Transcription of the Drosophila melanogaster 5S RNA gene requires an upstream promoter and four intragenic sequence elements

1988 ◽  
Vol 8 (3) ◽  
pp. 1266-1274
Author(s):  
S J Sharp ◽  
A D Garcia
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Start Site ◽  
Transcription Start ◽  
5S Rna ◽  
Upstream Promoter ◽  
5S Dna ◽  
Control Regions

Linker-scanning (LS) mutations were constructed spanning the length of the Drosophila melanogaster 5S RNA gene. In vitro transcription analysis of the LS 5S DNAs revealed five transcription control regions. One control region essential for transcription initiation was identified in the 5'-flanking sequence. The major sequence determinants of this upstream promoter region were located between coordinates -39 and -26 (-30 region), but important sequences extended to the transcription start site at position 1. Since mutations in the upstream promoter did not alter the ability of 5S DNA to sequester transcription factors into a stable transcription complex, it appears that this control region involved the interaction of RNA polymerase III. Active 5S DNA transcription additionally required the four intragenic control regions (ICRs) located between coordinates 3 and 18 (ICR I), 37 and 44 (ICR II), 48 and 61 (ICR III), and 78 and 98 (ICR IV). LS mutations in each ICR decreased the ability of 5S DNA to sequester transcription factors. ICR III, ICR IV, and the spacer sequence between were similar in sequence and position to the determinant elements of the multipartite ICR of Xenopus 5S DNA. The importance of ICR III and ICR IV in transcription initiation and in sequestering transcription factors suggests the presence of an activity in D. melanogaster similar to transcription factor TFIIIA of Xenopus laevis and HeLa cells. Transcription initiation of Drosophila 5S DNA was not eliminated by LS mutations in the spacer region even though these mutations reduced the ability of the TFIIIA-like activity to bind. The previously unidentified control regions ICR I and ICR II appear to be important for the interaction of a transcription factor activity, or multiple-factor activities, distinct from the TFIIIA-like activity. The interaction of this activity with ICR I directed the selection of the transcription start site.

scholarly journals Transcription of the Drosophila melanogaster 5S RNA gene requires an upstream promoter and four intragenic sequence elements.

1988 ◽  
Vol 8 (3) ◽  
pp. 1266-1274 ◽  
Author(s):  
S J Sharp ◽  
A D Garcia
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Start Site ◽  
Transcription Start ◽  
5S Rna ◽  
Upstream Promoter ◽  
5S Dna ◽  
Control Regions

Linker-scanning (LS) mutations were constructed spanning the length of the Drosophila melanogaster 5S RNA gene. In vitro transcription analysis of the LS 5S DNAs revealed five transcription control regions. One control region essential for transcription initiation was identified in the 5'-flanking sequence. The major sequence determinants of this upstream promoter region were located between coordinates -39 and -26 (-30 region), but important sequences extended to the transcription start site at position 1. Since mutations in the upstream promoter did not alter the ability of 5S DNA to sequester transcription factors into a stable transcription complex, it appears that this control region involved the interaction of RNA polymerase III. Active 5S DNA transcription additionally required the four intragenic control regions (ICRs) located between coordinates 3 and 18 (ICR I), 37 and 44 (ICR II), 48 and 61 (ICR III), and 78 and 98 (ICR IV). LS mutations in each ICR decreased the ability of 5S DNA to sequester transcription factors. ICR III, ICR IV, and the spacer sequence between were similar in sequence and position to the determinant elements of the multipartite ICR of Xenopus 5S DNA. The importance of ICR III and ICR IV in transcription initiation and in sequestering transcription factors suggests the presence of an activity in D. melanogaster similar to transcription factor TFIIIA of Xenopus laevis and HeLa cells. Transcription initiation of Drosophila 5S DNA was not eliminated by LS mutations in the spacer region even though these mutations reduced the ability of the TFIIIA-like activity to bind. The previously unidentified control regions ICR I and ICR II appear to be important for the interaction of a transcription factor activity, or multiple-factor activities, distinct from the TFIIIA-like activity. The interaction of this activity with ICR I directed the selection of the transcription start site.

scholarly journals Ordering promoter binding of class III transcription factors TFIIIC1 and TFIIIC2.

1988 ◽  
Vol 8 (8) ◽  
pp. 3017-3025 ◽  
Author(s):  
N Dean ◽  
A J Berk
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Class Iii ◽  
High Affinity ◽  
Necessary And Sufficient ◽  
Functional Components ◽  
Competition Assays

The separation of the mammalian class III transcription factor TFIIIC into two functional components, termed TFIIIC1 and TFIIIC2, enabled an analysis of their functions in transcription initiation. Template competition assays were used to define the order with which these factors interact in vitro to form stable preinitiation complexes on the adenovirus VAI and Drosophila melanogaster tRNA(Arg) genes. The interaction between these genes and TFIIIC2, the factor that binds with high affinity to the B block, was both necessary and sufficient for template commitment. When either the VAI or tRNA(Arg) gene was preincubated with TFIIIC2 alone, transcription of a second gene added subsequently was excluded, indicating that TFIIIC2 bound stably to the first template. Furthermore, the interaction between TFIIIC2 and these genes must occur prior to that of TFIIIC1 or TFIIIB. Once TFIIIC2 was bound, TFIIIC1 could bind to the tRNA(Arg) and VAI genes, although its interaction with the VAI gene was less stable than that with the tRNA(Arg) gene. TFIIIB activity bound stably to the complex of both genes with TFIIIC2. These results demonstrate that TFIIIC2 is the first transcription factor to bind to these genes and that TFIIIB and TFIIIC1 can then interact in either order to form a preinitiation complex.

Ordering promoter binding of class III transcription factors TFIIIC1 and TFIIIC2

1988 ◽  
Vol 8 (8) ◽  
pp. 3017-3025 ◽  
Author(s):  
N Dean ◽  
A J Berk
Keyword(s):  
Transcription Factor ◽  
Drosophila Melanogaster ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Class Iii ◽  
High Affinity ◽  
Necessary And Sufficient ◽  
Functional Components ◽  
Competition Assays

The separation of the mammalian class III transcription factor TFIIIC into two functional components, termed TFIIIC1 and TFIIIC2, enabled an analysis of their functions in transcription initiation. Template competition assays were used to define the order with which these factors interact in vitro to form stable preinitiation complexes on the adenovirus VAI and Drosophila melanogaster tRNA(Arg) genes. The interaction between these genes and TFIIIC2, the factor that binds with high affinity to the B block, was both necessary and sufficient for template commitment. When either the VAI or tRNA(Arg) gene was preincubated with TFIIIC2 alone, transcription of a second gene added subsequently was excluded, indicating that TFIIIC2 bound stably to the first template. Furthermore, the interaction between TFIIIC2 and these genes must occur prior to that of TFIIIC1 or TFIIIB. Once TFIIIC2 was bound, TFIIIC1 could bind to the tRNA(Arg) and VAI genes, although its interaction with the VAI gene was less stable than that with the tRNA(Arg) gene. TFIIIB activity bound stably to the complex of both genes with TFIIIC2. These results demonstrate that TFIIIC2 is the first transcription factor to bind to these genes and that TFIIIB and TFIIIC1 can then interact in either order to form a preinitiation complex.

scholarly journals The miR-203a Regulatory Network Affects the Proliferation of Chronic Myeloid Leukemia K562 Cells

2021 ◽  
Vol 9 ◽  
Author(s):  
Jinhua He ◽  
Zeping Han ◽  
Ziyi An ◽  
Yumin Li ◽  
Xingyi Xie ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Promoter Region ◽  
Target Genes ◽  
Mononuclear Cells ◽  
K562 Cells ◽  
Luciferase Reporter ◽  
Upstream Promoter ◽  
Upstream Promoter Region

To study the molecular mechanism by which miR-203a affects the development of CML, bioinformatics software was used to predict the upstream transcription factors and downstream target genes of miR-203a. A 5’-rapid amplification of cDNA ends assay was performed to detect gene transcription initiation sites. A chromatin immunoprecipitation assay was used to verify the binding of transcription factors and promoter regions. A double luciferase reporter gene vector was constructed to demonstrate the regulatory effect of miR-203a on target genes. Real-time PCR and western blotting were used to detect the relative expression levels of genes and proteins, respectively. The results showed that there was a binding site for the transcription factor EGR1 in the upstream promoter region of miR-203a. WT1, BMI1, and XIAP were identified as target genes regulated by miR-203a. EGR1 and miR-203a were downregulated in human peripheral blood mononuclear cells and the CML K562 cell line, while WT1, BMI1, and XIAP were upregulated. The transcription initiation site of miR-203a was identified in the upstream promoter region (G nucleotide at −339 bp), and the transcription factor EGR1 could bind to the promoter region (at −268 bp) of miR-203a and increase its expression. Over expression of miR-203a inhibited the proliferation of K562 cells. A rescue assay showed that overexpression of WT1, BMI1, and XIAP offset the antitumor effect of miR-203a. Conclusion, EGR1 positively regulated the expression of miR-203a, thus relieving the inhibition of miR-203a on the translation of its target genes (WT1, BMI1, and XIAP) and affecting the proliferation of K562 cells.

scholarly journals Interactions between RNA polymerase and the core recognition element are a determinant of transcription start site selection

2016 ◽  
Vol 113 (21) ◽  
pp. E2899-E2905 ◽  
Author(s):  
Irina O. Vvedenskaya ◽  
Hanif Vahedian-Movahed ◽  
Yuanchao Zhang ◽  
Deanne M. Taylor ◽  
Richard H. Ebright ◽  
...  
Keyword(s):  
Rna Polymerase ◽  
Transcription Start Site ◽  
Transcription Initiation ◽  
Specific Protein ◽  
Start Site ◽  
Transcription Start ◽  
Transcription Bubble ◽  
Recognition Element

During transcription initiation, RNA polymerase (RNAP) holoenzyme unwinds ∼13 bp of promoter DNA, forming an RNAP-promoter open complex (RPo) containing a single-stranded transcription bubble, and selects a template-strand nucleotide to serve as the transcription start site (TSS). In RPo, RNAP core enzyme makes sequence-specific protein–DNA interactions with the downstream part of the nontemplate strand of the transcription bubble (“core recognition element,” CRE). Here, we investigated whether sequence-specific RNAP–CRE interactions affect TSS selection. To do this, we used two next-generation sequencing-based approaches to compare the TSS profile of WT RNAP to that of an RNAP derivative defective in sequence-specific RNAP–CRE interactions. First, using massively systematic transcript end readout, MASTER, we assessed effects of RNAP–CRE interactions on TSS selection in vitro and in vivo for a library of 47 (∼16,000) consensus promoters containing different TSS region sequences, and we observed that the TSS profile of the RNAP derivative defective in RNAP–CRE interactions differed from that of WT RNAP, in a manner that correlated with the presence of consensus CRE sequences in the TSS region. Second, using 5′ merodiploid native-elongating-transcript sequencing, 5′ mNET-seq, we assessed effects of RNAP–CRE interactions at natural promoters in Escherichia coli, and we identified 39 promoters at which RNAP–CRE interactions determine TSS selection. Our findings establish RNAP–CRE interactions are a functional determinant of TSS selection. We propose that RNAP–CRE interactions modulate the position of the downstream end of the transcription bubble in RPo, and thereby modulate TSS selection, which involves transcription bubble expansion or transcription bubble contraction (scrunching or antiscrunching).

scholarly journals Contributions of UP Elements and the Transcription Factor FIS to Expression from the Seven rrn P1 Promoters inEscherichia coli

2001 ◽  
Vol 183 (21) ◽  
pp. 6305-6314 ◽  
Author(s):  
Christine A. Hirvonen ◽  
Wilma Ross ◽  
Christopher E. Wozniak ◽  
Erin Marasco ◽  
Jennifer R. Anthony ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Binding Sites ◽  
Promoter Activity ◽  
The Other ◽  
Multigene Families ◽  
Inescherichia Coli ◽  
Start Site ◽  
Transcription Start ◽  
Control Of Gene Expression ◽  
Up Elements

ABSTRACT The high activity of the rrnB P1 promoter inEscherichia coli results from acis-acting DNA sequence, the UP element, and atrans-acting transcription factor, FIS. In this study, we examine the effects of FIS and the UP element at the other sixrrn P1 promoters. We find that UP elements are present at all of the rrn P1 promoters, but they make different relative contributions to promoter activity. Similarly, FIS binds upstream of, and activates, all seven rrn P1 promoters but to different extents. The total number of FIS binding sites, as well as their positions relative to the transcription start site, differ at each rrn P1 promoter. Surprisingly, the FIS sites upstream of site I play a much larger role in transcription from most rrn P1 promoters compared to rrnBP1. Our studies indicate that the overall activities of the sevenrrn P1 promoters are similar, and the same contributors are responsible for these high activities, but these inputs make different relative contributions and may act through slightly different mechanisms at each promoter. These studies have implications for the control of gene expression of unlinked multigene families.

The T-box transcription factor Brachyury regulates expression of eFGF through binding to a non-palindromic response element

Development ◽  
1998 ◽  
Vol 125 (19) ◽  
pp. 3887-3894 ◽  
Author(s):  
E.S. Casey ◽  
M.A. O'Reilly ◽  
F.L. Conlon ◽  
J.C. Smith
Keyword(s):  
Transcription Factor ◽  
Site Selection ◽  
Target Genes ◽  
Biological Function ◽  
Start Site ◽  
Palindromic Sequence ◽  
Vertebrate Development ◽  
Transcription Start ◽  
T Box ◽  
Human And Mouse

Brachyury is a member of the T-box gene family and is required for formation of posterior mesoderm and notochord during vertebrate development. The ability of Brachyury to activate transcription is essential for its biological function, but nothing is known about its target genes. Here we demonstrate that Xenopus Brachyury directly regulates expression of eFGF by binding to an element positioned approximately 1 kb upstream of the eFGF transcription start site. This site comprises half of the palindromic sequence previously identified by binding site selection and is also present in the promoters of the human and mouse homologues of eFGF.

scholarly journals The macrophage transcription factor PU.1 directs tissue-specific expression of the macrophage colony-stimulating factor receptor.

1994 ◽  
Vol 14 (1) ◽  
pp. 373-381 ◽  
Author(s):  
D E Zhang ◽  
C J Hetherington ◽  
H M Chen ◽  
D G Tenen
Keyword(s):  
Transcription Factor ◽  
Transcription Factors ◽  
Transcription Initiation ◽  
Colony Stimulating Factor ◽  
Specific Expression ◽  
Tissue Specific ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Stimulating Factor

The macrophage colony-stimulating factor (M-CSF) receptor is expressed in a tissue-specific fashion from two distinct promoters in monocytes/macrophages and the placenta. In order to further understand the transcription factors which play a role in the commitment of multipotential progenitors to the monocyte/macrophage lineage, we have initiated an investigation of the factors which activate the M-CSF receptor very early during the monocyte differentiation process. Here we demonstrate that the human monocytic M-CSF receptor promoter directs reporter gene activity in a tissue-specific fashion. Since one of the few transcription factors which have been implicated in the regulation of monocyte genes is the macrophage- and B-cell-specific PU.1 transcription factor, we investigated whether PU.1 binds and activates the M-CSF receptor promoter. Here we demonstrate that both in vitro-translated PU.1 and PU.1 from nuclear extracts bind to a specific site in the M-CSF receptor promoter just upstream from the major transcription initiation site. Mutations in this site which eliminate PU.1 binding decrease M-CSF receptor promoter activity significantly in macrophage cell lines only. Furthermore, PU.1 transactivates the M-CSF receptor promoter in nonmacrophage cells. These results suggest that PU.1 plays a major role in macrophage gene regulation and development by directing the expression of a receptor for a key macrophage growth factor.

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