scholarly journals A nucleosome positioned in the distal promoter region activates transcription of the human U6 gene.

1997 ◽  
Vol 17 (8) ◽  
pp. 4397-4405 ◽  
Author(s):  
W Stünkel ◽  
I Kober ◽  
K H Seifart
Keyword(s):  
Micrococcal Nuclease ◽  
Deletion Mutants ◽  
Dna Fragments ◽  
Wild Type ◽  
Sequence Element ◽  
Proximal Sequence Element ◽  
Experimental Approaches ◽  
Type Gene ◽  
Distal Promoter ◽  
U6 Gene

To investigate the consequences of chromatin reconstitution for transcription of the human U6 gene, we assembled nucleosomes on both plasmids and linear DNA fragments containing the U6 gene. Initial experiments with DNA fragments revealed that U6 sequences located between the distal sequence element (DSE) and the proximal sequence element (PSE) lead to the positioning of a nucleosome partially encompassing these promoter elements. Furthermore, indirect end-labelling analyses of the reconstituted U6 wild-type plasmids showed strong micrococcal nuclease cuts near the DSE and PSE, indicating that a nucleosome is located between these elements. To investigate the influence that nucleosomes exert on U6 transcription, we used two different experimental approaches for chromatin reconstitution, both of which resulted in the observation that transcription of the U6 wild-type gene was enhanced after chromatin assembly. To ensure that the facilitated transcription of the nucleosomal templates is in fact due to a positioned nucleosome, we constructed mutants of the U6 gene in which the sequences between the DSE and PSE were progressively deleted. In contrast to what was observed with the wild-type genes, transcription of these deletion mutants was significantly inhibited when they were packaged into nucleosomes.

scholarly journals Transcription of the sea urchin U6 gene in vitro requires a TATA-like box, a proximal sequence element, and sea urchin USF, which binds an essential E box.

1994 ◽  
Vol 14 (3) ◽  
pp. 2191-2200 ◽  
Author(s):  
J M Li ◽  
R A Parsons ◽  
W F Marzluff
Keyword(s):  
Sea Urchin ◽  
Rna Polymerase Iii ◽  
Nuclear Extract ◽  
Major Protein ◽  
In Vitro Mutagenesis ◽  
Flanking Sequence ◽  
Sequence Element ◽  
Proximal Sequence Element ◽  
U6 Gene

The tandemly repeated gene set encoding the sea urchin U6 gene has been cloned from the sea urchin Strongylocentrotus purpuratus. The U6 gene is transcribed by RNA polymerase III in a sea urchin nuclear extract. Like that of the vertebrate U6 genes, transcription of the sea urchin U6 gene does not require any internal sequences or 3' sequences but requires only 5' flanking sequences. Only 88 nucleotides of 5' flanking sequence are required for maximal expression in vitro. Mutagenesis experiments demonstrated the requirement for three elements, a CACGTG element at -80, a proximal sequence element at about -55, and the TATA-like box at -25. The major protein in sea urchin extracts that interacts with the CACGTG element is sea urchin USF, and immunodepletion of sea urchin USF greatly reduces transcription. The USF binding site in the U6 gene is highly homologous (11 of 13 nucleotides) with the USF binding sites found in the promoter of the S. purpuratus spec genes.

scholarly journals PFP1, a Gene Encoding an Epc-N Domain-Containing Protein, Is Essential for Pathogenicity of the Barley Pathogen Rhynchosporium commune

2014 ◽  
Vol 13 (8) ◽  
pp. 1026-1035 ◽  
Author(s):  
Sylvia Siersleben ◽  
Daniel Penselin ◽  
Claudia Wenzel ◽  
Sylvie Albert ◽  
Wolfgang Knogge
Keyword(s):  
Upstream Region ◽  
Insertion Mutagenesis ◽  
Deletion Mutants ◽  
Wild Type ◽  
Homologous Proteins ◽  
Gene Encoding ◽  
Fungal Virulence ◽  
Content Type ◽  
Wild Type Level ◽  
Type Gene

ABSTRACTScald caused byRhynchosporium communeis an important foliar disease of barley. Insertion mutagenesis ofR. communegenerated a nonpathogenic fungal mutant which carries the inserted plasmid in the upstream region of a gene namedPFP1. The characteristic feature of the gene product is an Epc-N domain. This motif is also found in homologous proteins shown to be components of histone acetyltransferase (HAT) complexes of fungi and animals. Therefore, PFP1 is suggested to be the subunit of a HAT complex inR. communewith an essential role in the epigenetic control of fungal pathogenicity. TargetedPFP1disruption also yielded nonpathogenic mutants which showed wild-type-like growthex planta, except for the occurrence of hyphal swellings. Complementation of the deletion mutants with the wild-type gene reestablished pathogenicity and suppressed the hyphal swellings. However, despite wild-type-levelPFP1expression, the complementation mutants did not reach wild-type-level virulence. This indicates that the function of the protein complex and, thus, fungal virulence are influenced by a position-affected long-range control ofPFP1expression.

scholarly journals Transcription of the sea urchin U6 gene in vitro requires a TATA-like box, a proximal sequence element, and sea urchin USF, which binds an essential E box

1994 ◽  
Vol 14 (3) ◽  
pp. 2191-2200
Author(s):  
J M Li ◽  
R A Parsons ◽  
W F Marzluff
Keyword(s):  
Sea Urchin ◽  
Rna Polymerase Iii ◽  
Nuclear Extract ◽  
Major Protein ◽  
In Vitro Mutagenesis ◽  
Flanking Sequence ◽  
Sequence Element ◽  
Proximal Sequence Element ◽  
U6 Gene

The tandemly repeated gene set encoding the sea urchin U6 gene has been cloned from the sea urchin Strongylocentrotus purpuratus. The U6 gene is transcribed by RNA polymerase III in a sea urchin nuclear extract. Like that of the vertebrate U6 genes, transcription of the sea urchin U6 gene does not require any internal sequences or 3' sequences but requires only 5' flanking sequences. Only 88 nucleotides of 5' flanking sequence are required for maximal expression in vitro. Mutagenesis experiments demonstrated the requirement for three elements, a CACGTG element at -80, a proximal sequence element at about -55, and the TATA-like box at -25. The major protein in sea urchin extracts that interacts with the CACGTG element is sea urchin USF, and immunodepletion of sea urchin USF greatly reduces transcription. The USF binding site in the U6 gene is highly homologous (11 of 13 nucleotides) with the USF binding sites found in the promoter of the S. purpuratus spec genes.

scholarly journals Identification and Topological Arrangement ofDrosophila Proximal Sequence Element (PSE)-Binding Protein Subunits That Contact the PSEs of U1 and U6 Small Nuclear RNA Genes

1998 ◽  
Vol 18 (3) ◽  
pp. 1570-1579 ◽  
Author(s):  
Yan Wang ◽  
William E. Stumph
Keyword(s):  
Rna Polymerase ◽  
Rna Polymerase Ii ◽  
Sea Urchins ◽  
Binding Protein ◽  
Rna Polymerase Iii ◽  
Specific Protein ◽  
Cross Linking ◽  
Sequence Element ◽  
Proximal Sequence Element ◽  
U6 Gene

ABSTRACT Most small nuclear RNAs (snRNAs) are synthesized by RNA polymerase II, but U6 and a few others are synthesized by RNA polymerase III. Transcription of snRNA genes by either polymerase is dependent on a proximal sequence element (PSE) located upstream of position −40 relative to the transcription start site. In contrast to findings in vertebrates, sea urchins, and plants, the RNA polymerase specificity ofDrosophila snRNA genes is intrinsically encoded in the PSE sequence itself. We have investigated the differential interaction of the Drosophila melanogaster PSE-binding protein (DmPBP) with U1 and U6 gene PSEs. By using a site specific protein-DNA photo-cross-linking assay, we identified three polypeptide subunits of DmPBP with apparent molecular masses of 95, 49, and 45 kDa that are in close proximity to the DNA and two additional putative polypeptides of 230 and 52 kDa that may be integral to the complex. The 95-kDa subunit cross-linked at positions spanning the entire length of the PSE, but the 49- and 45-kDa subunits cross-linked only to the 3′ half of the PSE. The same polypeptides cross-linked to both the U1 and U6 PSE sequences. However, there were significant differences in the cross-linking patterns of these subunits at a subset of the phosphate positions, depending on whether binding was to a U1 or U6 gene PSE. These data suggest that RNA polymerase specificity is associated with distinct modes of interaction of DmPBP with the DNA at U1 and U6 promoters.

scholarly journals Identification ofvib-1, a Locus Involved in Vegetative Incompatibility Mediated byhet-cinNeurospora crassa

Genetics ◽  
2002 ◽  
Vol 162 (1) ◽  
pp. 89-101 ◽  
Author(s):  
Qijun Xiang ◽  
N Louise Glass
Keyword(s):  
Acid Phosphatase ◽  
Recognition System ◽  
Deletion Mutants ◽  
Wild Type ◽  
Vegetative Incompatibility ◽  
Death Rates ◽  
Self Recognition ◽  
Allelic Differences ◽  
Native Gel ◽  
Type Strains

AbstractA non-self-recognition system called vegetative incompatibility is ubiquitous in filamentous fungi and is genetically regulated by het loci. Different fungal individuals are unable to form viable heterokaryons if they differ in allelic specificity at a het locus. To identify components of vegetative incompatibility mediated by allelic differences at the het-c locus of Neurospora crassa, we isolated mutants that suppressed phenotypic aspects of het-c vegetative incompatibility. Three deletion mutants were identified; the deletions overlapped each other in an ORF named vib-1 (vegetative incompatibility blocked). Mutations in vib-1 fully relieved growth inhibition and repression of conidiation conferred by het-c vegetative incompatibility and significantly reduced hyphal compartmentation and death rates. The vib-1 mutants displayed a profuse conidiation pattern, suggesting that VIB-1 is a regulator of conidiation. VIB-1 shares a region of similarity to PHOG, a possible phosphate nonrepressible acid phosphatase in Aspergillus nidulans. Native gel analysis of wild-type strains and vib-1 mutants indicated that vib-1 is not the structural gene for nonrepressible acid phosphatase, but rather may regulate nonrepressible acid phosphatase activity.

The DNA Binding Protein Rfg1 Is a Repressor of Filamentation inCandida albicans

Genetics ◽  
2001 ◽  
Vol 157 (4) ◽  
pp. 1503-1512 ◽  
Author(s):  
Roy A Khalaf ◽  
Richard S Zitomer
Keyword(s):  
Dna Binding ◽  
Binding Protein ◽  
Hyphal Growth ◽  
Dna Binding Protein ◽  
Homozygous Deletion ◽  
Wild Type ◽  
Pathogenic Yeast ◽  
Repression Complex ◽  
Type Gene ◽  
Repression Domain

AbstractWe have identified a repressor of hyphal growth in the pathogenic yeast Candida albicans. The gene was originally cloned in an attempt to characterize the homologue of the Saccharomyces cerevisiae Rox1, a repressor of hypoxic genes. Rox1 is an HMG-domain, DNA binding protein with a repression domain that recruits the Tup1/Ssn6 general repression complex to achieve repression. The C. albicans clone also encoded an HMG protein that was capable of repression of a hypoxic gene in a S. cerevisiae rox1 deletion strain. Gel retardation experiments using the purified HMG domain of this protein demonstrated that it was capable of binding specifically to a S. cerevisiae hypoxic operator DNA sequence. These data seemed to indicate that this gene encoded a hypoxic repressor. However, surprisingly, when a homozygous deletion was generated in C. albicans, the cells became constitutive for hyphal growth. This phenotype was rescued by the reintroduction of the wild-type gene on a plasmid, proving that the hyphal growth phenotype was due to the deletion and not a secondary mutation. Furthermore, oxygen repression of the hypoxic HEM13 gene was not affected by the deletion nor was this putative ROX1 gene regulated positively by oxygen as is the case for the S. cerevisiae gene. All these data indicate that this gene, now designated RFG1 for Repressor of Filamentous Growth, is a repressor of genes required for hyphal growth and not a hypoxic repressor.

scholarly journals Isolation of a gene required for programmed initiation of development by Aspergillus nidulans.

1992 ◽  
Vol 12 (9) ◽  
pp. 3827-3833 ◽  
Author(s):  
T H Adams ◽  
W A Hide ◽  
L N Yager ◽  
B N Lee
Keyword(s):  
Life Cycle ◽  
Aspergillus Nidulans ◽  
Optimal Growth ◽  
Growth Conditions ◽  
Nutrient Deprivation ◽  
Deletion Mutants ◽  
Wild Type ◽  
Microbial Development ◽  
Type Strains ◽  
Posttranscriptional Level

In contrast to many other cases in microbial development, Aspergillus nidulans conidiophore production initiates primarily as a programmed part of the life cycle rather than as a response to nutrient deprivation. Mutations in the acoD locus result in "fluffy" colonies that appear to grow faster than the wild type and proliferate as undifferentiated masses of vegetative cells. We show that unlike wild-type strains, acoD deletion mutants are unable to make conidiophores under optimal growth conditions but can be induced to conidiate when growth is nutritionally limited. The requirement for acoD in conidiophore development occurs prior to activation of brlA, a primary regulator of development. The acoD transcript is present both in vegetative hyphae prior to developmental induction and in developing cultures. However, the effects of acoD mutations are detectable only after developmental induction. We propose that acoD activity is primarily controlled at the posttranscriptional level and that it is required to direct developmentally specific changes that bring about growth inhibition and activation of brlA expression to result in conidiophore development.

scholarly journals Occurrence of Repeat Induced Point Mutation in Long Segmental Duplications of Neurospora

Genetics ◽  
1997 ◽  
Vol 147 (1) ◽  
pp. 125-136 ◽  
Author(s):  
David D Perkins ◽  
Brian S Margolin ◽  
Eric U Selker ◽  
S D Haedo
Keyword(s):  
Point Mutation ◽  
Sexual Cycle ◽  
Segmental Duplications ◽  
Wild Type ◽  
Single Chain ◽  
Base Pairs ◽  
Gene Size ◽  
Type Gene ◽  
Repeat Induced Point Mutation

Abstract Previous studies of repeat induced point mutation (RIP) have typically involved gene-size duplications resulting from insertion of transforming DNA at ectopic chromosomal positions. To ascertain whether genes in larger duplications are subject to RIP, progeny were examined from crosses heterozygous for long segmental duplications obtained using insertional or quasiterminal translocations. Of 17 distinct mutations from crossing 11 different duplications, 13 mapped within the segment that was duplicated in the parent, one was closely linked, and three were unlinked. Half of the mutations in duplicated segments were at previously unknown loci. The mutations were recessive and were expressed both in haploid and in duplication progeny from Duplication × Normal, suggesting that both copies of the wild-type gene had undergone RIP. Seven transition mutations characteristic of RIP were found in 395 base pairs (bp) examined in one ro-11 allele from these crosses and three were found in ~750 bp of another. A single chain-terminating C to T mutation was found in 800 bp of arg-6. RIP is thus responsible. These results are consistent with the idea that the impaired fertility that is characteristic of segmental duplications is due to inactivation by RIP of genes needed for progression through the sexual cycle.

scholarly journals Identification and Characterization of a Peptidoglycan Hydrolase, MurA, of Listeria monocytogenes, a Muramidase Needed for Cell Separation

2003 ◽  
Vol 185 (23) ◽  
pp. 6801-6808 ◽  
Author(s):  
Shannon A. Carroll ◽  
Torsten Hain ◽  
Ulrike Technow ◽  
Ayub Darji ◽  
Philippos Pashalidis ◽  
...  
Keyword(s):  
Cell Wall ◽  
Listeria Monocytogenes ◽  
Cell Separation ◽  
Bacterial Species ◽  
Surface Protein ◽  
Wild Type ◽  
Terminal Domain ◽  
Cell Wall Hydrolase ◽  
Characteristic Features ◽  
Type Gene

ABSTRACT A novel cell wall hydrolase encoded by the murA gene of Listeria monocytogenes is reported here. Mature MurA is a 66-kDa cell surface protein that is recognized by the well-characterized L. monocytogenes-specific monoclonal antibody EM-7G1. MurA displays two characteristic features: (i) an N-terminal domain with homology to muramidases from several gram-positive bacterial species and (ii) four copies of a cell wall-anchoring LysM repeat motif present within its C-terminal domain. Purified recombinant MurA produced in Escherichia coli was confirmed to be an authentic cell wall hydrolase with lytic properties toward cell wall preparations of Micrococcus lysodeikticus. An isogenic mutant with a deletion of murA that lacked the 66-kDa cell wall hydrolase grew as long chains during exponential growth. Complementation of the mutant strain by chromosomal reintegration of the wild-type gene restored expression of this murein hydrolase activity and cell separation levels to those of the wild-type strain. Studies reported herein suggest that the MurA protein is involved in generalized autolysis of L. monocytogenes.

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