Cruciform DNA Sequences in Gene Promoters Can Impact Transcription upon Oxidative Modification of 2′-Deoxyguanosine

Abstract FKLF-2, a novel Krüppel-type zinc finger protein, was cloned from murine yolk sac. The deduced polypeptide sequence of 289 amino acids has 3 contiguous zinc fingers at the near carboxyl-terminal end, an amino-terminal domain characterized by its high content of alanine and proline residues and a carboxyl-terminal domain rich in serine residues. By Northern blot hybridization, the human homologue of FKLF-2 is expressed in the bone marrow and striated muscles and not in 12 other human tissues analyzed. FKLF-2 is constitutively expressed in established cell lines with an erythroid phenotype, but it is inconsistently expressed in cell lines with myeloid or lymphoid phenotypes. The expression of FKLF-2 messenger RNA (mRNA) is up-regulated after induction of mouse erythroleukemia cells. In luciferase assays, FKLF-2 activates predominantly the γ, and to a lesser degree, the ɛ and β globin gene promoters. The activation of γ gene promoter does not depend on the presence of an HS2 enhancer. FKLF-2 activates the γ promoter predominantly by interacting with the γ CACCC box, and to a lesser degree through interaction with the TATA box or its surrounding DNA sequences. FKLF-2 also activated all the other erythroid specific promoters we tested (GATA-1, glycophorin B, ferrochelatase, porphobilinogen deaminase, and 5-aminolevulinate synthase). These results suggest that in addition to globin, FKLF-2 may be involved in activation of transcription of a wide range of genes in the cells of the erythroid lineage.

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Enzymatic Protein Biopolymers as a Tool to Synthetize Eukaryotic Messenger Ribonucleic Acid (mRNA) with Uses in Vaccination, Immunotherapy and Nanotechnology

Polymers ◽

10.3390/polym12081633 ◽

2020 ◽

Vol 12 (8) ◽

pp. 1633

Author(s):

Fabiola Urbina ◽

Sebastián Morales-Pison ◽

Edio Maldonado

Keyword(s):

Transcription Factors ◽

Rna Polymerase Ii ◽

Dna Sequences ◽

Gene Promoter ◽

Test Tube ◽

Gene Promoters ◽

Human Beings ◽

Promoter Elements ◽

Messenger Ribonucleic Acid

Multi-subunit enzymes are protein biopolymers that are involved in many cellular processes. The enzyme that carries out the process of transcription of mRNAs is RNA polymerase II (RNAPII), which is a multi-subunit enzyme in eukaryotes. This protein biopolymer starts the transcription from specific sites and is positioned by transcription factors, which form a preinitiation complex (PIC) on gene promoters. To recognize and position the RNAPII and the transcription factors on the gene promoters are needed specific DNA sequences in the gene promoters, which are named promoter elements. Those gene promoter elements can vary and therefore several kinds of promoters exist, however, it appears that all promoters can use a similar pathway for PIC formation. Those pathways are discussed in this review. The in vitro transcribed mRNA can be used as vaccines to fight infectious diseases, e.g., in immunotherapy against cancer and in nanotechnology to deliver mRNA for a missing protein into the cell. We have outlined a procedure to produce an mRNA vaccine against the SARS-CoV-2 virus, which is the causing agent of the big pandemic, COVID-19, affecting human beings all over the world. The potential advantages of using eukaryotic RNAPII to synthetize large transcripts are outlined and discussed. In addition, we suggest a method to cap the mRNA at the 5′ terminus by using enzymes, which might be more effective than cap analogs. Finally, we suggest the construction of a future multi-talented RNAPII, which would be able to synthetize large mRNA and cap them in the test tube.

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Comparison of Human and Monkey Pigment Gene Promoters to Evaluate DNA Sequences Proposed to Govern L:M Cone Ratio

Normal and Defective Colour Vision ◽

10.1093/acprof:oso/9780198525301.003.0006 ◽

2003 ◽

pp. 51-60 ◽

Cited By ~ 2

Author(s):

C. McMahon ◽

J. Neitz ◽

M. Neitz

Keyword(s):

Dna Sequences ◽

Gene Promoters

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Spacer promoters are orientation-dependent activators of pre-rRNA transcription in Drosophila melanogaster

Molecular and Cellular Biology ◽

10.1128/mcb.10.9.4667-4677.1990 ◽

1990 ◽

Vol 10 (9) ◽

pp. 4667-4677

Author(s):

G Grimaldi ◽

P Fiorentini ◽

P P Di Nocera

Keyword(s):

Drosophila Melanogaster ◽

Dna Sequences ◽

Transcription Initiation ◽

Gene Promoter ◽

Large Set ◽

Gene Promoters ◽

Stimulatory Action ◽

Rrna Transcription ◽

Tandem Arrays

In Drosophila melanogaster, 240-base-pair (bp) repeats, clustered in tandem arrays within the ribosomal DNA nontranscribed spacer region, include sites of RNA polymerase I-dependent transcription initiation and elements that stimulate the rate of transcription from the downstream precursor rRNA (pre-rRNA) promoter. We have analyzed the in vivo transcriptional activity of a large set of recombinant constructs in which tandem arrays of distinct segments derived from a 240-bp repeat were inserted upstream of the pre-rRNA promoter. The results indicate that activating spacer elements are confined to a region of 70 bp. Enhancing units overlap with spacer promoters, since DNA segments that stimulate transcription at the gene promoter also efficiently drive transcription initiation. The finding that artificial spacer arrays invariably stimulate pre-rRNA transcription initiation in an orientation-dependent fashion suggest that spacer-initiated transcription is involved in the enhancement process. The minimal spacer activating segment includes a perfect copy of a core domain of the gene promoter extending from -24 to +10 flanked by poorly homologous upstream DNA sequences. Spacer and gene promoters are functionally interchangeable as activating units. However, the different combination of DNA elements within the two determines a functional hierarchy, as only the pre-rRNA promoter is responsive to the stimulatory action of upstream units.

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Domain analysis of the plant DNA-binding protein GT1a: requirement of four putative alpha-helices for DNA binding and identification of a novel oligomerization region.

Molecular and Cellular Biology ◽

10.1128/mcb.15.2.1014 ◽

1995 ◽

Vol 15 (2) ◽

pp. 1014-1020 ◽

Cited By ~ 18

Author(s):

E Lam

Keyword(s):

Dna Binding ◽

Dna Sequences ◽

Transcription Initiation ◽

Binding Protein ◽

Dna Binding Protein ◽

Cdna Clones ◽

Protein Oligomerization ◽

Gene Promoters ◽

Alpha Helices

Light is an important environmental signal that can influence diverse developmental processes in plants. Many plant nuclear genes respond to light at the level of transcription initiation. GT-1 and GT2 are nuclear factors which interact with DNA sequences in many light-responsive gene promoters. cDNA clones which encode proteins with sequence binding specificities similar to those of these two factors have been isolated. They show significant amino acid sequence similarities within three closely spaced, putative alpha-helices that were predicted by secondary structure analysis but do not show significant homologies with any other reported DNA-binding protein. In this work, N- and C-terminal deletions of tobacco GT1a were generated by in vitro transcription and translation, and their DNA-binding activities and subunit structures were studied. The results suggest that the C-terminal domain of GT1a is critical for protein oligomerization, while a region predicted to contain four closely spaced alpha-helices is required for DNA binding. Direct chemical cross-linking and gel filtration analyses of full-length and truncated derivatives of GT1a suggest that this factor can exist in solution as a homotetramer and that oligomerization is independent of DNA binding. This study thus establishes two independent functional domains in this class of eukaryotic trans-acting factors. Possible implications of the multimeric nature of GT1a in relation to the known characteristics of light-responsive promoter architecture are discussed.

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A member of the nuclear factor-1 family is involved in the pituitary repression of the human placental growth hormone genes

Biochemical Journal ◽

10.1042/bj3540387 ◽

2001 ◽

Vol 354 (2) ◽

pp. 387-395 ◽

Cited By ~ 5

Author(s):

Lisa Daly NORQUAY ◽

Yan JIN ◽

Rama Mohan SURABHI ◽

R. Daniel GIETZ ◽

Naoko TANESE ◽

...

Keyword(s):

Growth Hormone ◽

Protein Interactions ◽

Dna Sequences ◽

Human Growth ◽

Chromosome 17 ◽

Gene Promoters ◽

Nuclear Factor ◽

Sequence Elements ◽

Placental Growth Hormone ◽

Nuclear Factor 1

The human growth hormone (GH) gene family consists of five tandemly arranged and highly related genes, including the chorionic somatomammotropins (CSs), at a single locus on chromosome 17. Despite striking homologies in promoter and flanking DNA sequences, the genes within this locus have different tissue-specific patterns of expression: GH-N is expressed almost exclusively in the somatotrophs of the anterior pituitary; the remaining genes, including CS-A, are expressed in placental syncytiotrophoblast. Previously we proposed that active repression of the placental gene promoters in pituitary GC cells is mediated by upstream ‘P’ sequences and, specifically, a 263bp region containing two ‘P’ sequence elements (PSE-A and PSE-B) and corresponding factors (PSF-A and PSF-B). We have now examined the possibility that PSF-A and PSF-B are members of the nuclear factor (NF)-1 family. Transcripts of NF-1A, NF-1C and NF-1X, but not of NF-1B, were readily detected in GC cells. High-affinity binding of NF-1 to PSE-B, but not to PSE-A, was confirmed by competition of DNA–protein interactions by using NF-1 DNA elements and antibodies. Functionally, a NF-1 element was able to substitute for PSE-B as a promoter-specific repressor in GC cells after gene transfer. However, there was a difference in the magnitude of repression exerted by the NF-1 and PSF-B elements on the CS-A promoter and, with the use of mutations, this difference was shown to be consistent with variations in NF-1-binding sequences. These results indicate that PSF-B, but not PSF-A, is a member of the NF-1 family, which participates in the PSF complex and in the repression of the CS-A promoter in pituitary GC cells.

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Forkhead L2 Is Expressed in the Ovary and Represses the Promoter Activity of the Steroidogenic Acute Regulatory Gene

Endocrinology ◽

10.1210/en.2003-1141 ◽

2004 ◽

Vol 145 (7) ◽

pp. 3424-3433 ◽

Cited By ~ 136

Author(s):

Margareta D. Pisarska ◽

Jeehyeon Bae ◽

Cynthia Klein ◽

Aaron J. W. Hsueh

Keyword(s):

Dna Sequences ◽

Premature Ovarian Failure ◽

Ovarian Failure ◽

Dominant Negative ◽

Dominant Negative Effect ◽

Gene Promoters ◽

Wild Type ◽

Exact Function ◽

Steroidogenic Acute Regulatory ◽

Star Gene

Abstract Premature ovarian failure in a subgroup of women with blepharophimosis-ptosis-epicanthus inversus type 1 syndrome has been associated with nonsense mutations in the gene encoding a Forkhead transcription factor, Forkhead L2 (FOXL2). However, the exact function of FOXL2 in the ovary is unclear. We investigated the expression of FOXL2 in the mouse ovary during follicular development and maturation by RT-PCR and in situ hybridization. The FOXL2 mRNA is expressed in ovaries throughout development and adulthood and is localized to the undifferentiated granulosa cells in small and medium follicles as well as cumulus cells of preovulatory follicles. FOXL2 belongs to a group of transcription factors capable of interacting with specific DNA sequences in diverse gene promoters. With the presence of multiple putative forkhead DNA consensus sites, the promoter of the human steroidogenic acute regulatory (StAR) gene was used to test for regulation by FOXL2. Cotransfection studies revealed that wild-type FOXL2 represses the activity of the StAR promoter, and the first 95 bp upstream of the transcriptional start site of the StAR gene is sufficient for FOXL2 repression. EMSAs confirmed that FOXL2 interacts directly with this region. Analyses using FOXL2 mutants also demonstrated the importance of the entire alanine/proline-rich carboxyl terminus of FOXL2 for transcriptional repression. Furthermore, these mutations produce a protein with a dominant-negative effect that disables the transcriptional repressor activity of wild-type FOXL2. Dominant-negative mutations of FOXL2 could increase expression of StAR and other follicle differentiation genes in small and medium follicles to accelerate follicle development, resulting in increased initial recruitment of dormant follicles and thus the premature ovarian failure phenotype.

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Aptamer Embedded Arch-Cruciform DNA Assemblies on 2-D VS2 Scaffolds for Sensitive Detection of Breast Cancer Cells

Biosensors ◽

10.3390/bios11100378 ◽

2021 ◽

Vol 11 (10) ◽

pp. 378

Author(s):

Jinfeng Quan ◽

Yihan Wang ◽

Jialei Zhang ◽

Kejing Huang ◽

Xuemei Wang ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Dna Sequences ◽

Breast Cancer Cells ◽

Limit Of Detection ◽

Sensitive Detection ◽

Highly Sensitive ◽

Highly Sensitive Detection ◽

Cruciform Dna ◽

Mcf 7

Arch-cruciform DNA are self-assembled on AuNPs/VS2 scaffold as a highly sensitive and selective electrochemical biosensor for michigan cancer foundation-7 (MCF-7) breast cancer cells. In the construction, arch DNA is formed using two single-strand DNA sequences embedded with the aptamer for MCF-7 cells. In the absence of MCF-7 cells, a cruciform DNA labeled with three terminal biotin is bound to the top of arch DNA, which further combines with streptavidin-labeled horseradish peroxidase (HRP) to catalyze the hydroquinone-H2O2 reaction on the electrode surface. The presence of MCF-7 cells can release the cruciform DNA and reduce the amount of immobilized HRP, thus effectively inhibiting enzyme-mediated electrocatalysis. The electrochemical response of the sensor is negatively correlated with the concentration of MCF-7 cells, with a linear range of 10 − 1 × 105 cells/mL, and a limit of detection as low as 5 cells/mL (S/N = 3). Through two-dimensional materials and enzyme-based dual signal amplification, this biosensor may pave new ways for the highly sensitive detection of tumor cells in real samples.

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Distinct chromatin signatures of DNA hypomethylation in aging and cancer

10.1101/229476 ◽

2017 ◽

Author(s):

Raúl F. Pérez ◽

Juan Ramón Tejedor ◽

Gustavo F. Bayón ◽

Agustín F. Fernández ◽

Mario F. Fraga

Keyword(s):

Dna Methylation ◽

Dna Sequences ◽

Es Cells ◽

Gene Promoters ◽

Dna Hypomethylation ◽

Chromatin Signature ◽

Unknown Gene ◽

Genomic Regions ◽

Nih Roadmap

AbstractBackgroundCancer is an aging-associated disease but the underlying molecular links between these processes are still largely unknown. Gene promoters that become hypermethylated in aging and cancer share a common chromatin signature in ES cells. In addition, there is also global DNA hypomethylation in both processes. However, any similarities of the regions where this loss of DNA methylation occurs is currently not well characterized, nor is it known whether such regions also share a common chromatin signature in aging and cancer.ResultsTo address this issue we analysed TCGA DNA methylation data from a total of 2,311 samples, including control and cancer cases from patients with breast, kidney, thyroid, skin, brain and lung tumors and healthy blood, and integrated the results with histone, chromatin state and transcription factor binding site data from the NIH Roadmap Epigenomics and ENCODE projects. We identified 98,857 CpG sites differentially methylated in aging, and 286,746 in cancer. Hyper- and hypomethylated changes in both processes each had a similar genomic distribution across tissues and displayed tissue-independent alterations. The identified hypermethylated regions in aging and cancer shared a similar bivalent chromatin signature. In contrast, hypomethylated DNA sequences occurred in very different chromatin contexts. DNA hypomethylated sequences were enriched at genomic regions marked with the activating histone posttranslational modification H3K4me1 in aging, whilst in cancer, loss of DNA methylation was primarily associated with the repressive H3K9me3 mark.ConclusionsOur results suggest that the role of DNA methylation as a molecular link between aging and cancer is more complex than previously thought.

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Nuclear receptors for retinoic acid and thyroid hormone regulate transcription of keratin genes.

Cell Regulation ◽

10.1091/mbc.1.12.965 ◽

1990 ◽

Vol 1 (12) ◽

pp. 965-973 ◽

Cited By ~ 63

Author(s):

M Tomic ◽

C K Jiang ◽

H S Epstein ◽

I M Freedberg ◽

H H Samuels ◽

...

Keyword(s):

Retinoic Acid ◽

Thyroid Hormone ◽

Epithelial Cells ◽

Nuclear Receptors ◽

Dna Sequences ◽

Primary Cultures ◽

Direct Interaction ◽

Gene Promoters ◽

Intermediate Filament Proteins ◽

Keratin Gene

In the epidermis, retinoids regulate the expression of keratins, the intermediate filament proteins of epithelial cells. We have cloned the 5' regulatory regions of four human epidermal keratin genes, K#5, K#6, K#10, and K#14, and engineered constructs in which these regions drive the expression of the CAT reporter gene. By co-transfecting the constructs into epithelial cells along with the vectors expressing nuclear receptors for retinoic acid (RA) and thyroid hormone, we have demonstrated that the receptors can suppress the promoters of keratin genes. The suppression is ligand dependent; it is evident both in established cell lines and in primary cultures of epithelial cells. The three RA receptors have similar effects on keratin gene transcription. Our data indicate that the nuclear receptors for RA and thyroid hormone regulate keratin synthesis by binding to negative recognition elements in the upstream DNA sequences of the keratin genes. RA thus has a twofold effect on epidermal keratin expression: qualitatively, it regulates the regulators that effect the switch from basal cell-specific keratins to differentiation-specific ones; and quantitatively, it determines the level of keratin synthesis within the cell by direct interaction of its receptors with the keratin gene promoters.

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