Simple screening protocol for identification of potential mycoremediation tools for the elimination of polycyclic aromatic hydrocarbons and phenols from hyperalkalophile industrial effluents.

A number of fungal strains belonging to the ascomycota, basidiomycota and zygomycota genera were subjected to an in vitro screening regime to assess their ligninolytic activity potential, with a view to their potential use in mycoremediation-based strategies to remove phenolic compounds and polycyclic aromatic hydrocarbons (PAHs) from industrial wastewaters. All six basidiomycetes completely decolorized remazol brilliant blue R (RBBR), while also testing positive in both the guaiacol and gallic acid tests indicating good levels of lignolytic activity. All the fungi were capable of tolerating phenanthrene, benzo-α- pyrene, phenol and p-chlorophenol in agar medium at levels of 10 ppm. Six of the fungal strains, Pseudogymnoascus sp., Aspergillus caesiellus, Trametes hirsuta IBB 450, Phanerochate chrysosporium ATCC 787, Pleurotus ostreatus MTCC 1804 and Cadophora sp. produced both laccase and Mn peroxidase activity in the ranges of 200-560 U/L and 6-152 U/L, respectively, in liquid media under nitrogen limiting conditions. The levels of adsorption of the phenolic and PAHs were negligible with 99% biodegradation being observed in the case of benzo-α-pyrene, phenol and p-chlorophenol. The aforementioned six fungal strains were also found to be able to effectively treat highly alkaline industrial wastewater (pH 12.4). When this wastewater was supplemented with 0.1 mM glucose, all of the tested fungi, apart from A. caesiellus, displayed the capacity to remove both the phenolic and PAH compounds. Based on their biodegradative capacity we found T. hirsuta IBB 450 and Pseudogymnoascus sp., to have the greatest potential for further use in mycoremediation based strategies to treat wastestreams containing phenolics and PAHs.

Exophiala chapopotensis sp. nov., an extremotolerant black yeast from an oil-polluted soil in Mexico; phylophenetic approach to species hypothesis in the Herpotrichiellaceae family.

Exophiala is a black fungi of the family Herpotrichiellaceae that can be found in a wide range of environments like soil, water and the human body as potential opportunistic pathogen. Some species are known to be extremophiles, thriving in harsh conditions such as deserts, glaciers, and polluted habitats. The identification of novel Exophiala species across diverse environments underlines the remarkable biodiversity within the genus. However, its classification using traditional phenotypic and phylogenetic analyses has posed a challenges. Here we describe a novel taxon, Exophiala chapopotensis sp. nov., strain LBMH1013, isolated from oil-polluted soil in Mexico, delimited according to combined morphological, molecular, evolutionary and statistics criteria. This species possesses the characteristic dark mycelia growing on PDA and tends to be darker in the presence of hydrocarbons. Its growth is dual with both yeast-like and hyphal forms. LBMH1013 differs from closely related species such as E. nidicola due to its larger aseptate conidia and could be distinguished from E. dermatitidis and E. heteromorpha by its inability to thrive above 37°C or 10% of NaCl. A comprehensive genomic analyses using up-to-date overall genome relatedness indices, several multigene phylogenies and molecular evolutionary analyzes using Bayesian speciation models, further validate its species-specific transition from all current Exophiala/Capronia species. Additionally, we applied the phylophenetic conceptual framework to delineate the species-specific hypothesis in order to incorporate this proposal within an integrative taxonomic framework. We believe that this approach to delimit fungal species will also be useful to our peers.

Extremely chaotolerant and kosmotolerant Aspergillus atacamensis - a metabolically versatile fungus suitable for recalcitrant biosolid treatment.

Obligate halophily is extremely rare in fungi. Nevertheless, Aspergillus atacamensis (strain EXF-6660), isolated from a salt water-exposed cave in the Coastal Range hills of the hyperarid Atacama Desert in Chile, is an obligate halophile, with a broad optimum range from 1.5 to 3.4 M of NaCl. When we tested its ability to grow at varied concentrations of both kosmotropic (NaCl, KCl, and sorbitol) and chaotropic (MgCl2, LiCl, CaCl2, and glycerol) solutes, stereoscopy and laser scanning microscopy revealed the formation of phialides and conidia. A. atacamensis EXF-6660 grew up to saturating levels of NaCl and at 2.0 M concentration of the chaotropic salt MgCl2. Our findings confirmed that A. atacamensis is an obligate halophile that can grow at substantially higher MgCl2 concentrations than 1.26 M, previously considered as the maximum limit supporting prokaryotic life. To assess the fungus' metabolic versatility, we used the phenotype microarray technology Biolog FF MicroPlates. In the presence of 2.0 M NaCl concentration, strain EXF-6660 metabolism was highly versatile. A vast repertoire of organic molecules (~95% of the substrates present in Biolog FF MicroPlates) was metabolized when supplied as sole carbon sources, including numerous polycyclic aromatic hydrocarbons, benzene derivatives, dyes, and several carbohydrates. Finally, the biotechnological potential of A. atacamensis for xenobiotic degradation and biosolid treatment was investigated. Interestingly, it could remove biphenyls, diphenyl ethers, different pharmaceuticals, phenols, and polyaromatic hydrocarbons. Our combined findings show that A. atacamensis EXF-6660 is a highly chaotolerant, kosmotolerant, and xerotolerant fungus, potentially useful for xenobiotic and biosolid treatments.

Surviving in the Brine: A Multi-Omics Approach for Understanding the Physiology of the Halophile Fungus Aspergillus sydowii at Saturated NaCl Concentration.

Although various studies have investigated osmoadaptations of halophilic fungi to saline conditions, only few analyzed the fungal mechanisms occurring at saturated NaCl concentrations. Halophilic Aspergillus sydowii is a model organism for the study of molecular adaptations of filamentous fungi to hyperosmolarity. For the first time a multi-omics approach (i.e., transcriptomics and metabolomics) was used to compare A. sydowii at saturated concentration (5.13 M NaCl) to optimal salinity (1 M NaCl). Analysis revealed 1,842 genes differentially expressed of which 704 were overexpressed. Most differentially expressed genes were involved in metabolism and signal transduction. A gene ontology multi-scale network showed that ATP binding constituted the main network node with direct interactions to phosphorelay signal transduction, polysaccharide metabolism, and transferase activity. Free amino acids significantly decreased and amino acid metabolism was reprogrammed at 5.13 M NaCl. mRNA transcriptional analysis revealed upregulation of genes involved in methionine and cysteine biosynthesis at extreme water deprivation by NaCl. No modifications of membrane fatty acid composition occurred. Upregulated genes were involved in high-osmolarity glycerol signal transduction pathways, biosynthesis of ß-1,3-glucans, and cross-membrane ion transporters. Downregulated genes were related to the synthesis of chitin, mannose, cell wall proteins, starvation, pheromone synthesis, and cell cycle. Non-coding RNAs represented the 20% of the total transcripts with 7% classified as long non-coding RNAs (lncRNAs). The 42% and 69% of the total lncRNAs and RNAs encoding transcription factors, respectively, were differentially expressed. A network analysis showed that differentially expressed lncRNAs and RNAs coding transcriptional factors were mainly related to the regulation of metabolic processes, protein phosphorylation, protein kinase activity, and plasma membrane composition. Metabolomic analyses revealed more complex and unknown metabolites at saturated NaCl concentration than at optimal salinity. This study is the first attempt to unravel the molecular ecology of an ascomycetous fungus at extreme water deprivation by NaCl (5.13 M). This work also represents a pioneer study to investigate the importance of lncRNAs and transcriptional factors in the transcriptomic response to high NaCl stress in halophilic fungi.

Physcomitrium patens Infection by Colletotrichum gloeosporioides: Understanding the Fungal-Bryophyte Interaction by Microscopy, Phenomics and RNA Sequencing.

Anthracnose caused by the hemibiotroph fungus Colletotrichum gloeosporioides is a devastating plant disease with an extensive impact on plant productivity. The process of colonization and disease progression of C. gloeosporioides has been studied in a number of angiosperm crops. To better understand the evolution of the plant response to pathogens, the study of this complex interaction has been extended to bryophytes. The model moss Physcomitrium patens Hedw. B&S (former Physcomitrella patens) is sensitive to known bacterial and fungal phytopathogens, including C. gloeosporioides, which cause infection and cell death. P. patens responses to these microorganisms resemble that of the angiosperms. However, the molecular events during the interaction of P. patens and C. gloeosporioides have not been explored. In this work, we present a comprehensive approach using microscopy, phenomics and RNA-seq analysis to explore the defense response of P. patens to C. gloeosporioides. Microscopy analysis showed that appressoria are already formed at 24 h after inoculation (hai) and tissue colonization and cell death occur at 24 hai and is massive at 48 hai. Consequently, the phenomics analysis showed progressing browning of moss tissues and impaired photosynthesis from 24 to 48 hai. The transcriptomic analysis revealed that more than 1200 P. patens genes were differentially expressed in response to Colletotrichum infection. The analysis of differentially expressed gene function showed that the C. gloeosporioides infection led to a transcription reprogramming in P. patens that upregulated the genes related to pathogen recognition, secondary metabolism, cell wall reinforcement and regulation of gene expression. In accordance with the observed phenomics results, some photosynthesis and chloroplast-related genes were repressed, indicating that, under attack, P. patens changes its transcription from primary metabolism to defend itself from the pathogen.

Transcriptome-Based Identification of a Functional Fasciola hepatica Carboxylesterase B.

Bioinformatics analysis of the complete transcriptome of Fasciola hepatica, identified a total of ten putative carboxylesterase transcripts, including a 3146 bp mRNA transcript coding a 2205 bp open reading frame that translates into a protein of 735 amino acids, resulting in a predicted protein mass of 83.5 kDa and a putative carboxylesterase B enzyme. The gene coding for this enzyme was found in two reported F. hepatica complete genomes stretching 23,230 bp, containing two exons of 1282 and 1864 bp, respectively, as well as a 20,084 bp intron between the exons. The enzymatic activity was experimentally assayed on F. hepatica protein extracts by SDS-PAGE zymograms using synthetic chromogenic substrates, confirming both the theoretical molecular weight and carboxylesterase enzymatic activity. Further bioinformatics predicted that this enzyme is an integral component of the cellular membrane that should be active as a 167 kDa homodimer complex and polyacrylamide gel electrophoresis (PAGE) zymograms experiments confirmed the analysis. Additional bioinformatics analysis showed that DNA sequences that code for this particular enzyme are highly conserved in other parasitic trematodes, although they are labeled hypothetical proteins.

Osmolyte Signatures for the Protection of Aspergillus sydowii Cells under Halophilic Conditions and Osmotic Shock.

Aspergillus sydowii is a moderate halophile fungus extensively studied for its biotechnological potential and halophile responses, which has also been reported as a coral reef pathogen. In a recent publication, the transcriptomic analysis of this fungus, when growing on wheat straw, showed that genes related to cell wall modification and cation transporters were upregulated under hypersaline conditions but not under 0.5 M NaCl, the optimal salinity for growth in this strain. This led us to study osmolyte accumulation as a mechanism to withstand moderate salinity. In this work, we show that A. sydowii accumulates trehalose, arabitol, mannitol, and glycerol with different temporal dynamics, which depend on whether the fungus is exposed to hypo- or hyperosmotic stress. The transcripts coding for enzymes responsible for polyalcohol synthesis were regulated in a stress-dependent manner. Interestingly, A. sydowii contains three homologs (Hog1, Hog2 and MpkC) of the Hog1 MAPK, the master regulator of hyperosmotic stress response in S. cerevisiae and other fungi. We show a differential regulation of these MAPKs under different salinity conditions, including sustained basal Hog1/Hog2 phosphorylation levels in the absence of NaCl or in the presence of 2.0 M NaCl, in contrast to what is observed in S. cerevisiae. These findings indicate that halophilic fungi such as A. sydowii utilize different osmoadaptation mechanisms to hypersaline conditions.

Aromatic Hydrocarbon Removal by Novel Extremotolerant Exophiala and Rhodotorula Spp. from an Oil Polluted Site in Mexico.

Since Aromatic hydrocarbons are recalcitrant and toxic, strategies to remove them are needed. The aim of this work was to isolate fungi capable of using aromatic hydrocarbons as carbon sources. Two isolates from an oil polluted site in Mexico were identified through morphological and molecular markers as a novel Rhodotorula sp. and an Exophiala sp. Both strains were able to grow in a wide range of pH media, from 4 to 12, showing their optimal growth at alkaline pH's and are both halotolerant. The Exophiala strain switched from hyphae to yeast morphotype in high salinity conditions. To the best of our knowledge, this is the first report of salt triggering dimorphism. The Rhodotorula strain, which is likely a new undescribed species, was capable of removing singled ringed aromatic compounds such as benzene, xylene, and toluene, but could not remove benzo[a] pyrene nor phenanthrene. Nevertheless, these hydrocarbons did not impair its growth. The Exophiala strain showed a different removal capacity. It could remove the polyaromatic hydrocarbons but performed poorly at removing toluene and xylene. Nevertheless, it still could grow well in the presence of the aromatic compounds. These strains could have a potential for aromatic compounds removal.

Haloadaptative Responses of Aspergillus sydowii to Extreme Water Deprivation: Morphology, Compatible Solutes, and Oxidative Stress at NaCl Saturation.

Water activity (aw) is critical for microbial growth, as it is severely restricted at aw < 0.90. Saturating NaCl concentrations (~5.0 M) induce extreme water deprivation (aw ≅ 0.75) and cellular stress responses. Halophilic fungi have cellular adaptations that enable osmotic balance and ionic/oxidative stress prevention to grow at high salinity. Here we studied the morphology, osmolyte synthesis, and oxidative stress defenses of the halophile Aspergillus sydowii EXF-12860 at 1.0 M and 5.13 M NaCl. Colony growth, pigmentation, exudate, and spore production were inhibited at NaCl-saturated media. Additionally, hyphae showed unpolarized growth, lower diameter, and increased septation, multicellularity and branching compared to optimal NaCl concentration. Trehalose, mannitol, arabitol, erythritol, and glycerol were produced in the presence of both 1.0 M and 5.13 M NaCl. Exposing A. sydowii cells to 5.13 M NaCl resulted in oxidative stress evidenced by an increase in antioxidant enzymes and lipid peroxidation biomarkers. Also, genes involved in cellular antioxidant defense systems were upregulated. This is the most comprehensive study that investigates the micromorphology and the adaptative cellular response of different non-enzymatic and enzymatic oxidative stress biomarkers in halophilic filamentous fungi.

Stress Reshapes the Physiological Response of Halophile Fungi to Salinity.

(1) Background: Mechanisms of cellular and molecular adaptation of fungi to salinity have been commonly drawn from halotolerant strains and few studies in basidiomycete fungi. These studies have been conducted in settings where cells are subjected to stress, either hypo- or hyperosmotic, which can be a confounding factor in describing physiological mechanisms related to salinity. (2) Methods: We have studied transcriptomic changes in Aspergillussydowii, a halophilic species, when growing in three different salinity conditions (No NaCl, 0.5 M, and 2.0 M NaCl). (3) Results: In this fungus, major physiological modifications occur under high salinity (2.0 M NaCl) and not when cultured under optimal conditions (0.5 M NaCl), suggesting that most of the mechanisms described for halophilic growth are a consequence of saline stress response and not an adaptation to saline conditions. Cell wall modifications occur exclusively at extreme salinity, with an increase in cell wall thickness and lamellar structure, which seem to involve a decrease in chitin content and an augmented content of alfa and beta-glucans. Additionally, three hydrophobin genes were differentially expressed under hypo- or hyperosmotic stress but not when the fungus grows optimally. Regarding compatible solutes, glycerol is the main compound accumulated in salt stress conditions, whereas trehalose is accumulated in the absence of salt. (4) Conclusions: Physiological responses to salinity vary greatly between optimal and high salt concentrations and are not a simple graded effect as the salt concentration increases. Our results highlight the influence of stress in reshaping the response of extremophiles to environmental challenges.

Characterization of Fungal Endophytes Isolated from the Metal Hyperaccumulator Plant Vachellia farnesiana Growing in Mine Tailings.

Heavy metal pollution has become an environmental and health problem worldwide. With the aim of finding novel strategies for metal bioremediation, endophytic fungi from the heavy metal hyperaccumulator plant Vachellia farnesiana were isolated and characterized. The plants were growing in mine tailings, rich in Zn, Pb, and Cu. Morphological and phylogenetic analyses indicated that the fungal strains belonged to Neocosmospora and Aspergillus genera. The Neocosmospora isolate belongs to the Fusarium solani species complex (FSSC) that groups phytopathogen species. However, in this case the plants from which it was isolated did not show any signs of disease. Both fungal strains were able to remove significant amounts of heavy metals from liquid cultures, either in a mixture of the three metals or each metal in a single culture. In response to lead exposure, the Neocosmospora sp. strain secreted specific novel phenolic compounds other than anthraquinones or naphtoquinones, which have been described in similar situations. The Aspergillus sp. dropped the pH in the medium. High-performance liquid chromatography determinations indicated that this strain secreted mainly glutamic acid in response to lead, a novel mechanism, which has not been reported elsewhere. Malic and succinic acids were also produced in response to lead exposure. Possibly, glutamic and succinic acids (synthesized in the Krebs cycle) can be used to cope with metal toxicity due to the plant providing photosynthates to the fungus. These fungi showed the potential to be used for bioremediation or restoration of metal-polluted environments.

Metagenomics of Atacama Lithobiontic Extremophile Life Unveils Highlights on Fungal Communities, Biogeochemical Cycles and Carbohydrate-Active Enzymes.

Halites, which are typically found in various Atacama locations, are evaporitic rocks that are considered as micro-scaled salterns. Both structural and functional metagenomic analyses of halite nodules were performed. Structural analyses indicated that the halite microbiota is mainly composed of NaCl-adapted microorganisms. In addition, halites appear to harbor a limited diversity of fungal families together with a biodiverse collection of protozoa. Functional analysis indicated that the halite microbiome possesses the capacity to make an extensive contribution to carbon, nitrogen, and sulfur cycles, but possess a limited capacity to fix nitrogen. The halite metagenome also contains a vast repertory of carbohydrate active enzymes (CAZY) with glycosyl transferases being the most abundant class present, followed by glycosyl hydrolases (GH). Amylases were also present in high abundance, with GH also being identified. Thus, the halite microbiota is a potential useful source of novel enzymes that could have biotechnological applicability. This is the first metagenomic report of fungi and protozoa as endolithobionts of halite nodules, as well as the first attempt to describe the repertoire of CAZY in this community. In addition, we present a comprehensive functional metagenomic analysis of the metabolic capacities of the halite microbiota, providing evidence for the first time on the sulfur cycle in Atacama halites.

A Review on Viral Metagenomics in Extreme Environments.

Viruses are the most abundant biological entities in the biosphere, and have the ability to infect Bacteria, Archaea, and Eukaryotes. The virome is estimated to be at least ten times more abundant than the microbiome with 107 viruses per milliliter and 109 viral particles per gram in marine waters and sediments or soils, respectively. Viruses represent a largely unexplored genetic diversity, having an important role in the genomic plasticity of their hosts. Moreover, they also play a significant role in the dynamics of microbial populations. In recent years, metagenomic approaches have gained increasing popularity in the study of environmental viromes, offering the possibility of extending our knowledge related to both virus diversity and their functional characterization. Extreme environments represent an interesting source of both microbiota and their virome due to their particular physicochemical conditions, such as very high or very low temperatures and >1 atm hydrostatic pressures, among others. Despite the fact that some progress has been made in our understanding of the ecology of the microbiota in these habitats, few metagenomic studies have described the viromes present in extreme ecosystems. Thus, limited advances have been made in our understanding of the virus community structure in extremophilic ecosystems, as well as in their biotechnological potential. In this review, we critically analyze recent progress in metagenomic based approaches to explore the viromes in extreme environments and we discuss the potential for new discoveries, as well as methodological challenges and perspectives.

First demonstration that ascomycetous halophilic fungi (Aspergillus sydowii and Aspergillus destruens) are useful in xenobiotic mycoremediation under high salinity conditions.

Polycyclic aromatic hydrocarbons (PAH) and pharmaceutical compounds (PhC) are xenobiotics present in many saline wastewaters. Although fungi are known for their ability to remove xenobiotics, the potential of halophilic fungi to degrade highly persistent pollutants was not yet investigated. The use of two halophilic fungi, Aspergillus sydowii and Aspergillus destruens, for the elimination of PAH and PhC at saline conditions was studied. In saline synthetic medium both fungi used benzo-α-pyrene and phenanthrene as sole carbon source and removed over 90% of both PAH, A. sydowii due to biodegradation and A. destruens to bioadsorption. They removed 100% of a mixture of fifteen PAH in saline biorefinery wastewater. Test using Cucumis sativus demonstrated that wastewater treated with the two fungi lowered considerably the phytotoxicity. This study is the first demonstration that ascomycetous halophilic fungi, in contrast to other fungi (and in particular basidiomycetes) can be used for mycotreatments under salinity conditions.

Isolation and characterization of endophytes from nodules of Mimosa pudica with biotechnological potential.

Legumes establish symbiotic relationships with different microorganisms, which could function as plant growth promotion microorganisms (PGPM). The finding of new PGPM strains is important to increase plant production avoiding or diminishing the use of industrial fertilizers. Thus, in this work we evaluated the plant growth promotion traits of ten strains isolated from Mimosa pudica root nodules. According to the 16S rDNA sequence, the microorganisms were identified as Enterobacter sp. and Serratia sp. To the best of our knowledge this is the first report describing and endophytic interaction between Mimosa pudica and Enterobacter sp. These strains have some plant growth promoting traits such as phosphate solubilization, auxin production and cellulase and chitinase activity. Strains identified as Serratia sp. inhibited the growth of the phytopathogenic fungi Fusarium sp., and Alternaria solani and the oomycete Phytophthora capsici. According to their biochemical characteristics, three strains were selected to test their plant growth promoting activity in a medium with an insoluble phosphate source. These bacteria show low specificity for their hosts as endophytes, since they were able to colonize two very different legumes: Phaseolus vulgaris and M. pudica. Seedlings of P. vulgaris were inoculated and grown for fifteen days. Enterobacter sp. NOD1 and NOD10, promoted growth as reflected by an increase in shoot height as well as an increase in the size and emergence of the first two trifolia. We could localize NOD5 as an endophyte in roots in P. vulgaris by transforming the strain with a Green Fluorescent Protein carrying plasmid. Experiments of co-inoculation with different Rhizobium etli strains allowed us to discard that NOD5 can fix nitrogen in the nodules formed by a R. etli Fix- strain. The isolates described in this work show biotechnological potential for plant growth promoting activity and production of indoleacetic acid and siderophores.

Understanding the importance of mRNA transport in memory.

RNA localization is an important mechanism to sort proteins to specific subcellular domains. In neurons, several mRNAs are localized in dendrites and their presence allows autonomous control of local translation in response to stimulation of specific synapses. Active constitutive and activity-induced mechanisms of mRNA transport have been described that represent critical steps in the establishment and maintenance of synaptic plasticity. In recent years, the molecular composition of different transporting units has been reported and the identification of proteins and mRNAs in these RNA granules contributes to our understanding of the key steps that regulate mRNA transport and translation. Although RNA granules are heterogeneous, several proteins are common to different RNA granule populations, suggesting that they play important roles in the formation of the granules and/or their regulation during transport and translation. About 1-4% of the neuron transcriptome is found in RNA granules and the characterization of bound mRNAs reveal that they encode proteins of the cytoskeleton, the translation machinery, vesicle trafficking, and/or proteins involved in synaptic plasticity. Non-coding RNAs and microRNAs are also found in dendrites and likely regulate RNA translation. These mechanisms of mRNA transport and local translation are critical for synaptic plasticity mediated by activity or experience and memory.

The first description of a hormone-sensitive lipase from a basidiomycete: Structural insights and biochemical characterization revealed Bjerkandera adusta BaEstB as a novel esterase.

The heterologous expression and characterization of a Hormone-Sensitive Lipases (HSL) esterase (BaEstB) from the Basidiomycete fungus Bjerkandera adusta is reported for the first time. According to structural analysis, amino acid similarities and conservation of particular motifs, it was established that this enzyme belongs to the (HSL) family. The cDNA sequence consisted of 969 nucleotides, while the gene comprised 1133, including three introns of 57, 50, and 57 nucleotides. Through three-dimensional modeling and phylogenetic analysis, we conclude that BaEstB is an ortholog of the previously described RmEstB-HSL from the phylogenetically distant fungus Rhizomucor miehei. The purified BaEstB was characterized in terms of its specificity for the hydrolysis of different acyl substrates confirming its low lipolytic activity and a noticeable esterase activity. The biochemical characterization of BaEstB, the DLS analysis and the kinetic parameters determination revealed this enzyme as a true esterase, preferentially found in a dimeric state, displaying activity under alkaline conditions and relative low temperature (pH = 10, 20°C). Our data suggest that BaEstB is more active on substrates with short acyl chains and bulky aromatic moieties. Phylogenetic data allow us to suggest that a number of fungal hypothetical proteins could belong to the HSL family.

Characterization of lignocellulolytic activities from fungi isolated from the deep-sea sponge Stelletta normani.

Extreme habitats have usually been regarded as a source of microorganisms that possess robust proteins that help enable them to survive in such harsh conditions. The deep sea can be considered an extreme habitat due to low temperatures (<5°C) and high pressure, however marine sponges survive in these habitats. While bacteria derived from deep-sea marine sponges have been studied, much less information is available on fungal biodiversity associated with these sponges. Following screening of fourteen fungi isolated from the deep-sea sponge Stelletta normani sampled at a depth of 751 metres, three halotolerant strains (TS2, TS11 and TS12) were identified which displayed high CMCase and xylanase activities. Molecular based taxonomic approaches identified these strains as Cadophora sp. TS2, Emericellopsis sp. TS11 and Pseudogymnoascus sp. TS 12. These three fungi displayed psychrotolerance and halotolerant growth on CMC and xylan as sole carbon sources, with optimal growth rates at 20°C. They produced CMCase and xylanase activities, which displayed optimal temperature and pH values of between 50-70°C and pH 5-8 respectively, together with good thermostability and halotolerance. In solid-state fermentations TS2, TS11 and TS12 produced CMCases, xylanases and peroxidase/phenol oxidases when grown on corn stover and wheat straw. This is the first time that CMCase, xylanase and peroxidase/phenol oxidase activities have been reported in these three fungal genera isolated from a marine sponge. Given the biochemical characteristics of these ligninolytic enzymes it is likely that they may prove useful in future biomass conversion strategies involving lignocellulosic materials.

A novel expansin protein from the white-rot fungus Schizophyllum commune.

A novel expansin protein (ScExlx1) was found, cloned and expressed from the Basidiomycete fungus Schizophylum commune. This protein showed the canonical features of plant expansins. ScExlx1 showed the ability to form "bubbles" in cotton fibers, reduce the size of avicel particles and enhance reducing sugar liberation from cotton fibers pretreated with the protein and then treated with cellulases. ScExlx1 was able to bind cellulose, birchwood xylan and chitin and this property was not affected by different sodium chloride concentrations. A novel property of ScExlx1 is its capacity to enhance reducing sugars (N-acetyl glucosamine) liberation from pretreated chitin and further added with chitinase, which has not been reported for any expansin or expansin-like protein. To the best of our knowledge, this is the first report of a bona fide fungal expansin found in a basidiomycete and we could express the bioactive protein in Pichia pastoris.

The nuclear receptor NR4A1 induces a form of cell death dependent on autophagy in mammalian cells.

The control of cell death is a biological process essential for proper development, and for preventing devastating pathologies like cancer and neurodegeneration. On the other hand, autophagy regulation is essential for protein and organelle degradation, and its dysfunction is associated with overlapping pathologies like cancer and neurodegeneration, but also for microbial infection and aging. In the present report we show that two evolutionarily unrelated receptors--Neurokinin 1 Receptor (NK(1)R,) a G-protein coupled receptor, and Insulin-like Growth Factor 1 Receptor (IGF1R), a tyrosine kinase receptor--both induce non-apoptotic cell death with autophagic features and requiring the activity of the autophagic core machinery proteins PI3K-III, Beclin-1 and Atg7. Remarkably, this form of cell death occurs in apoptosis-competent cells. The signal transduction pathways engaged by these receptors both converged on the activation of the nuclear receptor NR4A1, which has previously been shown to play a critical role in some paradigms of apoptosis and in NK(1)R-induced cell death. The activity of NR4A1 was necessary for IGF1R-induced cell death, as well as for a canonical model of cell death by autophagy induced by the presence of a pan-caspase inhibitor, suggesting that NR4A1 is a general modulator of this kind of cell death. During cell death by autophagy, NR4A1 was transcriptionally competent, even though a fraction of it was present in the cytoplasm. Interestingly, NR4A1 interacts with the tumor suppressor p53 but not with Beclin-1 complex. Therefore the mechanism to promote cell death by autophagy might involve regulation of gene expression, as well as protein interactions. Understanding the molecular basis of autophagy and cell death mediation by NR4A1, should provide novel insights and targets for therapeutic intervention.