Nitrogen starvation or glucose limitation induces conidiation in constantly shaken liquid cultures of Neurospora crassa.

ABSTRACT We have identified a new gene encoding the G protein α subunit,gna-3, from the filamentous fungusNeurospora crassa. The predicted amino acid sequence of GNA-3 is most similar to the Gα proteins MOD-D, MAGA, and CPG-2 from the saprophytic fungus Podospora anserina and the pathogenic fungi Magnaporthe grisea and Cryphonectria parasitica, respectively. Deletion of gna-3 leads to shorter aerial hyphae and premature, dense conidiation during growth on solid medium or in standing liquid cultures and to inappropriate conidiation in submerged culture. The conidiation and aerial hypha defects of the Δgna-3 strain are similar to those of a previously characterized adenylyl cyclase mutant, cr-1. Supplementation with cyclic AMP (cAMP) restores wild-type morphology to Δgna-3 strains in standing liquid cultures. Solid medium augmented with exogenous cAMP suppresses the premature conidiation defect, but aerial hypha formation is still reduced. Submerged-culture conidiation is refractory to cAMP but is suppressed by peptone. In addition, Δgna-3 submerged cultures express the glucose-repressible gene, qa-2, to levels greatly exceeding those observed in the wild type under carbon-starved conditions. Δgna-3 strains exhibit reduced fertility in homozygous crosses during the sexual cycle; exogenous cAMP has no effect on this phenotype. Intracellular steady-state cAMP levels of Δgna-3 strains are decreased 90% relative to the wild type under a variety of growth conditions. Reduced intracellular cAMP levels in the Δgna-3 strain correlate with lower adenylyl cyclase activity and protein levels. These results demonstrate that GNA-3 modulates conidiation and adenylyl cyclase levels in N. crassa.

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Regulation of exocellular protease in Neurospora crassa: Induction and repression under conditions of nitrogen starvation

Archives of Biochemistry and Biophysics ◽

10.1016/0003-9861(77)90541-0 ◽

1977 ◽

Vol 182 (2) ◽

pp. 601-613 ◽

Cited By ~ 34

Author(s):

Bernard L. Cohen ◽

Harvey Drucker

Keyword(s):

Neurospora Crassa ◽

Nitrogen Starvation

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The wis1 signal transduction pathway is required for expression of cAMP-repressed genes in fission yeast

Journal of Cell Science ◽

10.1242/jcs.109.7.1927 ◽

1996 ◽

Vol 109 (7) ◽

pp. 1927-1935 ◽

Cited By ~ 13

Author(s):

S. Stettler ◽

E. Warbrick ◽

S. Prochnik ◽

S. Mackie ◽

P. Fantes

Keyword(s):

Protein Kinase ◽

Map Kinase ◽

Nitrogen Starvation ◽

Osmotic Shock ◽

Regulatory Subunit ◽

Loss Of Function ◽

Dependent Protein Kinase ◽

Glucose Limitation ◽

Camp Dependent Protein Kinase ◽

Dependent Protein

The wis1 protein kinase of Schizosaccharomyces pombe is a member of the MAP kinase kinase family. Loss of wis1 function has previously been reported to lead to a delay in the G2-mitosis transition, loss of viability in stationary phase, and hypersensitivity to osmotic shock. It acts at least in part by activating the MAP kinase homologue sty1; loss-of-function sty1 mutants share many phenotypes with wis1 deletion mutants. We show here that, in addition, loss of wis1 function leads to defective conjugation, and to suppression of the hyperconjugation phenotype of the pat1-114 mutation. Consistent with this, the induction of the mei2 gene, which is normally induced by nitrogen starvation, is defective in wis1 mutants. In wild-type cells, nitrogen starvation leads to mei2 induction through a fall in intracellular cyclic AMP (cAMP) level and activity of the cAMP-dependent protein kinase. We show here that wis1 function is required for mei2 induction following nitrogen starvation. Expression of the fbp1 gene is negatively regulated by cAMP in response to glucose limitation: induction of fbp1 also requires wis1 and sty1 function. Loss of wis1 is epistatic over increased fbp1 expression brought about by loss of adenylate cyclase (git2/cyr1) or cAMP-dependent protein kinase (pka1) function. These observations can be explained by a model in which the pka1 pathway negatively regulates the wis1 pathway, or the two pathways might act independently on downstream targets. The latter explanation is supported, at least as regards regulation of cell division, by the observation that loss of function of the regulatory subunit of the cAMP-dependent protein kinase (cgs1) brings about a modest increase in cell length at division in both wis1+ and wis1 delta genetic backgrounds.

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Shared and Independent Roles for a Gαi Protein and Adenylyl Cyclase in Regulating Development and Stress Responses in Neurospora crassa

Eukaryotic Cell ◽

10.1128/ec.1.4.634-642.2002 ◽

2002 ◽

Vol 1 (4) ◽

pp. 634-642 ◽

Cited By ~ 50

Author(s):

F. Douglas Ivey ◽

Ann M. Kays ◽

Katherine A. Borkovich

Keyword(s):

Neurospora Crassa ◽

Adenylyl Cyclase ◽

Stress Responses ◽

Growth And Development ◽

Submerged Culture ◽

Wild Type Strain ◽

Wild Type ◽

Liquid Cultures ◽

Aerial Hyphae ◽

Gαi Protein

ABSTRACT Growth and development are regulated using cyclic AMP (cAMP)-dependent and -independent pathways in Neurospora crassa. The cr-1 adenylyl cyclase mutant lacks detectable cAMP and exhibits numerous defects, including colonial growth habit, short aerial hyphae, premature conidiation on plates, inappropriate conidiation in submerged culture, and increased thermotolerance. Evidence suggests that the heterotrimeric Gα protein GNA-1 is a direct positive regulator of adenylyl cyclase. Δgna-1 strains are female-sterile, and Δgna-1 strains have reduced apical extension rates on normal and hyperosmotic medium, greater resistance to oxidative and heat stress, and stunted aerial hyphae compared to the wild-type strain. In this study, a Δgna-1 cr-1 double mutant was analyzed to differentiate cAMP-dependent and -independent signaling pathways regulated by GNA-1. Δgna-1 cr-1 mutants have severely restricted colonial growth and do not produce aerial hyphae on plates or in standing liquid cultures. Addition of cAMP to plates or standing liquid cultures rescues cr-1, but not Δgna-1 cr-1, defects, which is consistent with previous results demonstrating that Δgna-1 mutants do not respond to exogenous cAMP. The females of all strains carrying the Δgna-1 mutation are sterile; however, unlike cr-1 and Δgna-1 strains, the Δgna-1 cr-1 mutant does not produce protoperithecia. The Δgna-1 and cr-1 mutations were synergistic with respect to inappropriate conidiation during growth in submerged culture. Thermotolerance followed the order wild type < Δgna-1 < cr-1 = Δgna-1 cr-1, consistent with a cAMP-dependent process. Taken together, the results suggest that in general, GNA-1 and CR-1 regulate N. crassa growth and development using parallel pathways, while thermotolerance is largely dependent on cAMP.

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Pseudohyphal differentiation in Komagataella phaffii: investigating the FLO gene family

FEMS Yeast Research ◽

10.1093/femsyr/foaa044 ◽

2020 ◽

Vol 20 (5) ◽

Author(s):

Sonakshi De ◽

Corinna Rebnegger ◽

Josef Moser ◽

Nadine Tatto ◽

Alexandra B Graf ◽

...

Keyword(s):

Gene Family ◽

Nitrogen Starvation ◽

Slow Growth ◽

Transcriptional Analysis ◽

Glucose Limitation ◽

Pseudohyphal Growth ◽

Komagataella Phaffii ◽

Fusel Alcohols ◽

Adverse Environmental Conditions ◽

Pseudohyphal Differentiation

ABSTRACT Many yeasts differentiate into multicellular phenotypes in adverse environmental conditions. Here, we investigate pseudohyphal growth in Komagataella phaffii and the involvement of the flocculin (FLO) gene family in its regulation. The K. phaffii FLO family consists of 13 members, and the conditions inducing pseudohyphal growth are different from Saccharomyces cerevisiae. So far, this phenotype was only observed when K. phaffii was cultivated at slow growth rates in glucose-limited chemostats, but not upon nitrogen starvation or the presence of fusel alcohols. Transcriptional analysis identified that FLO11, FLO400 and FLO5-1 are involved in the phenotype, all being controlled by the transcriptional regulator Flo8. The three genes exhibit a complex mechanism of expression and repression during transition from yeast to pseudohyphal form. Unlike in S. cerevisiae, deletion of FLO11 does not completely prevent the phenotype. In contrast, deletion of FLO400 or FLO5-1 prevents pseudohyphae formation, and hampers FLO11 expression. FAIRE-Seq data shows that the expression and repression of FLO400 and FLO5-1 are correlated to open or closed chromatin regions upstream of these genes, respectively. Our findings indicate that K. phaffii Flo400 and/or Flo5-1 act as upstream signals that lead to the induction of FLO11 upon glucose limitation in chemostats at slow growth and chromatin modulation is involved in the regulation of their expression.

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Ultrastructure and Morphology of the Neurospora Crassa Cell Wall Mutants, Slime And Slime X, Using Tem and Sem

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100090257 ◽

1976 ◽

Vol 34 ◽

pp. 54-55

Author(s):

Karen S. Howard ◽

H. D. Braymer ◽

M. D. Socolofsky ◽

S. A. Milligan

Keyword(s):

Cell Wall ◽

Neurospora Crassa ◽

Wild Type ◽

Morphological Comparison ◽

Small Areas ◽

Solid Media ◽

Thin Sectioning ◽

X Cells ◽

Mutant Cells ◽

Isolated Cell

The recently isolated cell wall mutant slime X of Neurospora crassa was prepared for ultrastructural and morphological comparison with the cell wall mutant slime. The purpose of this article is to discuss the methods of preparation for TEM and SEM observations, as well as to make a preliminary comparison of the two mutants.TEM: Cells of the slime mutant were prepared for thin sectioning by the method of Bigger, et al. Slime X cells were prepared in the same manner with the following two exceptions: the cells were embedded in 3% agar prior to fixation and the buffered solutions contained 5% sucrose throughout the procedure.SEM: Two methods were used to prepare mutant and wild type Neurospora for the SEM. First, single colonies of mutant cells and small areas of wild type hyphae were cut from solid media and fixed with OSO4 vapors similar to the procedure used by Harris, et al. with one alteration. The cell-containing agar blocks were dehydrated by immersion in 2,2-dimethoxypropane (DMP).

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