scholarly journals A Novel Isoform of Liver Receptor Homolog-1 Is Regulated by Steroidogenic Factor-1 and the Specificity Protein Family in Ovarian Granulosa Cells

Endocrinology ◽  
2013 ◽  
Vol 154 (4) ◽  
pp. 1648-1660 ◽  
Author(s):  
Shinya Kawabe ◽  
Takashi Yazawa ◽  
Masafumi Kanno ◽  
Yoko Usami ◽  
Tetsuya Mizutani ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Granulosa Cells ◽  
Promoter Activity ◽  
The Novel ◽  
Start Site ◽  
Transcription Start ◽  
Steroidogenic Factor 1 ◽  
Exon 2 ◽  
Ovarian Granulosa Cells ◽  
Specificity Protein

Abstract Liver receptor homolog-1 (LRH-1) is a member of the nuclear receptor 5A (NR5A) subfamily. It is expressed in granulosa cells of the ovary and is involved in steroidogenesis and ovulation. To reveal the transcriptional regulatory mechanism of LRH-1, we determined its transcription start site in the ovary using KGN cells, a human granulosa cell tumor cell line. 5′-rapid amplification of cDNA ends PCR revealed that human ovarian LRH-1 was transcribed from a novel transcription start site, termed exon 2o, located 41 bp upstream of the reported exon 2. The novel LRH-1 isoform was expressed in the human ovary but not the liver. Promoter analysis and an EMSA indicated that a steroidogenic factor-1 (SF-1) binding site and a GC box upstream of exon 2o were required for promoter activity, and that SF-1 and specificity protein (Sp)-1/3 bind to the respective regions in ovarian granulosa cells. In KGN cells, transfection of SF-1 increased ovarian LRH-1 promoter activity and SF-1-dependent reporter activity was further enhanced when peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α) was cotransfected. In Drosophila SL2 cells, Sp1 was more effective than Sp3 in enhancing promoter activity, and co-transfection of the NR5A-family synergistically increased activity. Infection with adenoviruses expressing SF-1 or PGC-1α induced LRH-1 expression in KGN cells. These results indicate that the expression of human LRH-1 is regulated in a tissue-specific manner, and that the novel promoter region is controlled by the Sp-family, NR5A-family and PGC-1α in ovarian granulosa cells in a coordinated fashion.

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 Influence of cis Element Arrangement on Promoter Strength in Trichoderma reesei

2017 ◽  
Vol 84 (1) ◽  
Author(s):  
Daniel P. Kiesenhofer ◽  
Robert L. Mach ◽  
Astrid R. Mach-Aigner
Keyword(s):  
Trichoderma Reesei ◽  
Transcription Start Site ◽  
Binding Sites ◽  
Inverted Repeat ◽  
Promoter Activity ◽  
Promoter Strength ◽  
Start Site ◽  
Transcription Start ◽  
Content Type ◽  
Sheer Number

ABSTRACTTrichoderma reeseican produce up to 100 g/liter of extracellular proteins. The major and industrially relevant products are cellobiohydrolase I (CBHI) and the hemicellulase XYNI. The genes encoding both enzymes are transcriptionally activated by the regulatory protein Xyr1. The first 850 nucleotides of thecbh1promoter contain 14 Xyr1-binding sites (XBS), and 8 XBS are present in thexyn1promoter. Some of these XBS are arranged in tandem and others as inverted repeats. One suchciselement, an inverted repeat, plays a crucial role in the inducibility of thexyn1promoter. We investigated the impact of the properties of suchciselements by shuffling them by insertion, exchange, deletion, and rearrangement ofciselements in both thecbh1andxyn1promoter. A promoter-reporter assay using theAspergillus nigergoxAgene allowed us to measure changes in the promoter strength and inducibility. Most strikingly, we found that an inverted repeat of XBS causes an important increase incbh1promoter strength and allows induction by xylan or wheat straw. Furthermore, evidence is provided that the distances ofciselements to the transcription start site have important influence on promoter activity. Our results suggest that the arrangement and distances ofciselements have large impacts on the strength of thecbh1promoter, whereas the sheer number of XBS has only secondary importance. Ultimately, the biotechnologically importantcbh1promoter can be improved byciselement rearrangement.IMPORTANCEIn the present study, we demonstrate that the arrangement ofciselements has a major impact on promoter strength and inducibility. We discovered an influence on promoter activity by the distances ofciselements to the transcription start site. Furthermore, we found that the configuration ofciselements has a greater effect on promoter strength than does the sheer number of transactivator binding sites present in the promoter. Altogether, the arrangement ofciselements is an important factor that should not be overlooked when enhancement of gene expression is desired.

scholarly journals Transcription Regulation by Tandem-Bound FNR at Escherichia coli Promoters

2003 ◽  
Vol 185 (20) ◽  
pp. 5993-6004 ◽  
Author(s):  
Anne M. L. Barnard ◽  
Jeffrey Green ◽  
Stephen J. W. Busby
Keyword(s):  
Escherichia Coli ◽  
Transcription Factor ◽  
Binding Site ◽  
Transcription Regulation ◽  
Transcription Start Site ◽  
Promoter Activity ◽  
Start Site ◽  
Transcription Start ◽  
Substitution Mutant

ABSTRACT FNR is an Escherichia coli transcription factor that regulates the transcription of many genes in response to anaerobiosis. We have constructed a series of artificial FNR-dependent promoters, based on the melR promoter, in which a consensus FNR binding site was centered at position −41.5 relative to the transcription start site. A second consensus FNR binding site was introduced at different upstream locations, and promoter activity was assayed in vivo. FNR can activate transcription from these promoters when the upstream FNR binding site is located at many different positions. However, sharp repression is observed when the upstream-bound FNR is located near positions −85 or −95. This repression is relieved by the FNR G74C substitution mutant, previously identified as being defective in transcription repression at the yfiD promoter. A parallel series of artificial FNR-dependent promoters, carrying a consensus FNR binding site at position −61.5 and a second upstream DNA site for FNR, was also constructed. Again, promoter activity was repressed by FNR when the upstream-bound FNR was located at particular positions.

scholarly journals Role of the Transcription Start Site Core Region and Transcription Factor YY1 in Rous Sarcoma Virus Long Terminal Repeat Promoter Activity

1998 ◽  
Vol 72 (8) ◽  
pp. 6592-6601 ◽  
Author(s):  
Constance M. Mobley ◽  
Linda Sealy
Keyword(s):  
Transcription Factor ◽  
Transcription Start Site ◽  
Rous Sarcoma Virus ◽  
Promoter Activity ◽  
Transcriptional Activity ◽  
Start Site ◽  
Transcription Start ◽  
Rous Sarcoma ◽  
Sarcoma Virus ◽  
Transcription Factor Yy1

ABSTRACT The Rous sarcoma virus (RSV) long terminal repeat (LTR) contains a transcriptionally potent enhancer and promoter that functions in a variety of cell types. Previous studies have identified the viral sequences required for enhancer activity, and characterization of these elements has provided insight into the mechanism of RSV transcriptional activity. The objective of this study was to better define the RSV LTR promoter by examining the transcription start site core (TSSC) region. Deletion of the TSSC resulted in complete loss of transcriptional activity despite the presence of a functional TATA box, suggesting that the TSSC is required for viral expression. Homologies within the TSSC to the DNA binding motif of YY1 suggested that it might regulate promoter activity. YY1 has been shown to regulate transcription in some cellular genes and viral promoters by binding to sites overlapping the transcription start site. Gel shift assays using YY1 antibody identified YY1 as one of three complexes that bound to the TSSC. Mutation of the YY1 binding site reduced RSV transcriptional activity by more than 50%, suggesting that YY1, in addition to other TSSC-binding factors, regulates RSV transcription. Furthermore, in vitro transcription assays performed with Drosophila embryo extract (devoid of YY1 activity) showed decreased levels of RSV transcription, while transient transfection experiments overexpressing YY1 demonstrated that YY1 could transactivate the RSV LTR ∼6- to 7-fold. We propose that the TSSC plays a vital role in RSV transcription and that this function is partially carried out by the transcription factor YY1.

scholarly journals Genomic Characterization of Human DSPG3

1999 ◽  
Vol 9 (5) ◽  
pp. 449-456
Author(s):  
Michelle Deere ◽  
Jose L. Dieguez ◽  
Sung-Joo Kim Yoon ◽  
David Hewett-Emmett ◽  
Albert de la Chapelle ◽  
...  
Keyword(s):  
Transcription Start Site ◽  
Sequence Data ◽  
Stop Codon ◽  
Genomic Structure ◽  
Start Codon ◽  
Start Site ◽  
Transcription Start ◽  
Exon 2 ◽  
Ancestral Gene ◽  
Link Type

DSPG3, the human homolog to chick PG-Lb, is a member of the small leucine-rich repeat proteoglycan (SLRP) family, including decorin, biglycan, fibromodulin, and lumican. In contrast to the tissue distribution of the other SLRPs, DSPG3 is predominantly expressed in cartilage. In this study, we have determined that the human DSPG3 gene is composed of seven exons: Exon 2 ofDSPG3 includes the start codon, exons 4–7 code for the leucine-rich repeats, exons 3 and 7 contain the potential glycosaminoglycan attachment sites, and exon 7 contains the potential N-glycosylation sites and the stop codon. We have identified two polymorphic variations, an insertion/deletion composed of 19 nucleotides in intron 1 and a tetranucleotide (TATT)n repeat in intron 5. Analysis of 1.6 kb of upstream promoter sequence ofDSPG3 reveals three TATA boxes, one of which is 20 nucleotides before the transcription start site. The transcription start site precedes the translation start site by 98 nucleotides. There are 14 potential binding sites for SOX9, a transcription factor present in cartilage, in the promoter, and in the first intron of DSPG3. We have examined the evolution of the SLRP gene family and found that gene products clustered together in the evolutionary tree are encoded by genes with similarities in genomic structure. Hence, it appears that the majority of the introns in the SLRP genes were inserted after the differentiation of the SLRP genes from an ancestral gene that was most likely composed of 2–3 exons.[The sequence data described in this paper have been submitted to GenBank under accession nos.AF031658 and U63814.]

scholarly journals The R3 Region, One of Three Major Repetitive Regions of Human Herpesvirus 6, Is a Strong Enhancer of Immediate-Early Gene U95

2001 ◽  
Vol 75 (21) ◽  
pp. 10149-10160 ◽  
Author(s):  
Masaya Takemoto ◽  
Takuya Shimamoto ◽  
Yuji Isegawa ◽  
Koichi Yamanishi
Keyword(s):  
Transcription Start Site ◽  
Binding Sites ◽  
Promoter Activity ◽  
Human Herpesvirus ◽  
Early Gene ◽  
Immediate Early ◽  
Start Site ◽  
Human Herpesvirus 6 ◽  
Transcription Start ◽  
Herpesvirus 6

ABSTRACT An immediate-early (IE) gene of human herpesvirus 6 (HHV-6), U95, has similarity at the amino acid level to the murine cytomegalovirus (MCMV) IE2 gene and is related to the human cytomegalovirus (HCMV) US22 gene family. Sequence analyses of U95 cDNA clones revealed that the transcription start site was located about 1.6 kbp upstream of the putative initiating ATG and that the transcript consisted of two exons. A single intron extended from nucleotides 142589 to 144229, which contained ORF U94. A protein with a molecular mass of about 120 kDa was translated from this cDNA clone in an in vitro transcription-translation assay. The transcription start site was found to be 220 bp downstream of the R3 region by primer extension analysis. HHV-6 has three repetitive elements, R1, R2, and R3, in or near the IE-A locus. R3 is composed of 24 copies of a 104- to 107-bp sequence element, which contains multiple putative binding sites for cellular transcription factors such as AP2 and NF-κB, and its biological significance has yet to be elucidated. The region between −710 and +46 relative to the transcription start site of U95 was analyzed in this study. Deletion from −710 to −396, corresponding to three copies of an R3 unit, decreased the promoter activity by 15-fold, and coexpression of IκBα(S32A/S36A) repressed it to almost the same level. Electrophoretic mobility shift assays showed that NF-κB family members p50 and c-Rel bound to NF-κB sites derived from the R3 region. These results demonstrate that R3 strongly enhances the U95 promoter activity and that NF-κB and binding sites for NF-κB in the R3 region play an important role in its activation. Because U95 promoter activity correlated with the number of R3 units, which each contained an NF-κB site, the repetitive organization of R3 is important for regulating U95 transcription.

scholarly journals c-mos expression in mouse oocytes is controlled by initiator-related sequences immediately downstream of the transcription initiation site

1991 ◽  
Vol 11 (10) ◽  
pp. 5190-5196
Author(s):  
S K Pal ◽  
S S Zinkel ◽  
A A Kiessling ◽  
G M Cooper
Keyword(s):  
Transcription Start Site ◽  
Transcription Initiation ◽  
Promoter Activity ◽  
Consensus Sequence ◽  
Initiation Site ◽  
Transcription Initiation Site ◽  
Transcriptional Level ◽  
Start Site ◽  
Transcription Start ◽  
Mouse Oocytes

We have employed transient expression assays to analyze the sequences that direct c-mos transcription in mouse oocytes. Plasmids containing the chloramphenicol acetyltransferase (CAT) gene fused to either a 2.4-kb or a 731-bp fragment from the 5'-flanking region of c-mos produced similar levels of CAT activity when injected into nuclei of growing oocytes. BAL 31 deletions revealed that sequences up to 20 bp upstream of the major transcription start site could be removed without any significant loss of CAT activity. Promoter activity only decreased when these deletions closely approached the transcription start site, which was mapped at 53 nucleotides upstream of the first ATG in the c-mos open reading frame. On the other hand, deletion of sequences within 20 nucleotides downstream of the transcription initiation site resulted in a 10-fold reduction in CAT expression. A similar decrease in promoter activity was observed as a result of point mutations in these 5' untranslated sequences. Thus, sequences immediately downstream of the transcription start site, including a consensus sequence (PyPyCAPyPyPyPyPy) present in the initiator elements of several genes, appear to regulate c-mos expression in mouse oocytes. Reverse transcription-polymerase chain reaction analysis of RNA from injected oocytes showed that this regulation is manifest at the transcriptional level. Expression of c-mos in mouse oocytes thus appears to be directed by a simple promoter consisting only of sequences immediately surrounding the transcription start site, including an initiator element in the untranslated leader.

Sign in / Sign up

Export Citation Format

Share Document