scholarly journals Identification of a 119-bp Promoter of the Maize Sulfite Oxidase Gene (ZmSO) That Confers High-Level Gene Expression and ABA or Drought Inducibility in Transgenic Plants

2019 ◽  
Vol 20 (13) ◽  
pp. 3326 ◽  
Author(s):  
Ziwei Xu ◽  
Meiping Wang ◽  
Ziting Guo ◽  
Xianfeng Zhu ◽  
Zongliang Xia
Keyword(s):  
Drought Stress ◽  
Transgenic Plants ◽  
Transcription Start Site ◽  
Sulfite Oxidase ◽  
Upstream Region ◽  
Regulation Mechanism ◽  
Start Site ◽  
Transcription Start ◽  
Mutant Analysis ◽  
High Level

Drought adversely affects crop growth and yields. The cloning and characterization of drought- or abscisic acid (ABA)-inducible promoters is of great significance for their utilization in the genetic improvement of crop resistance. Our previous studies have shown that maize sulfite oxidase (SO) has a sulfite-oxidizing function and is involved in the drought stress response. However, the promoter of the maize SO gene has not yet been characterized. In this study, the promoter (ZmSOPro, 1194 bp upstream region of the translation initiation site) was isolated from the maize genome. The in-silico analysis of the ZmSOPro promoter identified several cis-elements responsive to the phytohormone ABA and drought stress such as ABA-responsive element (ABRE) and MYB binding site (MBS), besides a number of core cis-acting elements, such as TATA-box and CAAT-box. A 5′ RACE (rapid amplification of cDNA ends) assay identified an adenine residue as the transcription start site of the ZmSO. The ZmSOPro activity was detected by β-glucuronidase (GUS) staining at nearly all developmental stages and in most plant organs, except for the roots in transgenic Arabidopsis. Moreover, its activity was significantly induced by ABA and drought stress. The 5′-deletion mutant analysis of the ZmSOPro in tobacco plants revealed that a 119-bp fragment in the ZmSOPro (upstream of the transcription start site) is a minimal region, which is required for its high-level expression. Moreover, the minimal ZmSOPro was significantly activated by ABA or drought stress in transgenic plants. Further mutant analysis indicated that the MBS element in the minimal ZmSOPro region (119 bp upstream of the transcription start site) is responsible for ABA and drought-stress induced expression. These results improve our understanding of the transcriptional regulation mechanism of the ZmSO gene, and the characterized 119-bp promoter fragment could be an ideal candidate for drought-tolerant gene engineering in both monocot and dicot crops.

scholarly journals Characterization of Binding Sequences for Butyrolactone Autoregulator Receptors in Streptomycetes

1999 ◽  
Vol 181 (16) ◽  
pp. 5075-5080 ◽  
Author(s):  
Hiroshi Kinoshita ◽  
Tomohiro Tsuji ◽  
Hiroomi Ipposhi ◽  
Takuya Nihira ◽  
Yasuhiro Yamada
Keyword(s):  
Transcription Start Site ◽  
Dna Sequences ◽  
Target Genes ◽  
Start Codon ◽  
Receptor Protein ◽  
Upstream Region ◽  
Start Site ◽  
Transcription Start ◽  
S1 Nuclease ◽  
Nuclease Mapping

ABSTRACT BarA of Streptomyces virginiae is a specific receptor protein for a member of butyrolactone autoregulators which binds to an upstream region of target genes to control transcription, leading to the production of the antibiotic virginiamycin M1 and S. BarA-binding DNA sequences (BarA-responsive elements [BAREs]), to which BarA binds for transcriptional control, were restricted to 26 to 29-nucleotide (nt) sequences on barA and barBupstream regions by the surface plasmon resonance technique, gel shift assay, and DNase I footprint analysis. Two BAREs (BARE-1 and BARE-2) on the barB upstream region were located 57 to 29 bp (BARE-1) and 268 to 241 bp (BARE-2) upstream from the barBtranslational start codon. The BARE located on the barAupstream region (BARE-3) was found 101 to 76 bp upstream of thebarA start codon. High-resolution S1 nuclease mapping analysis revealed that BARE-1 covered the barBtranscription start site and BARE-3 covered an autoregulator-dependent transcription start site of the barA gene. Deletion and mutation analysis of BARE-2 demonstrated that at least a 19-nt sequence was required for sufficient BarA binding, and A or T residues at the edge as well as internal conserved nucleotides were indispensable. The identified binding sequences for autoregulator receptor proteins were found to be highly conserved among Streptomyces species.

scholarly journals Location of promoter elements necessary and sufficient to direct testis-specific expression of the Hst70/Hsp70.2 gene

2004 ◽  
Vol 379 (3) ◽  
pp. 739-747 ◽  
Author(s):  
Dorota ŚCIEGLIŃSKA ◽  
Natallia VYDRA ◽  
Zdzisław KRAWCZYK ◽  
Wiesława WIDŁAK
Keyword(s):  
Gene Expression ◽  
Transcription Start Site ◽  
Gene Promoter ◽  
Specific Gene ◽  
Start Site ◽  
Transcription Start ◽  
Mobility Shift ◽  
Specific Expression ◽  
High Level ◽  
Testis Specific Expression

The rat Hst70 gene and its mouse counterpart Hsp70.2 are expressed specifically in pachytene primary spermatocytes and spermatids. Here we demonstrate that a 165 bp fragment of the Hst70 gene promoter, containing the T1 transcription start site region, entire exon 1 and 42 bp 5´ region of the intron, is sufficient to drive testis-specific expression of the chloramphenicol acetyltransferase reporter gene in transgenic mice with the same developmentally regulated pattern as the endogenous Hsp70.2 gene. We show further that high-level tissue-specific gene expression requires additional sequences localized upstream of the T2 transcription start site. Electrophoretic mobility-shift assay analysis revealed that only testes of juvenile rats, when Hst70 gene expression is repressed, contain proteins that specifically bind to the Oct (octamer) sequence localized directly downstream of the T1 site.

The Role of XPB/Ssl2 dsDNA Translocase Processivity in Transcription Start-site Scanning

2021 ◽  
pp. 166813
Author(s):  
Eric J. Tomko ◽  
Olivia Luyties ◽  
Jenna K. Rimel ◽  
Chi-Lin Tsai ◽  
Jill O. Fuss ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Start Site ◽  
Transcription Start

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).

Mapping of the Human Renin Transcription Start Site: Evidence for a Single Functional Promoter

1988 ◽  
Vol 10 (6) ◽  
pp. 1313-1316
Author(s):  
Martin Paul ◽  
David W. Burt ◽  
Norifumi Nakamura ◽  
Richard E. Pratt ◽  
Victor J. Dzau
Keyword(s):  
Transcription Start Site ◽  
Start Site ◽  
Transcription Start ◽  
Human Renin

scholarly journals Sequence of the 5′-flanking region and promoter activity of the human mucin gene MUC5B in different phenotypes of colon cancer cells

2000 ◽  
Vol 348 (3) ◽  
pp. 675-686 ◽  
Author(s):  
Isabelle VAN SEUNINGEN ◽  
Michaël PERRAIS ◽  
Pascal PIGNY ◽  
Nicole PORCHET ◽  
Jean-Pierre AUBERT
Keyword(s):  
Gene Expression ◽  
Transcription Start Site ◽  
Promoter Activity ◽  
Luciferase Reporter ◽  
Nuclear Factor ◽  
Start Site ◽  
Transcription Start ◽  
Mucin Gene ◽  
Intron 1 ◽  
Flanking Region

Control of gene expression in intestinal cells is poorly understood. Molecular mechanisms that regulate transcription of cellular genes are the foundation for understanding developmental and differentiation events. Mucin gene expression has been shown to be altered in many intestinal diseases and especially cancers of the gastrointestinal tract. Towards understanding the transcriptional regulation of a member of the 11p15.5 human mucin gene cluster, we have characterized 3.55 kb of the 5ʹ-flanking region of the human mucin gene MUC5B, including the promoter, the first two exons and the first intron. We report here the promoter activity of successively 5ʹ-truncated sections of 956 bases of this region by fusing it to the coding region of a luciferase reporter gene. The transcription start site was determined by primer-extension analysis. The region upstream of the transcription start site is characterized by the presence of a TATA box at bases -32/-26, DNA-binding elements for transcription factors c-Myc, N-Myc, Sp1 and nuclear factor ĸB as well as putative activator protein (AP)-1-, cAMP-response-element-binding protein (CREB)-, hepatocyte nuclear factor (HNF)-1-, HNF-3-, TGT3-, gut-enriched Krüppel factor (GKLF)-, thyroid transcription factor (TTF)-1- and glucocorticoid receptor element (GRE)-binding sites. Intron 1 of MUC5B was also characterized, it is 2511 nucleotides long and contains a DNA segment of 259 bp in which are clustered eight tandemly repeated GA boxes and a CACCC box that bind Sp1. AP-2α and GATA-1 nuclear factors were also shown to bind to their respective cognate elements in intron 1. In transfection studies the MUC5B promoter showed a cell-specific activity as it is very active in mucus-secreting LS174T cells, whereas it is inactive in Caco-2 enterocytes and HT-29 STD (standard) undifferentiated cells. Within the promoter, maximal transcription activity was found in a segment covering the first 223 bp upstream of the transcription start site. Finally, in co-transfection experiments a transactivating effect of Sp1 on to MUC5B promoter was seen in LS174T and Caco-2 cells.

scholarly journals MAPCap: A fast and quantitative transcription start site profiling protocol

2019 ◽  
Author(s):  
Vivek Bhardwaj ◽  
Giuseppe Semplicio ◽  
Niyazi Umut Erdogdu ◽  
Asifa Akhtar
Keyword(s):  
High Resolution ◽  
Differential Expression ◽  
Transcription Start Site ◽  
Expression Analysis ◽  
Differential Expression Analysis ◽  
Start Site ◽  
Transcription Start ◽  
Transcription Start Sites

Abstract Below we present a simple and quick TSS quantification protocol, MAPCap (Multiplexed Affinity Purification of Capped RNA) that enables users to combine high-resolution detection of transcription start-sites and differential expression analysis. MAPCap can be used to profile TSS from dozens of samples in a multiplexed way, in 16-18 hours. MAPCap data can be analyzed using our easy-to-use software icetea (https://bioconductor.org/packages/icetea), which allows users to detect robust TSS using replicates, and perform differential TSS analysis.

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