Modeling Neurospora growth

Modeling Neurospora growt

Fluorescent staining of Neurospora nuclei with DAPI

Fluorescent staining of Neurospora nuclei with DAP

Effects of glucagon on the growth of Neurospora

Effects of glucagon on the growth of Neurospora

Antagonism between salicylate and the cAMP signal controls yeast cell survival and growth recovery from quiescence

Aspirin and its main metabolite salicylate are promising molecules in preventing cancer and metabolic diseases. S. cerevisiae cells have been used to study some of their effects: (i) salicylate induces the reversible inhibition of both glucose transport and the biosyntheses of glucose-derived sugar phosphates, (ii) Aspirin/salicylate causes apoptosis associated with superoxide radical accumulation or early cell necrosis in MnSOD-deficient cells growing in ethanol or in glucose, respectively. So, treatment with (acetyl)-salicylic acid can alter the yeast metabolism and is associated with cell death. We describe here the dramatic effects of salicylate on cellular control of the exit from a quiescence state. The growth recovery of long-term stationary phase cells was strongly inhibited in the presence of salicylate, to a degree proportional to the drug concentration. At high salicylate concentration, growth reactivation was completely repressed and associated with a dramatic loss of cell viability. Strikingly, both of these phenotypes were fully suppressed by increasing the cAMP signal without any variation of the exponential growth rate. Upon nutrient exhaustion, salicylate induced a premature lethal cell cycle arrest in the budded-G2/M phase that cannot be suppressed by PKA activation. We discuss how the dramatic antagonism between cAMP and salicylate could be conserved and impinge common targets in yeast and humans. Targeting quiescence of cancer cells with stem-like properties and their growth recovery from dormancy are major challenges in cancer therapy. If mechanisms underlying cAMP-salicylate antagonism will be defined in our model, this might have significant therapeutic implications

Role of guanine nucleotides in the regulation of the Ras/cAMP pathway in Saccharomyces cerevisiae

AbstractThe CDC25 gene product is a guanine nucleotide exchange factor for Ras proteins in yeast. Recently it has been suggested that the intracellular levels of guanine nucleotides may influence the exchange reaction. To test this hypothesis we measured the levels of nucleotides in yeast cells under different growth conditions and the relative amount of Ras2-GTP. The intracellular GTP/GDP ratio was found to be very sensitive to growth conditions: the ratio is high, close to that of ATP/ADP during exponential growth, but it decreases rapidly before the beginning of stationary phase, and it drops further under starvation conditions. The addition of glucose to glucose-starved cells causes a fast increase of the GTP/GDP ratio. The relative amount of Ras2-GTP changes in a parallel way suggesting that there is a correlation with the cytosolic GTP/GDP ratio. In addition ‘in vitro’ mixed-nucleotide exchange experiments done on purified Ras2 protein demonstrated that the GTP and GDP concentrations influence the extent of Ras2-GTP loading giving further support to their possible regulatory role

Competencia y crecimiento de árboles nativos establecidos espontáneamente en plantaciones de <i>Araucaria angustifolia</i> en el NE de Misiones

Entender los procesos vinculados a la regeneración de especies arbóreas nativas en plantaciones comerciales es fundamental para favorecer prácticas que permitan incorporar mayor biodiversidad y mejorar la eficiencia en el uso de los recursos. En este tipo de rodales, el crecimiento de las especies establecidas de manera espontánea depende de las relaciones de competencia que se establecen entre ellas y a su vez, la respuesta del crecimiento de los árboles puede diferir en especies con distinta densidad de la madera. Conociendo estas diferencias, y a través de un manejo apropiado de control de estas relaciones, podría lograrse un incremento apreciable del crecimiento de los árboles regenerados naturalmente. El objetivo de este trabajo fue evaluar el efecto de la competencia sobre el crecimiento de árboles nativos establecidos espontáneamente en plantaciones de Araucaria angustifolia (Bertol.) Kuntze en Misiones. El estudio se llevó a cabo a partir de medidas obtenidas en parcelas permanentes del Campo Anexo Manuel Belgrano (CAMB), en la localidad de San Antonio. Se utilizaron índices que reflejan las relaciones de competencia entre especies no implantadas respecto de la especie implantada. Las especies fueron clasificadas en dos clases de densidad de la madera (baja y media-alta). Teniendo en cuenta especie y densidad de la madera se implementó un modelo para analizar las diferencias de respuesta del crecimiento. De la misma manera se evaluaron las relaciones entre el volumen de copa, la competencia y el crecimiento para determinar si esta medida de copa podría reflejar el estado de desarrollo de las especies. En base a los resultados concluimos que los individuos de araucaria no presentan relaciones de competencia con individuos de las especies pertenecientes a la regeneración espontánea. Las especies con baja densidad de la madera resultaron menos tolerantes a la escasez de recursos respecto de especies de mayores densidades de la madera. El volumen de copa no tuvo relación con la competencia mientras que se relacionó positivamente con el crecimiento en especies de densidad de la madera media-alta. Se concluyó que, a través de la implementación de tratamientos silvícolas que determinen una disminución de la competencia en las especies de baja densidad de la madera, podría lograrse un mayor crecimiento de las mismas dada su sensibilidad a la competencia. Este tipo de prácticas puede favorecer el rendimiento de sistemas productivos que aumenten la complejidad y promuevan la regeneración de otras especies nativas.Facultad de Ciencias Agrarias y Forestale

Searching for antimicrobial photosensitizers among a panel of BODIPYs

In recent years, antimicrobial Photodynamic Therapy (aPDT) gained increasing attention for its potential to inhibit the growth and spread of microorganisms, both as free-living cells and/or embedded in biofilm communities. In this scenario, compounds belonging to the family of boron-dipyrromethenes (BODIPYs) represent a very promising class of photosensitizers for applications in antimicrobial field. In this study, twelve non-ionic and three cationic BODIPYs were assayed for the inactivation of Staphylococcus aureus, Pseudomonas aeruginosa and Candida albicans. As expected, S. aureus showed to be very sensitive to BODIPYs and mild conditions were sufficient to reach good rates of photoinactivation with both neutral and monocationic ones. Surprisingly, one neutral compound (named B9 in this study) resulted the best BODIPY to photoinactivate P. aeruginosa PAO1. The photoinactivation of C. albicans was reached with both neutral and mono-cationic BODIPYs. Furthermore, biofilms of the three model microorganisms were challenged with BODIPYs in light-based antimicrobial technique. S. aureus biofilms were successfully inhibited with milder conditions than those applied to P. aeruginosa and C. albicans. Notably, it was possible to eradicate 24-h-old biofilms of both S. aureus and P. aeruginosa. In conclusion, this study supports the potential of neutral BODIPYs as pan-antimicrobial PSs. Graphical abstract: [Figure not available: see fulltext.]

Analysis of the Secondary Structure of the Catalytic Domain of Mouse Ras Exchange Factor CDC25Mm

The minimal active domain GEF domain. of the mouse Ras exchange factor CDC25Mm was purified to homogeneity from recombinant Escherichia coli culture. The 256 amino acids polypeptide shows high activity in vitro and forms a stable complex with H-ras p21 in absence of guanine nucleotides. Circular dichroism CD. spectra in the far UV region indicate that this domain is highly structured with a high content of a-helix 42%.. Near UV CD spectra evidenced good signal due to phenylalanine and tyrosine while a poor contribution was elicited by the three tryptophan residues contained in this domain. The tryptophan fluorescence signal was scarcely affected by denaturation of the protein or by formation of the binary complex with H-ras p21, suggesting that the Trp residues, which are well conserved in the GEF domain of several Ras-exchange factors, were exposed to the surface of the protein and they are not most probably directly involved in the interaction with Ras proteins. q1998 Elsevier Science B.

The budding yeast RasGEF Cdc25 reveals an unexpected nuclear localization

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Activation State of the Ras2 Protein and Glucose-induced Signaling in Saccharomyces cerevisiae

The activity of adenylate cyclase in the yeast Saccharomyces cerevisiae is controlled by two G-protein systems, the Ras proteins and the Gα protein Gpa2. Glucose activation of cAMP synthesis is thought to be mediated by Gpa2 and its G-protein-coupled receptor Gpr1. Using a sensitive GTP-loading assay for Ras2 we demonstrate that glucose addition also triggers a fast increase in the GTP loading state of Ras2 concomitant with the glucose-induced increase in cAMP. This increase is severely delayed in a strain lacking Cdc25, the guanine nucleotide exchange factor for Ras proteins. Deletion of the Ras-GAPs IRA2 (alone or with IRA1) or the presence of RAS2Val19 allele causes constitutively high Ras GTP loading that no longer increases upon glucose addition. The glucose-induced increase in Ras2 GTP-loading is not dependent on Gpr1 or Gpa2. Deletion of these proteins causes higher GTP loading indicating that the two G-protein systems might directly or indirectly interact. Because deletion of GPR1 or GPA2 reduces the glucose-induced cAMP increase the observed enhancement of Ras2 GTP loading is not sufficient for full stimulation of cAMP synthesis. Glucose phosphorylation by glucokinase or the hexokinases is required for glucose-induced Ras2 GTP loading. These results indicate that glucose phosphorylation might sustain activation of cAWDP synthesis by enhancing Ras2 GTP loading likely through inhibition of the Ira proteins. Strains with reduced feedback inhibition on cAMP synthesis also display elevated basal and induced Ras2 GTP loading consistent with the Ras2 protein acting as a target of the feedback-inhibition mechanism