718896929
somatic segregation by microconidial isolation in synthesized heterokaryons of neurospora crassa. 194718899678
induced reversions of biochemical mutants in neurospora crassa. 194818909961
mapping the interaction sites of aspergillus nidulans phytochrome fpha with the global regulator vea and the white collar protein lreb.aspergillus nidulans senses red and blue-light and employs a phytochrome and a neurospora crassa white collar (wc) homologous system for light perception and transmits this information into developmental decisions. under light conditions it undergoes asexual development and in the dark it develops sexually. the phytochrome fpha consists of a light sensory domain and a signal output domain, consisting of a histidine kinase and a response regulator domain. previously it was shown that the phytochr ...200918936976
distinct structural elements dictate the specificity of the type iii pentaketide synthase from neurospora crassa.the fungal type iii polyketide synthase 2'-oxoalkylresorcylic acid synthase (oras) primes with a range of acyl-coenzyme a thioesters (c4-c20) and extends using malonyl-coenzyme a to produce pyrones, resorcinols, and resorcylic acids. to gain insight into this unusual substrate specificity and product profile, we have determined the crystal structures of oras to 1.75 a resolution, the phe-252-->gly site-directed mutant to 2.1 a resolution, and a binary complex of oras with eicosanoic acid to 2.0 ...200818940668
a point mutation in the two-component histidine kinase bcos-1 gene confers dicarboximide resistance in field isolates of botrytis cinerea.abstract partial dna fragments of botrytis cinerea field isolates encoding the putative osmosensor histidine kinase gene (bcos1) were cloned by polymerase chain reaction amplification and the predicted amino acid sequences were compared between dicarboximide-sensitive and resistant field isolates. the predicted bcos1p is highly homologous to osmosensor histidine kinase os1p from neurospora crassa including the n-terminal six tandem repeats of approximately 90 amino acids. four dicarboximide-resi ...200218944142
the mycoparasite ampelomyces quisqualis expresses exga encoding an exo-beta-1,3-glucanase in culture and during mycoparasitism.abstract ampelomyces quisqualis, a mycoparasite of fungi causing powdery mildews, exhibited high levels of extracellular exo-beta-1,3-glucanase activity in culture compared with neurospora crassa and gliocladium roseum. a. quisqualis culture filtrates affected powdery mildew caused by sphaerotheca fusca in a manner indicative of cell wall degradation, as determined by microscopic examination. a gene encoding an exo-beta-1,3-glucanase in a. quisqualis, designated exga, was isolated and sequenced. ...199918944674
the cyanide hydratase from neurospora crassa forms a helix which has a dimeric repeat.the fungal cyanide hydratases form a functionally specialized subset of the nitrilases which catalyze the hydrolysis of cyanide to formamide with high specificity. these hold great promise for the bioremediation of cyanide wastes. the low resolution (3.0 nm) three-dimensional reconstruction of negatively stained recombinant cyanide hydratase fibers from the saprophytic fungus neurospora crassa by iterative helical real space reconstruction reveals that enzyme fibers display left-handed d(1) s(5. ...200918946669
atf-1 transcription factor regulates the expression of ccg-1 and cat-1 genes in response to fludioxonil under os-2 map kinase in neurospora crassa.the atf/creb family transcriptional factors are regulated by stress-activated map kinase in yeast. the disruptants of the atf-1 gene, which encodes an atf/creb family transcriptional factor, were isolated and characterized in neurospora crassa. the characteristic phenotypes in the os-2 map kinase strain, such as osmotic sensitivity and fludioxonil resistance, were not observed in the deltaatf-1 strain; however, like the os-2 strain, up-regulation of the catalase gene cat-1 and the clock-controll ...200818948219
identification of all fk506-binding proteins from neurospora crassa.immunophilins are intracellular receptors of immunosuppressive drugs, carrying peptidyl-prolyl cis-trans isomerase activity, with a general role in protein folding but also involved in specific regulatory mechanisms. four immunophilins of the fkbp-type (fk506-binding proteins) were identified in the genome of neurospora crassa. previously, fkbp22 has been located in the endoplasmic reticulum as part of chaperone/folding complexes and fkbp13 has been found to have a dual location in the cytoplasm ...200818948221
genome-wide high throughput analysis of dna methylation in eukaryotes.cytosine methylation is the quintessential epigenetic mark. two well-established methods, bisulfite sequencing and methyl-dna immunoprecipitation (medip) lend themselves to the genome-wide analysis of dna methylation by high throughput sequencing. here we provide an overview and brief review of these methods. we summarize our experience with medip followed by high throughput illumina/solexa sequencing, exemplified by the analysis of the methylated fraction of the neurospora crassa genome ("methy ...200918950712
stress-induced cell death is mediated by ceramide synthesis in neurospora crassa.the combined stresses of moderate heat shock (45 degrees c) and analog-induced glucose deprivation constitute a lethal stress for neurospora crassa. we found that this cell death requires fatty acid synthesis and the cofactor biotin. in the absence of the cofactor, the stressed cells are particularly sensitive to exogenous ceramide, which is lethal at low concentrations. when we extracted endogenous sphingolipids, we found that unique ceramides were induced (i) by the inhibitory glucose analog 2 ...200818952903
cell fusion in the filamentous fungus, neurospora crassa.hyphal fusion occurs at different stages in the vegetative and sexual life cycle of filamentous fungi. similar to cell fusion in other organisms, the process of hyphal fusion requires cell recognition, adhesion, and membrane merger. analysis of the hyphal fusion process in the model organism neurospora crassa using fluorescence and live cell imaging as well as cell and molecular biological techniques has begun to reveal its complex cellular regulation. several genes required for hyphal fusion ha ...200818979236
functional characterization of tzn1 and tzn2-zinc transporter genes in neurospora crassa.previous work from our laboratory involved the description of the neurospora metal transportome, which included seven hypothetical zinc transporters belonging to the zip family. the aim of the present study was to make a comparative functional evaluation of two hypothetical zinc transporters named tzn1 (ncu07621.3) and tzn2 (ncu11414.3). phenotypic analysis of tzn1 and tzn2 mutants and a double mutant (tzn1tzn2) revealed that the deletion of tzn1 causes aconidiation and a greater defect in growt ...200918989627
closing the circadian negative feedback loop: frq-dependent clearance of wc-1 from the nucleus.in neurospora crassa, a transcription factor, wcc, activates the transcription of frq. frq forms homodimers as well as complexes with an rna helicase, frh, and the wcc, and translocates into the nucleus to inactivate the wcc, closing the time-delayed negative feedback loop. the detailed mechanism for closing this loop, however, remains incompletely understood. in particular within the nucleus, the low amount of frq compared with that of wc-1 creates a conundrum: how can the nuclear frq inactivat ...200818997062
c(35)-apocarotenoids in the yellow mutant neurospora crassa ylo.the neurospora crassa mutant ylo exhibits a yellow phenotype instead of the red-orange pigmentation of the wild type. recently, it was shown that the mutant ylo is defective in a specific aldehyde dehydrogenase which catalyses the last step of carotenogenesis to the formation of neurosporaxanthin [estrada, a.f., youssar, l., scherzinger, d., al-babili, s., avalos, j., 2008. the ylo-1 gene encodes an aldehyde dehydrogenase responsible for the last reaction in the neurospora carotenoid pathway. mo ...200819007949
the group iii two-component histidine kinase of filamentous fungi is involved in the fungicidal activity of the bacterial polyketide ambruticin.we have shown that the plant pathogen alternaria brassicicola exhibited very high susceptibility to ambruticin vs4 and to a lesser extent to the phenylpyrrole fungicide fludioxonil. these compounds are both derived from natural bacterial metabolites with antifungal properties and are thought to exert their toxicity by interfering with osmoregulation in filamentous fungi. disruption of the osmosensor group iii histidine kinase gene abnik1 (for a. brassicola nik1) resulted in high levels of resist ...200919011080
the fungus neurospora crassa displays telomeric silencing mediated by multiple sirtuins and by methylation of histone h3 lysine 9.silencing of genes inserted near telomeres provides a model to investigate the function of heterochromatin. we initiated a study of telomeric silencing in neurospora crassa, a fungus that sports dna methylation, unlike most other organisms in which telomeric silencing has been characterized.200819014414
ripcal: a tool for alignment-based analysis of repeat-induced point mutations in fungal genomic sequences.repeat-induced point mutation (rip) is a fungal-specific genome defence mechanism that alters the sequences of repetitive dna, thereby inactivating coding genes. repeated dna sequences align between mating and meiosis and both sequences undergo c:g to t:a transitions. in most fungi these transitions preferentially affect cpa di-nucleotides thus altering the frequency of certain di-nucleotides in the affected sequences. the majority of previously published in silico analyses were limited to the c ...200819014496
the het-c heterokaryon incompatibility gene in aspergillus niger.heterokaryon incompatibility among aspergillus niger strains is a widespread phenomenon that is observed as the inability to form stable heterokaryons. the genetic basis of heterokaryon incompatibility reactions is well established in some sexual filamentous fungi but largely unknown in presumed asexual species, such as a. niger. to test whether the genes that determine heterokaryon incompatibility in neurospora crassa, such as het-c, vib-1 and pin-c, have a similar function in a. niger, we perf ...200919015029
a high-density single nucleotide polymorphism map for neurospora crassa.we report the discovery and validation of a set of single nucleotide polymorphisms (snps) between the reference neurospora crassa strain oak ridge and the mauriceville strain (fgsc 2555), of sufficient density to allow fine mapping of most loci. sequencing of mauriceville cdnas and alignment to the completed genomic sequence of the oak ridge strain identified 19,087 putative snps. of these, a subset was validated by cleaved amplified polymorphic sequence (caps), a simple and robust pcr-based ass ...200919015548
[cloning and bioinformatic analysis of calcineurin b gene in setosphaeria turcica].to clone and identify cnb gene from setosphaeria turcica and to perform the primary bioinformatic analysis.200819062640
the saccharomyces cerevisiae prm1 homolog in neurospora crassa is involved in vegetative and sexual cell fusion events but also has postfertilization functions.cell-cell fusion is essential for a variety of developmental steps in many eukaryotic organisms, during both fertilization and vegetative cell growth. although the molecular mechanisms associated with intracellular membrane fusion are well characterized, the molecular mechanisms of plasma membrane merger between cells are poorly understood. in the filamentous fungus neurospora crassa, cell fusion events occur during both vegetative and sexual stages of its life cycle, thus making it an attractiv ...200919064710
a splice variant of the neurospora crassa hex-1 transcript, which encodes the major protein of the woronin body, is modulated by extracellular phosphate and ph changes.the woronin body, a septal pore-associated organelle specific to filamentous ascomycetes, is crucial for preventing cytoplasmic bleeding after hyphal injury. in this study, we show that t1hex-1 transcript and a variant splicing t2hex-1 transcript are up-regulated at alkaline ph. we also show that both hex-1 transcripts are overexpressed in the preg(c), nuc-1(rip), and pacc(ko) mutant strains of neurospora crassa grown under conditions of phosphate shortage at alkaline ph, suggesting that hex-1 t ...200919071122
heterologous expression in escherichia coli of neurospora crassa neutral trehalase as an active enzyme.neutral trehalase from neurospora crassa was expressed in escherichia coli as a polypeptide of approximately 84 kda in agreement with the theoretical size calculated from the corresponding cdna. the recombinant neutral trehalase, purified by affinity chromatography exhibited a specific activity of 80-150 mu/mg protein. optima of ph and temperature were 7.0 and 30 degrees c, respectively. the enzyme was absolutely specific for trehalose, and was quite sensitive to incubation at 40 degrees c. the ...200919073263
the influence of sterols on the conformation of recombinant mitochondrial porin in detergent.mitochondrial porins (voltage-dependent anion-selective channels, vdac) are key contributors to cellular metabolism. when isolated from mitochondria porins copurify with sterols, and some isolated forms of the protein require sterol for insertion into artificial membranes. nonetheless, the contributions of sterols to the folded state of mitochondrial porin are not understood. recently, with the goal of high-resolution structural studies, several laboratories have developed methods for folding re ...200819088801
relics of repeat-induced point mutation direct heterochromatin formation in neurospora crassa.both rnai-dependent and -independent mechanisms have been implicated in the establishment of heterochromatin domains, which may be stabilized by feedback loops involving chromatin proteins and modifications of histones and dna. neurospora crassa sports features of heterochromatin found in higher eukaryotes, namely cytosine methylation (5mc), methylation of histone h3 lysine 9 (h3k9me), and heterochromatin protein 1 (hp1), and is a model to investigate heterochromatin establishment and maintenanc ...200919092133
characterization of chromosome ends in the filamentous fungus neurospora crassa.telomeres and subtelomere regions have vital roles in cellular homeostasis and can facilitate niche adaptation. however, information on telomere/subtelomere structure is still limited to a small number of organisms. prior to initiation of this project, the neurospora crassa genome assembly contained only 3 of the 14 telomeres. the missing telomeres were identified through bioinformatic mining of raw sequence data from the genome project and from clones in new cosmid and plasmid libraries. their ...200919104079
structure-function relationships in fungal large-subunit catalases.neurospora crassa has two large-subunit catalases, cat-1 and cat-3. cat-1 is associated with non-growing cells and accumulates particularly in asexual spores; cat-3 is associated with growing cells and is induced under different stress conditions. it is our interest to elucidate the structure-function relationships in large-subunit catalases. here we have determined the cat-3 crystal structure and compared it with the previously determined cat-1 structure. similar to cat-1, cat-3 hydrogen peroxi ...200919109972
telomere organization in the ligninolytic basidiomycete pleurotus ostreatus.telomeres are structural and functional chromosome regions that are essential for the cell cycle to proceed normally. they are, however, difficult to map genetically and to identify in genome-wide sequence programs because of their structure and repetitive nature. we studied the telomeric and subtelomeric organization in the basidiomycete pleurotus ostreatus using a combination of molecular and bioinformatics tools that permitted us to determine 19 out of the 22 telomeres expected in this fungus ...200919114509
characterization and mrna expression profile of the tbnre1 gene of the ectomycorrhizal fungus tuber borchii.this study focuses on the cloning and characterization of the major nitrogen regulator element from the ectomycorrhizal fungus tuber borchii, tbnre1. sequence analysis of the predicted protein and complementation experiments in neurospora crassa demonstrated that the cloned gene is orthologous to area/nit-2 gene. transcriptional expression investigations by real-time rt-pcr showed tbnre1 up-regulation in the presence of nitrate or in the absence of nitrogen during free-living mycelium growth. on ...200919115055
dimerization of the yeast eukaryotic translation initiation factor 5a requires hypusine and is rna dependent.post-translational modification of the highly conserved k51 residue of the saccharomyces cerevisiae eukaryotic translation initiation factor 5a (eif5a) to form hypusine, is essential for its many functions including the binding of specific mrnas. we characterized hypusinated yeast eif5a by size-exclusion chromatography and native page, showing that the protein exists as a homodimer. a k51r mutant, which was not functional in vivo eluted as a monomer and inhibition of hypusination abolished dimer ...200919120453
rhythmic conidiation in constant light in vivid mutants of neurospora crassa.in neurospora crassa, a circadian rhythm of conidiation (asexual spore formation) can be seen on the surface of agar media. this rhythm has a period of 22 hr in constant darkness (d/d). under constant illumination (l/l), no rhythm is visible and cultures show constant conidiation. however, here we report that strains with a mutation in the vivid (vvd) gene, previously shown to code for the photoreceptor involved in photo-adaptation, exhibit conidiation rhythms in l/l as well as in d/d. the perio ...200919139144
protein phosphatase pp1 is required for normal dna methylation in neurospora.covalent modifications of histones integrate intracellular and extracellular cues to regulate the genome. h3 lys 9 methylation (h3k9me) can direct heterochromatin formation and dna methylation, while phosphorylation of h3 ser 10 (h3s10p) drives gene activation and chromosome condensation. to examine the relationship between h3s10p, h3k9me, and dna methylation in neurospora crassa, we built and tested mutants of the putative h3s10 phosphatase, pp1. a pp1-impaired mutant showed increased h3s10p an ...200819141471
understanding the plasticity of the alpha/beta hydrolase fold: lid swapping on the candida antarctica lipase b results in chimeras with interesting biocatalytic properties.the best of both worlds. long molecular dynamics (md) simulations of candida antarctica lipase b (calb) confirmed the function of helix alpha5 as a lid structure. replacement of the helix with corresponding lid regions from calb homologues from neurospora crassa and gibberella zeae resulted in new calb chimeras with novel biocatalytic properties. the figure shows a snapshot from the md simulation. the candida antarctica lipase b (calb) has found very extensive use in biocatalysis reactions. long ...200919156649
tools for fungal proteomics: multifunctional neurospora vectors for gene replacement, protein expression and protein purification.the completion of genome-sequencing projects for a number of fungi set the stage for detailed investigations of proteins. we report the generation of versatile expression vectors for detection and isolation of proteins and protein complexes in the filamentous fungus neurospora crassa. the vectors, which can be adapted for other fungi, contain c- or n-terminal flag, ha, myc, gfp, or hat-flag epitope tags with a flexible poly-glycine linker and include sequences for targeting to the his-3 locus in ...200919171944
a sorghum 85kda heat stress-modulated protein shows calmodulin-binding properties and cross-reactivity to anti-neurospora crassa hsp 80 antibodies.the present study, carried out to identify stress-modulated calmodulin (cam)-binding proteins in sorghum, resulted in the isolation of several proteins, which showed binding to cam-sepharose matrix. calmodulin gel overlay assay and maldi-tof ms analysis revealed that an 85kda protein (hsp85), which interacted with calmodulin, cross-reacted with anti-n. crassa hsp80 antibodies. since these antibodies bind to plant hsp90, sorghum hsp85 is likely to be a member of the hsp90 family. this study provi ...200919174162
inflated kinetic isotope effects in the branched mechanism of neurospora crassa 2-nitropropane dioxygenase.catalytic turnover of neurospora crassa 2-nitropropane dioxygenase with nitroethane as substrate occurs through both nonoxidative and oxidative pathways. the ph dependence of the kinetic isotope effects with [1,1-(2)h(2)]nitroethane as substrate was measured in the current study by monitoring the formation of the nitronate product in the nonoxidative pathway. the kinetic isotope effect on the second-order rate constant for nitronate formation, k(cat)/k(m), decreased from an upper limiting value ...200919199786
chromosome segment duplications in neurospora crassa: barren crosses beget fertile science.studies on neurospora chromosome segment duplications (dps) performed since the publication of perkins's comprehensive review in 1997 form the focus of this article. we present a brief summary of perkins's seminal work on chromosome rearrangements, specifically, the identification of insertional and quasiterminal translocations that can segregate dp progeny when crossed with normal sequence strains (i.e., t x n). we describe the genome defense process called meiotic silencing by unpaired dna tha ...200919204993
transcriptional changes in the nuc-2a mutant strain of neurospora crassa cultivated under conditions of phosphate shortage.the molecular mechanism that controls the response to phosphate shortage in neurospora crassa involves four regulatory genes -nuc-2, preg, pgov, and nuc-1. phosphate shortage is sensed by the nuc-2 gene, the product of which inhibits the functioning of the preg-pgov complex. this allows the translocation of the transcriptional factor nuc-1 into the nucleus, which activates the transcription of phosphate-repressible phosphatases. the nuc-2a mutant strain of n. crassa carries a loss-of-function mu ...200919230635
catabolite repression directly affects transcription of the qa-y gene of neurospora crassa.the quinic acid (qa) gene cluster of neurospora crassa is subject to two levels of gene control: a primary system which responds to the presence of quinic acid via the qa-1s repressor protein, and a secondary system which represses transcription of qa genes in the presence of a preferred carbon source, e.g. glucose. rna blot analysis revealed that transcription of the qa-y gene, which encodes a quinic acid permease, was strongly repressed in the presence of glucose even in the absence of the qa- ...200919236936
the bem46-like protein appears to be essential for hyphal development upon ascospore germination in neurospora crassa and is targeted to the endoplasmic reticulum.the bud emergence (bem)46 proteins are evolutionarily conserved members of the alpha/beta-hydrolase super family, but their exact role remains unknown. to better understand the cellular role of bem46 and its homologs, we used the model organism neurospora crassa in conjunction with bem46 rnai, over-expression vectors, and repeat induced point mutation analyzes. we clearly demonstrated that bem46 is required for cell type-specific hyphal growth, which indicates a role for bem46 in maintaining pol ...200919238386
effects of mitochondrial complex iii disruption in the respiratory chain of neurospora crassa.in mitochondria from most organisms, including neurospora crassa, dimeric complex iii was found associated with complex i. additional association of complex iv with this core structure leads to the formation of a respirasome. it was recently described for bacteria and mammals that complex iii is needed for the assembly/stability of complex i. to elucidate the role of complex iii in the organization of the respiratory chain of n. crassa, we analysed strains devoid of either the rieske iron-sulphu ...200919239619
quelling targets the rdna locus and functions in rdna copy number control.rna silencing occurs in a broad range of organisms. although its ancestral function is probably related to the genome defense mechanism against repetitive selfish elements, it has been found that rna silencing regulates different cellular processes such as gene expression and chromosomal segregation. in neurospora crassa, a rna silencing mechanism, called quelling, acts to repress the expression of transgenes and transposons, but until now no other cellular functions have been shown to be regula ...200919243581
transient responses during hyperosmotic shock in the filamentous fungus neurospora crassa.fungal cells maintain an internal hydrostatic pressure (turgor) of about 400-500 kpa. in the filamentous fungus neurospora crassa, the initial cellular responses to hyperosmotic treatment are loss of turgor, a decrease in relative hyphal volume per unit length (within 1 min) and cell growth arrest; all recover over a period of 10-60 min due to increased net ion uptake and glycerol production. the electrical responses to hyperosmotic treatment are a transient depolarization of the potential (with ...200919246761
genome-wide analysis of light-inducible responses reveals hierarchical light signalling in neurospora.white collar-1 (wc-1) and white collar-2 (wc-2) are essential for light-mediated responses in neurospora crassa, but the molecular mechanisms underlying gene induction and the roles of other real and putative photoreceptors remain poorly characterized. unsupervised hierarchical clustering of genome-wide microarrays reveals 5.6% of detectable transcripts, including several novel mediators, that are either early or late light responsive. evidence is shown for photoreception in the absence of the d ...200919262566
a novel polyketide biosynthesis gene cluster is involved in fruiting body morphogenesis in the filamentous fungi sordaria macrospora and neurospora crassa.during fungal fruiting body development, hyphae aggregate to form multicellular structures that protect and disperse the sexual spores. analysis of microarray data revealed a gene cluster strongly upregulated during fruiting body development in the ascomycete sordaria macrospora. real time pcr analysis showed that the genes from the orthologous cluster in neurospora crassa are also upregulated during development. the cluster encodes putative polyketide biosynthesis enzymes, including a reducing ...200919277664
carotenoids and carotenogenic genes in podospora anserina: engineering of the carotenoid composition extends the life span of the mycelium.carotenoids have been identified in the fungus podospora anserina and a parallel pathway to neurosporene and beta-carotene was established. three genes for the beta-carotene branch have been cloned and their function elucidated. they correspond to the al-1, al-2 and al-3 genes from neurospora crassa. they were individually and in combinations over-expressed in p. anserina in order to modify the carotenoid composition qualitatively and quantitatively. in the resulting transformants, carotenoid sy ...200919277665
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