Features Of Transcriptional Regulation Research Articles

Downregulation of LHCGR Attenuates COX-2 Expression and Induces Luteinized Unruptured Follicle Syndrome in Endometriosis.

An association between endometriosis and luteinized unruptured follicle syndrome (LUFs) has long been identified. Although inactivating mutation of luteinizing hormone/choriogonadotropin receptor (LHGCR) results in LUFs, whether LHCGR contributes to promoting LUFs in endometriosis remains elusive. To investigate the effect of LHCGR signaling in the development of endometriosis-associated LUFs and dissect the underlying mechanism in vivo mouse endometriosis model was established to measure the effect on ovarian folliculogenesis. In vitro cultures of primary human GCs collected from patients undergoing in vitro fertilization were performed and treated with human chorionic gonadotropin (hCG), dibutyryl cyclic-AMP (db-cAMP), LHCGR or CCAAT/enhancer binding protein-α (C/EBPα) small interfering RNA to identify the potential mechanisms. KGN cell line was used to investigate the mechanistic features of transcriptional regulation. Results showed an increased incidence of LUFs was observed in mice with endometriosis. The expression of LHCGR was decreased in the GCs of endometriosis mice. In in vitro cell models, LHCGR signaling increased the expression of C/EBPα and cyclooxygenase-2(COX-2), while inhibiting C/EBPα mitigated the induced COX-2 expression. Mechanically, C/EBPα bounded to the promoter region of COX-2 and increased the transcriptional activity under the stimulation of hCG or db-cAMP. Taken together, this study demonstrated that the LHCGR signaling was reduced in GCs of endometriosis and resulted in a decrease in gonadotropin-induced COX-2 expression. Our study might provide new insights into the dysfunction of GCs in endometriosis.

CAPTURE: In Situ Analysis of Chromatin Composition of Endogenous Genomic Loci by Biotinylated dCas9

Cis-regulatory elements (CREs) play a pivotal role in spatiotemporal control of tissue-specific gene expression, yet the molecular composition of the vast majority of CREs in native chromatin remains unknown. In this article, we describe the clustered regularly interspaced short palindromic repeats (CRISPR) affinity purification in situ of regulatory elements (CAPTURE) approach to simultaneously identify locus-specific chromatin-regulating protein complexes and long-range DNA interactions. Using an in vivo biotinylated nuclease-deficient Cas9 (dCas9) protein and programmable single guide RNAs (sgRNAs), this approach allows for high-resolution and locus-specific isolation of protein complexes and long-range chromatin looping associated with single copy CREs in mammalian cells. Unbiased analysis of the compositional structure of developmentally regulated or disease-associated CREs identifies new features of transcriptional regulation. Hence, CAPTURE provides a versatile platform to study genomic locus-regulating chromatin composition in a mammalian genome. © 2018 by John Wiley & Sons, Inc.

Identification of Promoters of Esherichia coli and Phage in Transcription Section Plasmid pSA850

Publisher Summary This chapter explores the transcriptional regulation of structural genes as an important aspect by which Escherichia coli cells control the catabolism and anabolism of genes. One of the advantages of this system is that gal promoters or any promoter of interest can be studied in vitro under different physiological conditions in the absence of any competing promoter for RNA polymerase. The chapter identifies several major promoters that have enabled to compare physiological effects on these promoters and to better understand certain features of transcriptional regulation, which were not obvious using minicircles. A promoter is also identified in the bacterial attachment site. This promoter may be involved in site-specific recombination between the phage attachment site and bacterial attachment site. The chapter presents a detailed account of how the 5´ and 3´ ends, along with the direction of unknown transcripts with the use of in vitro transcription and primer extension assays.