scholarly journals Upstream domains of the Xenopus laevis rDNA promoter are revealed in microinjected oocytes.

1986 ◽  
Vol 6 (4) ◽  
pp. 1228-1234 ◽  
Author(s):  
J Windle ◽  
B Sollner-Webb
Keyword(s):  
Xenopus Laevis ◽  
Dna Sequences ◽  
Promoter Region ◽  
Core Promoter ◽  
Rrna Genes ◽  
Deletion Mutants ◽  
Low Concentrations ◽  
Upstream Promoter ◽  
Polymerase I ◽  
Vector Dna

The DNA sequences involved in promoting transcription of the Xenopus laevis rRNA genes were determined by microinjecting a series of deletion mutants into oocyte nuclei. A very small promoter region is sufficient to direct efficient transcription when templates are microinjected at high rDNA concentration, since 5'delta- 9 and 3'delta +6 templates are fully active. However, as the concentration of injected template is decreased, an increasing requirement for upstream domains, extending to nucleotide approximately -170, is observed. The major downstream border of the required region does not change. This apparently expanding 5' promoter border results from the fact that, as the rDNA concentration is decreased, transcription from templates lacking the upstream promoter domain falls off much more sharply than does transcription from a complete promoter. In fact, the deleted promoters are virtually inactive below a threshold rDNA concentration. It is indeed the rDNA concentration that is important, for coinjected vector DNA does not increase the level of transcription obtained from low concentrations of the 5' deletions. From these data we conclude that polymerase I transcription factors can recognize and initiate transcription from a small core promoter domain, but that sequences extending upstream to nucleotide approximately -170 increase the efficiency of initiation. A model is presented that could account for these results.

Upstream domains of the Xenopus laevis rDNA promoter are revealed in microinjected oocytes

1986 ◽  
Vol 6 (4) ◽  
pp. 1228-1234
Author(s):  
J Windle ◽  
B Sollner-Webb
Keyword(s):  
Xenopus Laevis ◽  
Dna Sequences ◽  
Promoter Region ◽  
Core Promoter ◽  
Rrna Genes ◽  
Deletion Mutants ◽  
Low Concentrations ◽  
Upstream Promoter ◽  
Polymerase I ◽  
Vector Dna

The DNA sequences involved in promoting transcription of the Xenopus laevis rRNA genes were determined by microinjecting a series of deletion mutants into oocyte nuclei. A very small promoter region is sufficient to direct efficient transcription when templates are microinjected at high rDNA concentration, since 5'delta- 9 and 3'delta +6 templates are fully active. However, as the concentration of injected template is decreased, an increasing requirement for upstream domains, extending to nucleotide approximately -170, is observed. The major downstream border of the required region does not change. This apparently expanding 5' promoter border results from the fact that, as the rDNA concentration is decreased, transcription from templates lacking the upstream promoter domain falls off much more sharply than does transcription from a complete promoter. In fact, the deleted promoters are virtually inactive below a threshold rDNA concentration. It is indeed the rDNA concentration that is important, for coinjected vector DNA does not increase the level of transcription obtained from low concentrations of the 5' deletions. From these data we conclude that polymerase I transcription factors can recognize and initiate transcription from a small core promoter domain, but that sequences extending upstream to nucleotide approximately -170 increase the efficiency of initiation. A model is presented that could account for these results.

scholarly journals Molecular Characterization of the eis Promoter of Mycobacterium tuberculosis

2004 ◽  
Vol 186 (16) ◽  
pp. 5410-5417 ◽  
Author(s):  
Esteban A. Roberts ◽  
Amanda Clark ◽  
Sarah McBeth ◽  
Richard L. Friedman
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Promoter Region ◽  
Core Promoter ◽  
Random Mutagenesis ◽  
Core Promoter Region ◽  
E Coli ◽  
The Core ◽  
Upstream Promoter ◽  
Promoter Deletion Analysis ◽  
Group A

ABSTRACT To further understand Mycobacterium tuberculosis pathogenesis, the regulation of potential virulence genes needs to be investigated. The eis gene of M. tuberculosis H37Rv enhances the intracellular survival of Mycobacterium smegmatis, which does not contain eis, within macrophages (J. Wei, J. L. Dahl, J. W. Moulder, E. A. Roberts, P. O'Gaora, D. B. Young, and R. L. Friedman, J. Bacteriol. 182:377-384, 2000). Experiments were done to characterize the eis promoter in M. smegmatis and M. tuberculosis H37Ra. The putative −10 and −35 regions matched the Escherichia coli σ70 consensus 67 and 83%, respectively, making it a group A/SigA-like mycobacterial promoter. Expression of site-directed variants of the core promoter region, determined by flow cytometry using gfp as a reporter, showed that the putative −10 region is essential for eis expression. In addition, site-directed alteration of the eis promoter to the consensus E. coli σ70 promoter elements increased gfp transcription to levels similar to that driven by the heat shock promoter, phsp60, of Mycobacterium bovis BCG. Upstream promoter deletion analysis showed that a 200- and 412-bp region of the promoter was necessary for maximum expression of gfp in M. smegmatis and M. tuberculosis H37Ra, respectively. Random mutagenesis of the 412-bp eis promoter, using a catechol 2,3-dioxygenase screen and activity assay, defined nucleotides upstream of the core promoter region that are essential to eis expression in both M. smegmatis and M. tuberculosis H37Ra, including a region homologous to a DinR cis element.

Two distant and precisely positioned domains promote transcription of Xenopus laevis rRNA genes: analysis with linker-scanning mutants

1986 ◽  
Vol 6 (12) ◽  
pp. 4585-4593
Author(s):  
J J Windle ◽  
B Sollner-Webb
Keyword(s):  
Xenopus Laevis ◽  
Promoter Activity ◽  
Promoter Sequence ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Unexpected Result ◽  
Base Pairs ◽  
S1 Nuclease ◽  
Transcriptional Initiation ◽  
Upstream Promoter

To examine the internal organization of the promoter of the Xenopus laevis rRNA gene, we constructed a series of linker-scanning mutants that traverse the rDNA initiation region. The mutant genes, which have 3 to 11 clustered base substitutions set within an otherwise unaltered rDNA promoter sequence, were injected into Xenopus oocyte nuclei, and their transcriptional capacity was assessed by S1 nuclease analysis of the resultant RNA. The data demonstrate that there are two essential promoter domains, the distal boundaries of which coincide with the promoter boundaries established previously by analysis of 5' and 3' deletion mutants. The upstream promoter domain is relatively small and extends from residues ca. -140 to -128. The downstream domain is considerably larger, encompassing residues ca. -36 to +10, and exactly corresponds in both size and position to the mammalian minimal promoter region. The Xenopus rDNA sequence between these two essential domains has a much smaller effect on the level of transcriptional initiation. In light of the fact that a large portion of this intervening region consists of a segment (residues -114 to -72) that is duplicated many times in the upstream spacer to form an rDNA enhancer sequence, it is noteworthy that a "-115/-77 linker scanner," in which virtually this entire segment is replaced by a polylinker sequence, has full promoter activity in the injected Xenopus borealis oocytes. Analysis of a parallel series of spacing change linker-scanning mutants revealed the unexpected result that the relative positions of the upstream and downstream promoter domains are very critical: all spacing alterations of more than 2 base pairs within this 100-base-pair region virtually abolish promoter activity. We conclude that the factors that bind to these two distant promoter domains must interact in a very precise stereospecific manner.

scholarly journals Two distant and precisely positioned domains promote transcription of Xenopus laevis rRNA genes: analysis with linker-scanning mutants.

1986 ◽  
Vol 6 (12) ◽  
pp. 4585-4593 ◽  
Author(s):  
J J Windle ◽  
B Sollner-Webb
Keyword(s):  
Xenopus Laevis ◽  
Promoter Activity ◽  
Promoter Sequence ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Unexpected Result ◽  
Base Pairs ◽  
S1 Nuclease ◽  
Transcriptional Initiation ◽  
Upstream Promoter

To examine the internal organization of the promoter of the Xenopus laevis rRNA gene, we constructed a series of linker-scanning mutants that traverse the rDNA initiation region. The mutant genes, which have 3 to 11 clustered base substitutions set within an otherwise unaltered rDNA promoter sequence, were injected into Xenopus oocyte nuclei, and their transcriptional capacity was assessed by S1 nuclease analysis of the resultant RNA. The data demonstrate that there are two essential promoter domains, the distal boundaries of which coincide with the promoter boundaries established previously by analysis of 5' and 3' deletion mutants. The upstream promoter domain is relatively small and extends from residues ca. -140 to -128. The downstream domain is considerably larger, encompassing residues ca. -36 to +10, and exactly corresponds in both size and position to the mammalian minimal promoter region. The Xenopus rDNA sequence between these two essential domains has a much smaller effect on the level of transcriptional initiation. In light of the fact that a large portion of this intervening region consists of a segment (residues -114 to -72) that is duplicated many times in the upstream spacer to form an rDNA enhancer sequence, it is noteworthy that a "-115/-77 linker scanner," in which virtually this entire segment is replaced by a polylinker sequence, has full promoter activity in the injected Xenopus borealis oocytes. Analysis of a parallel series of spacing change linker-scanning mutants revealed the unexpected result that the relative positions of the upstream and downstream promoter domains are very critical: all spacing alterations of more than 2 base pairs within this 100-base-pair region virtually abolish promoter activity. We conclude that the factors that bind to these two distant promoter domains must interact in a very precise stereospecific manner.

Nucleotide polymorphisms in three genes support host and geographic speciation in tree pathogens belonging toGremmeniellaspp.

2002 ◽  
Vol 80 (11) ◽  
pp. 1151-1159 ◽  
Author(s):  
M Dusabenyagasani ◽  
G Laflamme ◽  
R C Hamelin
Keyword(s):  
North America ◽  
Dna Sequences ◽  
North American ◽  
Abies Balsamea ◽  
Host Specialization ◽  
Rrna Genes ◽  
Nucleotide Polymorphisms ◽  
Group I ◽  
Geographic Separation ◽  
Pinus Spp

We detected nucleotide polymorphisms within the genus Gremmeniella in DNA sequences of β-tubulin, glyceraldehyde phosphate dehydrogenase, and mitochondrial small subunit rRNA (mtSSU rRNA) genes. A group-I intron was present in strains originating from fir (Abies spp.) in the mtSSU rRNA locus. This intron in the mtSSU rRNA locus of strains isolated from Abies sachalinensis (Fridr. Schmidt) M.T. Mast in Asia was also found in strains isolated from Abies balsamea (L.) Mill. in North America. Phylogenetic analyses yielded trees that grouped strains by host of origin with strong branch support. Asian strains of Gremmeniella abietina (Lagerberg) Morelet var. abietina isolated from fir (A. sachalinensis) were more closely related to G. abietina var. balsamea from North America, which is found on spruce (Picea spp.) and balsam fir, and European and North American races of G. abietina var. abietina from pines (Pinus spp.) were distantly related. Likewise, North American isolates of Gremmeniella laricina (Ettinger) O. Petrini, L.E. Petrini, G. Laflamme, & G.B. Ouellette, a pathogen of larch, was more closely related to G. laricina from Europe than to G. abietina var. abietina from North America. These data suggest that host specialization might have been the leading evolutionary force shaping Gremmeniella spp., with geographic separation acting as a secondary factor.Key words: Gremmeniella, geographic separation, host specialization, mitochondrial rRNA, nuclear genes.

scholarly journals Correlation of methylation status in MTHFR promoter region with recurrent pregnancy loss

2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Mai Mahmoud Shaker ◽  
Taghreed Abdelmoniem shalabi ◽  
Khalda said Amr
Keyword(s):  
Dna Methylation ◽  
Dna Sequences ◽  
Promoter Region ◽  
Pregnancy Loss ◽  
Recurrent Pregnancy Loss ◽  
Methylation Status ◽  
Control Group ◽  
Case Group ◽  
Methyl Radical ◽  
Fertile Women

Abstract Background DNA methylation is an epigenetic process for modifying transcription factors in various genes. Methylenetetrahydrofolate reductase (MTHFR) stimulates synthesis of methyl radical in the homocysteine cycle and delivers methyl groups needed in DNA methylation. Furthermore, numerous studies have linked gene polymorphisms of this enzyme with a larger risk of recurrent pregnancy loss (RPL), yet scarce information is available concerning the association between epigenetic deviations in this gene and RPL. Hypermethylation at precise DNA sequences can function as biomarkers for a diversity of diseases. We aimed by this study to evaluate the methylation status of the promoter region of MTHFR gene in women with RPL compared to healthy fertile women. It is a case–control study. Hundred RPL patients and hundred healthy fertile women with no history of RPL as controls were recruited. MTHFR C677T was assessed by polymerase chain reaction-restriction fragment length polymorphism (RFLP). Quantitative evaluation of DNA methylation was performed by high-resolution melt analysis by real-time PCR. Results The median of percentage of MTHFR promoter methylation in RPL cases was 6.45 [0.74–100] vs. controls was 4.50 [0.60–91.7], P value < 0.001. In the case group, 57 hypermethylated and 43 normo-methylated among RPL patients vs. 40 hypermethylated and 60 normo-methylated among controls, P< 0.005. Frequency of T allele in C677T MTHFR gene among RPL patients was 29% vs. 23% among the control group; C allele vs. T allele: odds ratio (OR) = 1.367 (95% confidence interval (CI) 0.725–2.581). Conclusion Findings suggested a significant association between hypermethylation of the MTHFR promoter region in RPL patients compared to healthy fertile women.

scholarly journals The Ribosomal Gene Loci—The Power behind the Throne

Genes ◽  
2021 ◽  
Vol 12 (5) ◽  
pp. 763
Author(s):  
Konstantin I. Panov ◽  
Katherine Hannan ◽  
Ross D. Hannan ◽  
Nadine Hein
Keyword(s):  
Genome Stability ◽  
Cell Cycle Progression ◽  
Global Gene Expression ◽  
Cell Types ◽  
Ribosomal Gene ◽  
Cellular Growth ◽  
Rrna Genes ◽  
Dynamic Regulation ◽  
Pol I ◽  
Polymerase I

Nucleoli form around actively transcribed ribosomal RNA (rRNA) genes (rDNA), and the morphology and location of nucleolus-associated genomic domains (NADs) are linked to the RNA Polymerase I (Pol I) transcription status. The number of rDNA repeats (and the proportion of actively transcribed rRNA genes) is variable between cell types, individuals and disease state. Substantial changes in nucleolar morphology and size accompanied by concomitant changes in the Pol I transcription rate have long been documented during normal cell cycle progression, development and malignant transformation. This demonstrates how dynamic the nucleolar structure can be. Here, we will discuss how the structure of the rDNA loci, the nucleolus and the rate of Pol I transcription are important for dynamic regulation of global gene expression and genome stability, e.g., through the modulation of long-range genomic interactions with the suppressive NAD environment. These observations support an emerging paradigm whereby the rDNA repeats and the nucleolus play a key regulatory role in cellular homeostasis during normal development as well as disease, independent of their role in determining ribosome capacity and cellular growth rates.

scholarly journals The core promoter region of the tumor necrosis factor alpha gene confers phorbol ester responsiveness to upstream transcriptional activators.

1992 ◽  
Vol 12 (3) ◽  
pp. 1352-1356 ◽  
Author(s):  
D C Leitman ◽  
E R Mackow ◽  
T Williams ◽  
J D Baxter ◽  
B L West
Keyword(s):  
Transcription Factors ◽  
Tumor Necrosis Factor ◽  
Promoter Region ◽  
Tumor Necrosis ◽  
Core Promoter ◽  
Core Promoter Region ◽  
Necrosis Factor Alpha ◽  
The Core ◽  
Factor Alpha ◽  
Necrosis Factor

Activators of protein kinase C, such as 12-O-tetradecanoylphorbol 13-acetate (TPA), are known to regulate the expression of many genes, including the tumor necrosis factor alpha (TNF) gene, by affecting the level or activity of upstream transcription factors. To investigate the mechanism whereby TPA activates the TNF promoter, a series of 5'-deletion mutants of the human TNF promoter linked to chloramphenicol acetyltransferase was transfected into U937 human promonocytic cells. TPA produced a 7- to 11-fold activation of all TNF promoters tested, even those promoters truncated to contain only the core promoter with no upstream enhancer elements. The proximal TNF promoter containing only 28 nucleotides upstream and 10 nucleotides downstream of the RNA start site confers TPA activation to a variety of unrelated upstream enhancer elements and transcription factors, including Sp1, CTF/NF1, cyclic AMP-response element, GAL-E1a, and GAL-VP16. The level of activation by TPA depends on the TATA box structure, since the TPA response is greater in promoters containing the sequence TATAAA than in those containing TATTAA or TATTTA. These findings suggest that the core promoter region is a target for gene regulation by second-messenger pathways.

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