scholarly journals Relationship of the Major Constituents of the Neurospora crassa Cell Wall to Wild-Type and Colonial Morphology

1965 ◽  
Vol 90 (4) ◽  
pp. 1073-1081 ◽  
Author(s):  
P. R. Mahadevan ◽  
E. L. Tatum
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Wild Type ◽  
Colonial Morphology ◽  
Relationship Of

Transformation of Neurospora crassa with the cloned am (glutamate dehydrogenase) gene

1984 ◽  
Vol 4 (1) ◽  
pp. 117-122
Author(s):  
J A Kinsey ◽  
J A Rambosek
Keyword(s):  
Neurospora Crassa ◽  
Glutamate Dehydrogenase ◽  
Plasmid Vector ◽  
Wild Type ◽  
Mendelian Segregation ◽  
Dehydrogenase Gene ◽  
Mutant Strains ◽  
Colonial Morphology ◽  
Relationship Of ◽  
Lambda Dna

We used DNA containing the am gene of Neurospora crassa, cloned in the lambda replacement vector lambdaL-47 (this clone is designated lambdaC-10), and plasmid vector subclones of this DNA to transform am deletion and point mutant strains. By means of subcloning, all sequences required for transformation to am prototrophy and expression of glutamate dehydrogenase have been shown to reside on a 2.5-kilobase BamHI fragment. We also characterized several am+ strains that were obtained after transformation with lambdaC-10. These strains showed Mendelian segregation of the am+ gene, although less than 50% of the transformed strains showed the normal linkage relationship of am with inl. In all cases tested, the strains had incorporated lambda DNA as well. The lambda DNA also showed a Mendelian segregation pattern. In one case, the incorporation of am DNA in a novel position was associated with a mutagenic event producing a strain with a very tight colonial morphology. In all cases in which the am+ gene had become the resident of a new chromosome, glutamate dehydrogenase was produced to only 10 to 20% of the wild-type levels.

scholarly journals Transformation of Neurospora crassa with the cloned am (glutamate dehydrogenase) gene.

1984 ◽  
Vol 4 (1) ◽  
pp. 117-122 ◽  
Author(s):  
J A Kinsey ◽  
J A Rambosek
Keyword(s):  
Neurospora Crassa ◽  
Glutamate Dehydrogenase ◽  
Plasmid Vector ◽  
Wild Type ◽  
Mendelian Segregation ◽  
Dehydrogenase Gene ◽  
Mutant Strains ◽  
Colonial Morphology ◽  
Relationship Of ◽  
Lambda Dna

We used DNA containing the am gene of Neurospora crassa, cloned in the lambda replacement vector lambdaL-47 (this clone is designated lambdaC-10), and plasmid vector subclones of this DNA to transform am deletion and point mutant strains. By means of subcloning, all sequences required for transformation to am prototrophy and expression of glutamate dehydrogenase have been shown to reside on a 2.5-kilobase BamHI fragment. We also characterized several am+ strains that were obtained after transformation with lambdaC-10. These strains showed Mendelian segregation of the am+ gene, although less than 50% of the transformed strains showed the normal linkage relationship of am with inl. In all cases tested, the strains had incorporated lambda DNA as well. The lambda DNA also showed a Mendelian segregation pattern. In one case, the incorporation of am DNA in a novel position was associated with a mutagenic event producing a strain with a very tight colonial morphology. In all cases in which the am+ gene had become the resident of a new chromosome, glutamate dehydrogenase was produced to only 10 to 20% of the wild-type levels.

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

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).

scholarly journals PHASE-SPECIFIC GENES FOR MACROCONIDIATION IN NEUROSPORA CRASSA

Genetics ◽  
1974 ◽  
Vol 78 (2) ◽  
pp. 679-690 ◽  
Author(s):  
Claude P Selitrennikoff ◽  
Robert E Nelson ◽  
Richard W Siegel
Keyword(s):  
Cell Wall ◽  
Life Cycle ◽  
Neurospora Crassa ◽  
Sexual Life ◽  
Wild Type ◽  
A Cell ◽  
Autolytic Activity ◽  
Sexual Life Cycle ◽  
Mutant Genes ◽  
Active Formation

ABSTRACT Two new mutant genes in Neurospora crassa prevent the formation of free macroconidia from proconidial chains. These genes, called conidial separation-1 and conidial separation-2, are phase-specific, playing no role in either the sexual life cycle or other aspects of the asexual life cycle. A cell-wall-associated autolytic activity was found to increase in wild-type cultures at the time of active formation of free conidia from proconidial chains; no such increase was detected in mutant cultures. It appears that the products of these genes are both essential for and unique to macroconidiation.

Production of plant cell wall degrading enzymes by the exo-1 mutant and wild type Neurospora crassa

2010 ◽  
Vol 150 ◽  
pp. 513-514
Author(s):  
A.M. Polizeli ◽  
M.A. Moraes ◽  
J.A. Jorge ◽  
H.F. Terenzi ◽  
M.L.T.M. Polizeli
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Plant Cell ◽  
Plant Cell Wall ◽  
Wild Type ◽  
Cell Wall Degrading Enzymes ◽  
Degrading Enzymes

scholarly journals The Predicted Mannosyltransferase GT69-2 Antagonizes RFW-1 To Regulate Cell Fusion in Neurospora crassa

mBio ◽  
2021 ◽  
Vol 12 (2) ◽  
Author(s):  
Yang Li ◽  
Jens Heller ◽  
A. Pedro Gonçalves ◽  
N. Louise Glass
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Somatic Cell ◽  
Filamentous Fungi ◽  
Cell Fusion ◽  
Wild Type ◽  
Content Type ◽  
Asexual Sporulation ◽  
Somatic Cell Fusion ◽  
Cell Wall Remodeling

ABSTRACT Filamentous fungi undergo somatic cell fusion to create a syncytial, interconnected hyphal network which confers a fitness benefit during colony establishment. However, barriers to somatic cell fusion between genetically different cells have evolved that reduce invasion by parasites or exploitation by maladapted genetic entities (cheaters). Here, we identified a predicted mannosyltransferase, glycosyltransferase family 69 protein (GT69-2) that was required for somatic cell fusion in Neurospora crassa. Cells lacking GT69-2 prematurely ceased chemotropic signaling and failed to complete cell wall dissolution and membrane merger in pairings with wild-type cells or between Δgt69-2 cells (self fusion). However, loss-of-function mutations in the linked regulator of cell fusion and cell wall remodeling-1 (rfw-1) locus suppressed the self-cell-fusion defects of Δgt69-2 cells, although Δgt69-2 Δrfw-1 double mutants still failed to undergo fusion with wild-type cells. Both GT69-2 and RFW-1 localized to the Golgi apparatus. Genetic analyses indicated that RFW-1 negatively regulates cell wall remodeling-dependent processes, including cell wall dissolution during cell fusion, separation of conidia during asexual sporulation, and conidial germination. GT69-2 acts as an antagonizer to relieve or prevent negative functions on cell fusion by RFW-1. In Neurospora species and N. crassa populations, alleles of gt69-2 were highly polymorphic and fell into two discrete haplogroups. In all isolates within haplogroup I, rfw-1 was conserved and linked to gt69-2. All isolates within haplogroup II lacked rfw-1. These data indicated that gt69-2/rfw-1 are under balancing selection and provide new mechanisms regulating cell wall remodeling during cell fusion and conidial separation. IMPORTANCE Cell wall remodeling is a dynamic process that balances cell wall integrity versus cell wall dissolution. In filamentous fungi, cell wall dissolution is required for somatic cell fusion and conidial separation during asexual sporulation. In the filamentous fungus Neurospora crassa, allorecognition checkpoints regulate the cell fusion process between genetically different cells. Our study revealed two linked loci with transspecies polymorphisms and under coevolution, rfw-1 and gt69-2, which form a coordinated system to regulate cell wall remodeling during somatic cell fusion, conidial separation, and asexual spore germination. RFW-1 acts as a negative regulator of these three processes, while GT69-2 functions antagonistically to RFW-1. Our findings provide new insight into the mechanisms involved in regulation of fungal cell wall remodeling during growth and development.

scholarly journals Copper accumulation in the cell-wall-deficient slime variant of Neurospora crassa. Comparison with a wild-type strain

1987 ◽  
Vol 245 (2) ◽  
pp. 479-484 ◽  
Author(s):  
U A Germann ◽  
K Lerch
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Type Strain ◽  
Copper Concentration ◽  
Culture Medium ◽  
Wild Type Strain ◽  
Copper Accumulation ◽  
Growth Time ◽  
Copper Binding ◽  
Wild Type

The copper-uptake process in the cell-wall-deficient slime variant of the fungus Neurospora crassa was compared with that in a wild-type strain. In both organisms investigated most of the copper is taken up from the culture medium during the exponential growth period. The wild-type strain, however, accumulates much more copper than does the slime variant. The influence of the copper concentration in the culture medium on the amounts of copper accumulated intracellularly suggests separate ways of copper import used by the two morphologically different N. crassa strains. Copper analyses of three different cytosolic fractions as a function of growth time or exogenous copper concentration indicate both strains to share a very similar copper metabolism. All the data presented are consistent with a detoxification function of the low-Mr copper-binding fraction of N. crassa. Both copper-metallothionein and oxidized glutathione (GSSG) are co-eluted with this fraction. The possible involvement of glutathione in metallothionein biosynthesis is discussed.

scholarly journals α-1,6-Mannosylation of N-Linked Oligosaccharide Present on Cell Wall Proteins Is Required for Their Incorporation into the Cell Wall in the Filamentous Fungus Neurospora crassa

2010 ◽  
Vol 9 (11) ◽  
pp. 1766-1775 ◽  
Author(s):  
Abhiram Maddi ◽  
Stephen J. Free
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Mutant Cell ◽  
Cell Wall Protein ◽  
Cell Wall Proteins ◽  
Wild Type ◽  
Content Type ◽  
Fold Reduction ◽  
Total Lack ◽  
Mutant Cells

ABSTRACT The enzyme α-1,6-mannosyltransferase (OCH-1) is required for the synthesis of galactomannans attached to the N-linked oligosaccharides of Neurospora crassa cell wall proteins. The Neurospora crassa och-1 mutant has a tight colonial phenotype and a defective cell wall. A carbohydrate analysis of the och-1 mutant cell wall revealed a 10-fold reduction in the levels of mannose and galactose and a total lack of 1,6-linked mannose residues. Analysis of the integral cell wall protein from wild-type and och-1 mutant cells showed that the mutant cell wall had reduced protein content. The och-1 mutant was found to secrete 18-fold more protein than wild-type cells. Proteomic analysis of the proteins released by the mutant into the growth medium identified seven of the major cell wall proteins. Western blot analysis of ACW-1 and GEL-1 (two glycosylphosphatidylinositol [GPI]-anchored proteins that are covalently integrated into the wild-type cell wall) showed that high levels of these proteins were being released into the medium by the och-1 mutant. High levels of ACW-1 and GEL-1 were also released from the och-1 mutant cell wall by subjecting the wall to boiling in a 1% SDS solution, indicating that these proteins are not being covalently integrated into the mutant cell wall. From these results, we conclude that N-linked mannosylation of cell wall proteins by OCH-1 is required for their efficient covalent incorporation into the cell wall.

scholarly journals Inhibition of cell expansion enhances cortical microtubule stability in the root apex of Arabidopsis thaliana

2021 ◽  
Vol 28 (1) ◽  
Author(s):  
Veronica Giourieva ◽  
Emmanuel Panteris
Keyword(s):  
Arabidopsis Thaliana ◽  
Cell Wall ◽  
Cell Expansion ◽  
Cortical Microtubule ◽  
Root Apex ◽  
Cellulose Synthase ◽  
Cellulose Biosynthesis ◽  
Cortical Microtubules ◽  
Wild Type ◽  
Microtubule Stability

Abstract Background Cortical microtubules regulate cell expansion by determining cellulose microfibril orientation in the root apex of Arabidopsis thaliana. While the regulation of cell wall properties by cortical microtubules is well studied, the data on the influence of cell wall to cortical microtubule organization and stability remain scarce. Studies on cellulose biosynthesis mutants revealed that cortical microtubules depend on Cellulose Synthase A (CESA) function and/or cell expansion. Furthermore, it has been reported that cortical microtubules in cellulose-deficient mutants are hypersensitive to oryzalin. In this work, the persistence of cortical microtubules against anti-microtubule treatment was thoroughly studied in the roots of several cesa mutants, namely thanatos, mre1, any1, prc1-1 and rsw1, and the Cellulose Synthase Interacting 1 protein (csi1) mutant pom2-4. In addition, various treatments with drugs affecting cell expansion were performed on wild-type roots. Whole mount tubulin immunolabeling was applied in the above roots and observations were performed by confocal microscopy. Results Cortical microtubules in all mutants showed statistically significant increased persistence against anti-microtubule drugs, compared to those of the wild-type. Furthermore, to examine if the enhanced stability of cortical microtubules was due to reduced cellulose biosynthesis or to suppression of cell expansion, treatments of wild-type roots with 2,6-dichlorobenzonitrile (DCB) and Congo red were performed. After these treatments, cortical microtubules appeared more resistant to oryzalin, than in the control. Conclusions According to these findings, it may be concluded that inhibition of cell expansion, irrespective of the cause, results in increased microtubule stability in A. thaliana root. In addition, cell expansion does not only rely on cortical microtubule orientation but also plays a regulatory role in microtubule dynamics, as well. Various hypotheses may explain the increased cortical microtubule stability under decreased cell expansion such as the role of cell wall sensors and the presence of less dynamic cortical microtubules.

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