scholarly journals REGULATION OF PHOSPHATE METABOLISM IN NEUROSPORA CRASSA: IDENTIFICATION OF THE STRUCTURAL GENE FOR REPRESSIBLE ACID PHOSPHATASE

Genetics ◽  
1976 ◽  
Vol 84 (2) ◽  
pp. 183-192
Author(s):  
Robert E Nelson ◽  
John F Lehman ◽  
Robert L Metzenberg
Keyword(s):  
Acid Phosphatase ◽  
Neurospora Crassa ◽  
Structural Gene ◽  
Group Iv ◽  
Structural Genes ◽  
Wild Type ◽  
Linkage Groups ◽  
Michaelis Constant ◽  
The Right

ABSTRACT A mutant of Neurospora crassa with an altered repressible acid phosphatase has been isolated. The enzyme is much more thermolabile than that of wild type, and has an increased Michaelis constant. Tests of allelic interactions (in partial diploids) and in vitro mixing experiments were consistent with the mutation being in the structural gene for the enzyme. This gene, pho-3, was found to be located in the right arm of Linkage Group IV (LG IV). Thus, pho-3 and the structural gene for repressible alkaline phosphatase, pho-2 (LG V), map in separate linkage groups and cannot be part of the same operon. Neither of these structural genes is linked to the known regulatory genes, nuc-1 (LG I), nuc-2 (LG II), and preg (LG II).

scholarly journals COMPLEMENTATION ANALYSIS OF LINKED CIRCADIAN CLOCK MUTANTS OF NEUROSPORA CRASSA

Genetics ◽  
1976 ◽  
Vol 82 (1) ◽  
pp. 9-17 ◽  
Author(s):  
Jerry F Feldman ◽  
Marian N Hoyle
Keyword(s):  
Linkage Group ◽  
Circadian Clock ◽  
Neurospora Crassa ◽  
Period Length ◽  
Complementation Analysis ◽  
Wild Type ◽  
The Right

ABSTRACT A fourth mutant of Neurospora crassa, designated frq-4, has been isolated in which the period length of the circadian conidiation rhythm is shortened to 19.3 ± 0.3 hours. This mutant is tightly linked to the three previously isolated frq mutants, and all four map to the right arm of linkage group VII about 10 map units from the centromere. Complementation tests suggest, but do not prove, that all four mutations are allelic, since each of the four mutants is co-dominant with the frq  + allele—i.e., heterokaryons have period lengths intermediate between the mutant and wild-type—and since heterokaryons between pairs of mutants also have period lengths intermediate between those of the two mutants.

Production of the CYS3 regulator, a bZIP DNA-binding protein, is sufficient to induce sulfur gene expression in Neurospora crassa

1992 ◽  
Vol 12 (4) ◽  
pp. 1568-1577
Author(s):  
J V Paietta
Keyword(s):  
Gene Expression ◽  
Neurospora Crassa ◽  
Structural Gene ◽  
Control Point ◽  
Regulatory Protein ◽  
Negative Regulator ◽  
Temperature Sensitive ◽  
Cys3 Protein

The cys-3+ gene of Neurospora crassa encodes a bZIP (basic region-leucine zipper) regulatory protein that is essential for sulfur structural gene expression (e.g., ars-1+). Nuclear transcription assays confirmed that cys-3+ was under sulfur-regulated transcriptional control and that cys-3+ transcription was constitutive in sulfur controller (scon)-negative regulator mutants. Given these results, I have tested whether expression of cys-3+ under high-sulfur (repressing) conditions was sufficient to induce sulfur gene expression. The N. crassa beta-tubulin (tub) promoter was fused to the cys-3+ coding segment and used to transform a cys-3 deletion mutant. Function of the tub::cys-3 fusion in homokaryotic transformants grown under high-sulfur conditions was confirmed by Northern (RNA) and Western immunoblot analysis. The tub::cys-3 transformants showed arylsulfatase gene expression under normally repressing high-sulfur conditions. A tub::cys-3ts fusion encoding a temperature-sensitive CYS3 protein was used to confirm that the induced structural gene expression was due to CYS3 protein function. Constitutive CYS3 production did not induce scon-2+ expression under repressing conditions. In addition, a cys-3 promoter fusion to lacZ showed that CYS3 production was sufficient to induce its own expression and provides in vivo evidence for autoregulation. Finally, an apparent inhibitory effect observed with a strain carrying a point mutation at the cys-3 locus was examined by in vitro heterodimerization studies. These results support an interpretation of CYS3 as a transcriptional activator whose regulation is a crucial control point in the signal response pathway triggered by sulfur limitation.

scholarly journals Subforms and In Vitro Reconstitution of the NAD-Reducing Hydrogenase of Alcaligenes eutrophus

1998 ◽  
Vol 180 (5) ◽  
pp. 1023-1029 ◽  
Author(s):  
Christian Massanz ◽  
Silke Schmidt ◽  
Bärbel Friedrich
Keyword(s):  
Alcaligenes Eutrophus ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Monomeric Form ◽  
Structural Genes ◽  
Wild Type ◽  
Catalytic Function ◽  
C Terminus ◽  
In Vitro Reconstitution ◽  
Reducing Activity

The cytoplasmic, NAD-reducing hydrogenase (SH) of Alcaligenes eutrophus H16 is a heterotetrameric enzyme which contains several cofactors and undergoes a complex maturation during biogenesis. HoxH is the Ni-carrying subunit, and together with HoxY it forms the hydrogenase dimer. HoxF and HoxU represent the flavin-containing diaphorase moiety, which is closely related to NADH:ubiquinone oxidoreductase and mediates NADH oxidation. A variety of mutations were introduced into the four SH structural genes to obtain mutant enzymes composed of monomeric and dimeric forms. A deletion removing most ofhoxF, hoxU, and hoxY led to the expression of a HoxH monomer derivative which was proteolytically processed at the C terminus like the wild-type polypeptide. While the hydrogenase dimer, produced by a strain deleted of hoxF andhoxU, displayed H2-dependent dye-reducing activity, the monomeric form did not mediate the activation of H2, although nickel was incorporated into HoxH. Deletion ofhoxH and hoxY led to the production of HoxFU dimers which displayed NADH:oxidoreductase activity. Mixing the hydrogenase and the diaphorase moieties in vitro reconstituted the structure and catalytic function of the SH holoenzyme.

scholarly journals Deletion of the Mycobacterium tuberculosis α-Crystallin-Like hspX Gene Causes Increased Bacterial Growth In Vivo

2006 ◽  
Vol 74 (2) ◽  
pp. 861-868 ◽  
Author(s):  
Yanmin Hu ◽  
Farahnaz Movahedzadeh ◽  
Neil G. Stoker ◽  
Anthony R. M. Coates
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Constitutive Expression ◽  
Active Role ◽  
Structural Genes ◽  
Wild Type ◽  
Major Antigen ◽  
Oxide Stress ◽  
Unusual Phenotype

ABSTRACT Hypervirulent mutants of Mycobacterium tuberculosis, whose growth rates are higher in vivo, have now been reported to have mutations in both regulatory and structural genes, but the basis for this unusual phenotype is not understood. One hypervirulence gene, dosR (devR, Rv2031c), activates transcription of approximately 50 genes in this pathogen in response to hypoxia and nitric oxide stress. The most dramatic activation (∼80-fold) is activation of the hspX (acr, Rv2031c) gene, which encodes a 16-kDa α-crystallin-like protein that is a major antigen. In this study we found that a Δacr mutant exhibited increased growth following infection of BALB/c mice in vivo and in both resting and activated macrophages in vitro (as measured by the number of CFU). The increased growth in macrophages was equal to that of a ΔdosR mutant, while introduction of a constitutively expressed hspX gene reduced the ΔdosR virulence to wild-type levels. These results suggest that the increased number of CFU of the ΔdosR mutant was largely due to loss of hspX expression. We also confirmed that constitutive expression of hspX slows growth in vitro, and we propose that hspX plays an active role in slowing the growth of M. tuberculosis in vivo immediately following infection.

scholarly journals Functional characterization of an N-terminally-truncated mitochondrial porin expressed in Neurospora crassa

2017 ◽  
Vol 63 (8) ◽  
pp. 730-738 ◽  
Author(s):  
Sabbir R. Shuvo ◽  
Uliana Kovaltchouk ◽  
Abdullah Zubaer ◽  
Ayush Kumar ◽  
William A.T. Summers ◽  
...  
Keyword(s):  
Neurospora Crassa ◽  
Outer Membrane ◽  
Functional Characterization ◽  
Helical Structure ◽  
Terminal Segment ◽  
Wild Type ◽  
Mitochondrial Porin ◽  
N Terminus

Mitochondrial porin, which forms voltage-dependent anion-selective channels (VDAC) in the outer membrane, can be folded into a 19-β-stranded barrel. The N terminus of the protein is external to the barrel and contains α-helical structure. Targeted modifications of the N-terminal region have been assessed in artificial membranes, leading to different models for gating in vitro. However, the in vivo requirements for gating and the N-terminal segment of porin are less well-understood. Using Neurospora crassa porin as a model, the effects of a partial deletion of the N-terminal segment were investigated. The protein, ΔN2-12porin, is assembled into the outer membrane, albeit at lower levels than the wild-type protein. The resulting strain displays electron transport chain deficiencies, concomitant expression of alternative oxidase, and decreased growth rates. Nonetheless, its mitochondrial genome does not contain any significant mutations. Most of the genes that are expressed in high levels in porin-less N. crassa are expressed at levels similar to that of wild type or are slightly increased in ΔN2-12porin strains. Thus, although the N-terminal segment of VDAC is required for complete function in vivo, low levels of a protein lacking part of the N terminus are able to rescue some of the defects associated with the absence of porin.

scholarly journals Heterogeneity in recombination frequencies in Neurospora crassa

1961 ◽  
Vol 2 (1) ◽  
pp. 43-62 ◽  
Author(s):  
L. C. Frost
Keyword(s):  
Neurospora Crassa ◽  
Wild Type ◽  
Linkage Groups ◽  
Genetic Determination ◽  
Wild Strains ◽  
Genetically Determined ◽  
Homogeneous Data

The available data on heterogeneity in centromere distances for a number of loci in several linkage groups are analysed and interpreted. When the crosses are grouped according to wild-type ancestry, heterogeneity is eliminated in any one group except those which consist of backcrosses or intercrosses. Abbott 4 and 12 are shown to be the source of the heterogeneity while Lindegren and probably Chilton wild strains give consistent, homogeneous distances. In a cross between Abbott 12 and Lindegren wild-types, the centromere distances of mt and asco show heterogeneous values between the spore pairs in an ascus indicating that significantly different distances are genetically determined and that the factors concerned show segregation. The genetic determination differs in the various wild strains; the data suggest that at least three factors are involved. In random spore analyses heterogeneity is present in recombination frequencies between linked markers either proximal or distal to their centromere. The mechanism by which heterogeneity in the data might arise is discussed. To obtain homogeneous data it is suggested that all markers used should be repeatedly backcrossed to the Lindegren wild-type.

scholarly journals Nonsense mutations of the ornithine decarboxylase structural gene of Neurospora crassa

1987 ◽  
Vol 7 (3) ◽  
pp. 1122-1128
Author(s):  
R H Davis ◽  
L V Hynes ◽  
P Eversole-Cire
Keyword(s):  
Neurospora Crassa ◽  
Ornithine Decarboxylase ◽  
Structural Gene ◽  
Wild Type Strain ◽  
Normal Growth ◽  
Wild Type ◽  
Carboxy Terminus ◽  
Nonsense Mutations ◽  
Polyamine Synthesis ◽  
Weak Activity

Ornithine decarboxylase (ODC) (EC 4.1.1.17) is an early enzyme of polyamine synthesis, and its activity rises quickly at the onset of growth and differentiation in most eucaryotes. Some have speculated that the enzyme protein may have a role in the synthesis of rRNA in addition to its role in catalyzing the decarboxylation of ornithine (G. D. Kuehn and V. J. Atmar, Fed. Proc. 41:3078-3083, 1982; D. H. Russell, Proc. Natl. Acad. Sci. USA 80:1318-1321, 1983). To test this possibility, we sought mutational evidence for the indispensability of the ODC protein for normal growth of Neurospora crassa. We found three new, ODC-deficient mutants that lacked ODC protein. Among these and by reversion analysis of an earlier set of mutants, we found that two ODC-deficient mutants carried nonsense mutations in the ODC structural gene, spe-1. Allele LV10 imparted a complete deficiency for enzyme activity (less than 0.006% of normal) and had no detectable ODC antigen. Allele PE4 imparted a weak activity to cells (0.1% of derepressed spe+ cultures) and encoded a lower-molecular-weight ODC subunit (Mr = 43,000) in comparison to that of the wild-type strain (Mr = 53,000). Strains carrying either mutation, like other spe-1 mutants, grew at a normal rate in exponential culture if the medium was supplemented with spermidine, the main end product of the polyamine pathway in N. crassa. Unless an antigenically silent, N-terminal fragment with an indispensable role persists in the LV10-bearing mutant, we conclude that the ODC protein has no role in the vegetative growth of this organism other than the synthesis of polyamines. The data extend earlier evidence that spe-1 is the structural gene for ODC in N. crassa. The activity found in mutants bearing allele PE4 suggests that the amino acids nearest the carboxy terminus do not contribute to the active site of the enzyme.

scholarly journals Identification of a putative structural gene for cathepsin D in Caenorhabditis elegans.

Genetics ◽  
1988 ◽  
Vol 119 (2) ◽  
pp. 355-363
Author(s):  
L A Jacobson ◽  
L Jen-Jacobson ◽  
J M Hawdon ◽  
G P Owens ◽  
M A Bolanowski ◽  
...  
Keyword(s):  
Caenorhabditis Elegans ◽  
Linkage Group ◽  
Structural Gene ◽  
Cathepsin D ◽  
Normal Growth ◽  
Mutant Gene ◽  
Wild Type ◽  
Type Activity ◽  
Group Ii ◽  
The Right

Abstract Mutants of Caenorhabditis elegans having about 10% of wild-type activity of the aspartyl protease cathepsin D have been isolated by screening. Mutant homozygotes have normal growth rates and no obvious morphological or developmental abnormalities. The mutant gene (cad-1) has been mapped to the right extremity of linkage group II. Heterozygous animals (cad-1/+) show intermediate enzyme levels and animals heterozygous for chromosomal deficiencies of the right extremity of linkage group II have 50% of wild-type activity. Cathepsin D purified from a mutant strain has a lower activity per unit mass of pure enzyme. These data suggest that cad-1 is a structural gene for cathepsin D.

The structural gene for a phosphorus-repressible phosphate permease in Neurospora crassa can complement a mutation in positive regulatory gene nuc-1

1988 ◽  
Vol 8 (3) ◽  
pp. 1376-1379
Author(s):  
B J Mann ◽  
R A Akins ◽  
A M Lambowitz ◽  
R L Metzenberg
Keyword(s):  
Neurospora Crassa ◽  
Structural Gene ◽  
Regulatory Gene ◽  
Cosmid Clone ◽  
Wild Type ◽  
Gene Encoding ◽  
Regulatory Locus

van+, a gene encoding a phosphorus-repressible phosphate permease, was isolated by its ability to complement nuc-1, a positive regulatory locus that normally regulates van+ expression. This was unexpected because the nuc-1 host already contained a resident van+ gene. Plasmids carrying van+ complemented a nuc-2 mutation as well. Probing of RNA from untransformed wild-type (nuc-1+) and constitutive (nuc-1c) strains by van+ probes indicated that levels of the van+ transcript were subject to control by nuc-1+. Probing of the same RNAs with a cosmid clone, containing approximately 15 kilobases of upstream and downstream DNA, revealed no other detectable phosphorus-regulated transcripts within this 40-kilobase region of the chromosome.

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