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October 1982 - October 1983
Education
September 1977 - June 1982
September 1974 - June 1977
Publications
Publications (74)
Functional symbiosis with fungal endophytes can help plants adapt to environmental stress. Diaporthe atlantica is one of the most abundant fungal taxa associated with roots of Festuca rubra subsp. pruinosa, a grass growing in sea cliffs. This study aimed to investigate the ability of a strain of this fungus to ameliorate the impact of drought stres...
Copper ion homeostasis involves a finely tuned and complex multi-level response system. This study expands on various aspects of the system in the model filamentous fungus Aspergillus nidulans. An RNA-seq screen in standard growth and copper toxicity conditions revealed expression changes in key copper response elements, providing an insight into t...
Tolerance of microorganisms to abiotic stress is enabled by regulatory mechanisms that coordinate the expression and activity of resistance genes. Alkalinity and high salt concentrations are major environmental physicochemical stresses. Here, we analyzed the roles of sodium-extrusion family (ENA) transporters EnaA, EnaB and EnaC in the response to...
Copper is a metal ion that is required as a micronutrient for growth and proliferation. However, copper accumulation generates toxicity by multiple mechanisms, potentially leading to cell death. Due to its toxic nature at high concentrations, different chemical variants of copper have been extensively used as antifungal agents in agriculture and me...
FluG is a long recognized early regulator of asexual development in Aspergillus nidulans. fluG null mutants show profuse aerial growth and no conidial production. Initial studies reported sequence homology of FluG with a prokaryotic type I glutamine synthetase, but catalytic activity has not been demonstrated. In this study, we conducted an in-dept...
Microbial cells interact with the environment by adapting to external changes. Signal transduction pathways participate in both sensing and responding in the form of modification of gene expression patterns, enabling cell survival. The filamentous fungal-specific SltA pathway regulates tolerance to alkalinity, elevated cation concentrations and, as...
Copper homeostasis has been extensively studied in mammals, bacteria, and yeast, but it has not been well-documented in filamentous fungi. In this report, we investigated the basis of copper tolerance in the model fungus Aspergillus nidulans. Three genes involved in copper homeostasis have been characterized. First, crpA the A. nidulans ortholog of...
crpA rescue phenotype. WT, ΔcrpA, and crpA+ complemented mutant strain characterization in solid medium supplemented with the indicated CuSO4 concentrations.
Copper homeostasis has been extensively studied in mammals, bacteria, and yeast, but it has not been well-documented in filamentous fungi. In this report, we investigated the basis of copper tolerance in the model fungus Aspergillus nidulans. Three genes involved in copper homeostasis have been characterized. First, crpA the A. nidulans ortholog of...
The mycelium is a colony form which combines the ability to adapt its morphology to the underlying substrate and conditions while maintaining functional unity through interconnectedness. Thus, regions that access nutrient-rich substrates may support other’s undertaking exploration or differentiation processes.
This display of versatility has led to...
In Aspergillus nidulans, asexual differentiation requires the presence of the transcription factor FlbB at the cell tip and apical nuclei. Understanding the relationship between these two pools is crucial for elucidating the biochemical processes mediating conidia production. Tip-to-nucleus communication was demonstrated by photo-convertible FlbB::...
In the model fungus Aspergillus nidulans, asexual development is induced from vegetative hyphae by a set of early regulators including the bZIP-type transcription factor FlbB. To determine the range of genes under the influence of the transcriptional activity of FlbB and to characterize their role in fungal development, we sequenced and compared th...
The mycelium is an organised cellular network that develops according to a functionally coherent plan. As it expands, the mycelium is capable of modulating the relative abundance of different cell types to suit the prevailing environmental conditions. This versatile pattern of multicellular development involves sophisticated environmental sensing a...
Morphogenesis encompasses programmed changes in gene expression that lead to the development of specialized cell types. In the model fungus Aspergillus nidulans, asexual development involves the formation of characteristic cell types, collectively known as the conidiophore. With the aim of determining the transcriptional changes that occur upon ind...
Aspergillus nidulans asexual differentiation is induced by Upstream Developmental Activators (UDAs) that include the bZIP-type Transcription Factor (TF) FlbB. A 2D-PAGE/MS-MS-coupled screen for proteins differentially expressed in the presence and absence of FlbB identified 18 candidates. Most candidates belong to GO term classes involved in osmoti...
Oligonucleotides used in this study.
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Characterization of the GmcA sequence and ortholog search.
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Genes identified by mass spectroscopy.
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Compositional characteristics of GMCs and 27 putative Ascomycete GmcA orthologs with BLAST E = 0.
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Germination of Aspergillus nidulans conidia in liquid cultures was progressively inhibited at inoculum loads above 1×10(5)conidiamL(-1). High conidial densities also inhibited growth of neighbouring mycelia. The eight-carbon oxylipin 1-octen-3-ol was identified as the main inhibitor in a fraction also containing 3-octanone and 3-octanol. These thre...
Alignment of the 40 Pezizomycotina FlbB orthologs used in this study. Anidu FlbB is in bold. Motifs B1, B2, B3 and B4 are labeled and highlighted in green or yellow in the nine sequences that were used to generate the motifs. The five signature residues of the bZip DNA binding domain are highlighted in purple. The first four heptads of the bZip dim...
Alignment of the 40 Pezizomycotina FlbE orthologs used in this study. Anidu FlbE is in bold. Conserved motifs E1, E2, E3, E4 and E5 are labeled and highlighted in green or yellow. The acidic region is highlighted in purple. Residues flanking intron locations are in bold italic.
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Aspergillus nidulans is a filamentous fungus widely used as a model for biotechnological and clinical research. It is also used as a platform for the study of basic eukaryotic developmental processes. Previous studies identified and partially characterized a set of proteins controlling cellular transformations in this ascomycete. Among these protei...
Development in the ascomycete A. nidulans is principally determined by environmental signals. Adaptability to oxidative stimuli can derive in changes of growth patterns and /or the activation of sexual or asexual reproductive cycles but this model fungus might also respond to high osmotic or salt concentrations, the redox state, the availability an...
Asexual development in Aspergillus nidulans begins in superficial hyphae as the programmed emergence of successive pseudohyphal modules, collectively known as the conidiophore, and is completed by a layer of specialized cells (phialides) giving rise to chains of aerial spores. A discrete number of regulatory factors present in hyphae play different...
Asexual development (conidiation) in Aspergillus is governed by multiple regulators. Here, we characterize the upstream developmental activator FlbC in Aspergillus nidulans. flbC mRNA is detectable throughout the life cycle, at relatively high levels during vegetative growth, early asexual and late sexual developmental phases. The deletion of flbC...
Fungi are capable of generating diverse cell types through developmental processes that stem from hyphae, acting as pluripotent cells. The formation of mitospores on emergence of hyphae to the air involves the participation of transcription factors, which co-ordinate the genesis of new cell types, eventually leading to spore formation.
In this inve...
Asexual development in the filamentous fungus Aspergillus nidulans is governed by the timely expression and cellular localization of multiple transcription factors. Hence, factors mediating import and export across the nuclear pore complexes (karyopherins) are expected to play a key role in coordinating the developmental programme. Here we characte...
Fungal colonies expand through apically extending peripheral hyphae. These long and relatively unbranched leaders leave behind cells that show a higher degree of branching and lower growth rates. Towards the centre of the colony, quiescent, but viable cells sustain processes such as transport of water, solutes and organelles, and support the concer...
Conidiophore formation in Aspergillus nidulans involves a developmental programme in which vegetative hyphae give rise to an ordered succession of differentiated cells: foot cell, stalk, vesicle, metulae, phialides and conidia. The developmental transition requires factors that are expressed in vegetative hyphae that activate the expression of the...
Aspergillus nidulans switches from vegetative growth to conidiation when aerial hyphae make contact with the atmosphere, or are subjected to specific environmental stress. The activation of the central conidiation pathway led by the transcription factor brlA is a critical milestone in this morphogenetic transition. A number of upstream developmenta...
In digital control, discrete-time signal reconstruction is usualy carried out by the zero-order hold (ZOH), although other options exist which exhibit interesting properties. A remarkable alternative is the fractional-order hold (FROH), which provokes no intersample ripple under steady state and, if properly tuned, can often place the discretized p...
The fungal colony is a complex multicellular unit consisting of various cell types and functions. Asexual spore formation
(conidiation) is integrated through sensory and regulatory elements into the general morphogenetic plan, in which the activation
of the transcription factor BrlA is the first determining step. A number of early regulatory elemen...
In digital control, discrete-time signal reconstruction is usualy carried out by the zero-order hold (ZOH), although other options exist which exhibit interesting properties. A remarkable alternative is the fractional-order hold (FROH), which provokes no intersample ripple under steady state and, if properly tuned, can often place the discretized p...
It is well established that the FRactional Order Hold (FROH), when properly tuned, enhances the performance of digital control systems. Indeed, FROH reconstruction can bring the zeros of the z-domain model closer to the origin than zero order hold reconstruction. This work applies numerical analysis techniques in order to obtain as stable as possib...
Zero-order hold devices (ZOH) are mainly used in digital control applications to obtain the output analog signals of digitally implemented controllers. However, it is proved that special hold devices such as the fractional order hold device (FROH) can improve, if properly tuned, the performance of hybrid control systems. Although the applicability...
Recently much experimental evidence has accumulated concerning intracellular calcium and its fundamental role as a regulator in eukaryotic cells. The literature relating to Ca2+ in fungi is large and diverse and this paper draws together the available information and discusses the particular functions of the ion in this group of organisms.Uptake me...
Asexual spores or conidia are dispersive propagules produced as an alternative to vegetative growth by a diverse group of filamentous fungi. The cellular development programmes which govern conidiation have been intensely studied in the last few decades, although important gaps stand in the way of our understanding of this phenomenon, namely in the...
The final step in the secretory pathway, which is the fusion event between secretory vesicles and the plasma membrane, was reconstructed using highly purified secretory vesicles and cytoplasmic-side-out plasma membrane vesicles from the yeast Saccharomyces cerevisiae. Both organelle preparations were obtained from a sec 6-4 temperature-sensitive mu...
Single Cell Protein (SCP) is a term coined in the 1960´s to embrace microbial biomass products which were produced by fermentation. SCP production technologies arose as a promising way to solve the problem of worldwide protein shortage. They evolved as bioconversion processes which turned low value by-products, often wastes, into products with adde...
The filamentous fungus Penicillium cyclopium conidiates in the presence of calcium ions in submerged culture without nutrient limitation according to a precisely timed
program. Conidiation could be prematurely induced in a nutritionally sufficient medium which had previously supported growth,
suggesting that a metabolite which influenced the proces...
The isolation and structure determination of conidiogenol and conidiogenone, tetracyclic diterpenes with a novel carbon skeleton, from extracts of the fermentation broth of Penicillium cyclopium is reported. Conidiogenol and conidiogenone are potent and selective inducers of conidiogenesis in P. cyclopium in liquid culture, and relay information ab...
A diploid strain of Saccharomyces cerevisiae able to metabolize lactose with high efficiency has been obtained. Haploid strains of Saccharomyces able to grow on lactose were constructed by cotransformation with two genes of Kluyveromyces lactis required for the utilization of the sugar, LAC4 and LAC12, encoding beta-galactosidase and lactose permea...
The amount of acid or base consumed in yeast cultures has been recently assigned to the pathway of nitrogen assimilation under respiratory conditions with no contribution by carbon metabolism (Castrillo et al., 1995). In this investigation, experiments under respirofermentative conditions have shown that production or consumption of ethanol does no...
Whey proteins were hydrolyzed in a membrane recycle reactor (MRR) in the presence of Alcalase 0.6L under varying enzyme to substrate ratios and residence times. Substrate conversion was directly dependent on these two parameters, and at a fixed enzyme-substrate ratio of 10%, conversion levels could be controlled through residence time, resulting in...
In this investigation, a method for the accurate quantitative determination of net proton production or consumption in biological cultures has been devised. Cells are cultured under constant pH conditions. The specific rate of proton production or consumption by the culture (qH+, mmol h−1 per g biomass) is proportional to the mmol of base or acid r...
Highly purified plasma membrane vesicles were prepared from yeast protoplasts by a combination of osmotic lysis, differential centrifugation, and separation in an aqueous dextran/polyethylene glycol two-phase system. The vesicles were predominantly (85-90%) of cytoplasmic side-out orientation and displayed large ATP-dependent proton pumping activit...
The pattern of energy metabolism of different types of yeasts (obligate aerobes and facultative anaerobes) in aerobic chemostat cultures has been evaluated and interpreted on the basis of a coupling of metabolic fluxes between glycolytic and oxidative components. A model has been formulated which defines glycolytic and oxidative subunits through wh...
A whey protein hydrolysate was prepared by incubation of reconstituted whey or a whey protein concentrate with Alcalase 0.6L. The proteolytic degradation of α-lactalbumin and β-lactoglobulin initially resulted in 6-kDa and, later, 2.5-kDa degradation products, quickly followed by the appearance of multiple peptides of 1 kDa or smaller. The hydrolys...
Addition of Ca2+ (1 to 10 mM) to submerged cultures of Penicillium cyclopium induces conidiation. Ca2+ induced an increase in cytosolic pH from approximately 7.00 to > 7.60 in less than 10 min, as determined with the fluorescent pH probe fluorescein. Measurement of the H(+)-ATPase activity in total membrane fractions did not show any stable activat...
The influence of physiological parameters such as carbon substrate flux and O2 uptake rates on energy metabolism are reported with reference to biomass productivity in whey chemostat culture. The combined results show that oxidoreductive energy metabolism may be attained independently of the yeast reaching its maximum respiratory capacity. A novel...
A plasma membrane fraction was obtained by the combined use of differential centrifugation and aqueous polymer two-phase partitioning techniques. Vanadate-inhibited ATPase and glucan synthase activities were highly enriched in this fraction, although the presence of ATPase activity which was not inhibited by vanadate, nitrate, molybdate, anyimycin...
The behavior of Kluyveromyces marxianus in aerobic chemostat cultures of deproteinated whey was studied in order to determine the characteristics of energy metabolism. The combined results show that K. marxianus presents a different pattern of behavior compared to Saccharomyces cereuisiae. A mathematical model has been constructed where energy meta...
A procedure for the production of plasma membrane vesicles, consisting of enzyme-assisted physical homogenization, differential centrifugation and partitioning in an aqueous polymer two phase system resulted in a fraction containing highly purified plasma membranes, as determined by vanadate-inhibited H+ ATPase and near absence of intracellular mem...
A new simple procedure for the production of protoplasts of Penicillium cyclopium with high regeneration rates and efficient transport activity is described, involving the use of a preparation of Novozym 234 with very low protease activity. The combination of a heat pretreatment at 55°C for 15 min with the use of the protease inhibitor aprotinin re...
Cell wall-free protoplasts of P. cyclopium could regenerate a cell wall and form mycelia in liquid culture with high rates of viability. When calcium was added to the medium, protoplasts displayed biphasic accumulation with an immediate metabolism-independent adsorption phase, followed by slow metabolism-dependent uptake. Exposure of the protoplast...
Penicillium notatum IMI 15378 sporulated profusely in submerged culture containing 10 mM-Ca2+, whilst the response of strain NRRL 832 was minimal up to 40 mm concentration of the cation.Calcium accumulation by mycelia of both strains was biphasic and involved rapid initial energy-independent binding, with the strains showing similar affinities for...
The respiratory properties of isolated mitochondria from P. cyclopium were studied with particular attention to their response to calcium ions. The results obtained indicate concentration dependent stimulation of NADH oxidation by calcium ions. Similar effects could also be obtained with other divalent cations.
The conversion of whey to single-cell protein by yeasts was investigated. The most suitable organism tested wasKluyveromyces
marxianus NCYC 1424. The efficiency of whey conversion to biomass was directly related to higher oxygen availability in the medium.
Uptake of calcium ions by Penicillium cyclopium in submerged culture was biphasic. The ion underwent rapid initial binding to the mycelium with a dissociation constant (KD) value of 0.114 mM-Ca2+ and binding capacity (Yt) of 3.5 nmol-Ca2+ (mg dry wt)−1. Metabolismindependent chemical adsorption of calcium was followed by metabolism-dependent active...