Regulation of Chicken FABP4 Transcription by Toll-Like Receptor 3 Activation in DF-1 Cells

소재령,김수정,송기덕 한국가금학회 2023 韓國家禽學會誌 Vol.50 No.4

Long-chain fatty acids (LCFAs) are vital in cellular compartments, primarily regulating lipid metabolism. Fatty Acid-Binding Proteins (FABPs) facilitate LCFA transport, lipid synthesis, storage, and act as signaling molecules influencing various pathways, including inflammation. FABP4, in particular, is linked to vascular and cardio-related diseases, and it plays a role in macrophage-mediated inflammatory responses. Previous studies have identified FABP4 as not only a representative biomarker for lipogenesis but also as having correlations with immune responses. This study aims to investigate the regulation of the chicken FABP4 (chFABP4) gene by toll-like receptor 3 (TLR3) activation and determine the signaling pathways that are involved in chFABP4 transcriptional regulation. We analyzed the transcriptional regulation of chFABP4 in TLR3-stimulated DF-1 cells. The results showed that chFABP4 was up-regulated upon stimulation with polyinosinic-polycytidylic acid (PIC), a TLR3 ligand. Notably, chFABP4 transcription was independently regulated in the NF-κB signaling pathway. It was up-regulated in p38 inhibition, demonstrating that the p38 signaling pathway might suppress the transcription of chFABP4 within TLR3-activated DF-1 cells. In contrast, chFABP4 expression was down-regulated in JNK signaling pathway inhibition, suggesting the positive regulation of JNK signaling pathway for chFABP4 transcription in DF-1 cells in response to TLR3 activation, consistent with findings in macrophages. MEK pathway inhibition resulted in a similar regulation to NF-κB signaling. These results suggest that each MAPK contributes differentially to the transcriptional regulation of chFABP4 by in DF-1 cells in response to TLR3 activation.

Post-transcriptional Regulation of NK Cell Activation

김태돈,박주영,최인표 대한면역학회 2009 Immune Network Vol.9 No.4

Natural killer (NK) cells play key roles in innate and adaptive immune defenses. NK cell responses are mediated by two major mechanisms: the direct cytolysis of target cells, and immune regulation by production of various cytokines. Many previous reports show that the complex NK cell activation process requires de novo gene expression regulated at both transcriptional and post-transcriptional levels. Specialized un-translated regions (UTR) of mRNAs are the main mechanisms of post-transcriptional regulation. Analysis of posttranscriptional regulation is needed to clearly understand NK cell biology and, furthermore, harness the power of NK cells for therapeutic aims. This review summarizes the current understanding of mRNA metabolism during NK cell activation, focusing primarily on post-transcriptional regulation. Natural killer (NK) cells play key roles in innate and adaptive immune defenses. NK cell responses are mediated by two major mechanisms: the direct cytolysis of target cells, and immune regulation by production of various cytokines. Many previous reports show that the complex NK cell activation process requires de novo gene expression regulated at both transcriptional and post-transcriptional levels. Specialized un-translated regions (UTR) of mRNAs are the main mechanisms of post-transcriptional regulation. Analysis of posttranscriptional regulation is needed to clearly understand NK cell biology and, furthermore, harness the power of NK cells for therapeutic aims. This review summarizes the current understanding of mRNA metabolism during NK cell activation, focusing primarily on post-transcriptional regulation.

Post-transcriptional Regulation of NK Cell Activation

Kim, Tae-Don,Park, Ju-Yeong,Choi, In-Pyo The Korean Association of Immunobiologists 2009 Immune Network Vol.9 No.4

Natural killer (NK) cells play key roles in innate and adaptive immune defenses. NK cell responses are mediated by two major mechanisms: the direct cytolysis of target cells, and immune regulation by production of various cytokines. Many previous reports show that the complex NK cell activation process requires de novo gene expression regulated at both transcriptional and post-transcriptional levels. Specialized un-translated regions (UTR) of mRNAs are the main mechanisms of post-transcriptional regulation. Analysis of posttranscriptional regulation is needed to clearly understand NK cell biology and, furthermore, harness the power of NK cells for therapeutic aims. This review summarizes the current understanding of mRNA metabolism during NK cell activation, focusing primarily on post-transcriptional regulation.

Novel function of the poly(c)-binding protein α-CP2 as a transcriptional activator that binds to single-stranded DNA sequences

KANG, DUK-HEE,SONG, KYU YOUNG,WEI, LI-NA,LAW, PING-YEE,LOH, HORACE H.,CHOI, HACK SUN D.A. Spandidos 2013 International journal of molecular medicine Vol.32 No.5

<P>α-complex protein 2 (α-CP2) is known as an RNA-binding protein that interacts in a sequence-specific manner with single-stranded polycytosine [poly(C)]. This protein is involved in various post-transcriptional regulations, such as mRNA stabilization and translational regulation. In this study, the full-length mouse α-CP2 gene was expressed in an insoluble form with an N-terminal histidine tag in <I>Escherichia coli</I> and purified for homogeneity using affinity column chromatography. Its identity was confirmed using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Recombinant α-CP2 was expressed and refolded. The protein folding conditions for denatured α-CP2 were optimized. DNA and RNA electrophoretic mobility shift assays demonstrated that the recombinant α-CP2 is capable of binding to both single-stranded DNA and RNA poly(C) sequences. Furthermore, plasmids expressing α-CP2 activated the expression of a luciferase reporter when co-transfected with a single-stranded (pGL-SS) construct containing a poly(C) sequence. To our knowledge, this study demonstrates for the first time that α-CP2 functions as a transcriptional activator by binding to a single-stranded poly(C) sequence.</P>

Development of a novel cellulase biosensor that detects crystalline cellulose hydrolysis using a transcriptional regulator

Kwon, Kil Koang,Yeom, Soo-Jin,Lee, Dae-Hee,Jeong, Ki Jun,Lee, Seung-Goo Elsevier 2018 Biochemical and biophysical research communication Vol.495 No.1

<P><B>Abstract</B></P> <P>Successful utilization of cellulose as renewable biomass depends on the development of economically feasible technologies, which can aid in enzymatic hydrolysis. In this study, we developed a whole-cell biosensor for detecting cellulolytic activity that relies on the recognition of cellobiose using the transcriptional factor CelR from <I>Thermobifida fusca</I> and transcriptional activation of its downstream <I>gfp</I> reporter gene. The fluorescence intensity of whole-cell biosensor, which was named as cellobiose-detectible genetic enzyme screening system (CBGESS), was directly proportional to the concentration of cellobiose. The strong fluorescence intensity of CBGESS demonstrated the ability to detect cellulolytic activity with two cellulosic substrates, carboxymethyl cellulose and <I>p</I>-nitrophenyl β-D-cellobioside in cellulase-expressing <I>Escherichia coli</I>. In addition, CBGESS easily sensed crystalline cellulolytic activity when commercial Celluclast 1.5L was dropped on an Avicel plate. Therefore, CBGESS is a powerful tool for detecting cellulolytic activity with high sensitivity in the presence of soluble or insoluble cellulosic substrates. CBGESS may be further applied to excavate novel cellulases or microbes from both genetic libraries and various environments.</P> <P><B>Highlights</B></P> <P> <UL> <LI> A novel biosensor for detecting cellulolytic activity is developed using cellobiose-dependent transcriptional regulator. </LI> <LI> CBGESS shows quantitative response to cellobiose with hyper sensitivity and specificity. </LI> <LI> Cellulase activity including hydrolysis of crystalline cellulose is measured by CBGESS <I>via</I> intracellular fluorescence. </LI> </UL> </P>

Decreased Growth and Antibiotic Production in Streptomyces coelicolor A3(2) by Deletion of a Highly Conserved DeoR Family Regulator, SCO1463

전종민,최태림,이보람,서주현,송훈석,정혜림,양수연,박준영,김은정,김병기,양영헌 한국생물공학회 2019 Biotechnology and Bioprocess Engineering Vol.24 No.4

Streptomyces spp. have been isolated from different environmental niches and are known to exhibit diversity in secondary metabolism. They have complex regulation system to control secondary metabolism, however it is difficult to prioritize the importance of specific regulator among the thousands of global or cluster-situated genes that may regulate secondary metabolism in different Streptomyces strains. Here we suggest a simple homologybased selection method to find out important regulators by comparing seven Streptomyces strains finding highly homologous regulators in various Streptomyces strains and showed highly homologous 11 regulators containing four well known regulators such as BldM, IclR, WhiD and NdgR. Among various regulators, we showed a putative transcriptional regulator in Streptomyces coelicolor A3(2), SCO1463 playing a pivotal role in growth, antibiotic production (actinorhodin[ACT] and undecylprodigiosin [RED] production), and production/utilization of organic acids such as propionate and succinate by making comparisons between the deletion mutant and the wild type strain. Although high homology in various strains does not always mean the importance of a gene, we suggested a criterion on which regulator should be studied first and which would be more important among more than one thousand regulators.

Ralstonia eutropha JMP134에서 페놀분해에 관여하는 조절유전자의 Subcloning 및 염기서열 분석

김기황,김영준,Subramanian Chitra 한국미생물학회 2002 미생물학회지 Vol.38 No.4

Ralstonia eutropha JMP134로부터 페놀대사의 조절에 관여하는 유전자부위를 cloning하여 염기서열을 파악하였으며 그 특성을 조사하였다. 염기서열 분석 결과 두 개의 open reading frame (ORF1 & ORF2)들을 발견하였다. ORF1은 페놀분해 구조유전자중의 마지막 인자인phlX의 stop codon으로부터 454 bp 아래에서 시작하여 총 501 개의 아미노산으로 구성되었으며 ORF2는 ORF1의 stop codon으로부터 1 bP 위쪽에서 4개의 염기쌍과 중첩된 상태에서 시작하여 총 232개의 아미노산으로 구성되어 있는 것으로 나타났다. 단백질 배열을 분석해본 결과 ORF1은 transcriptional activator로 작용하는 NtrC family에 속하는 것으로 확인되었으며, ORF2는 negative regulator로 알려진 GntR family에 속하는 것으로 나타났다. ORF1과 ORF2를 encoding하는 유전자를 각각 phlR2 와 phlA로 명명하였으며 이들의 가능한 조절기작을 고찰하였다. In this study, chromosomal DNA fragment related to the regulation of phenol metabolism in Ralstonia eutropha JMP 134 was cloned and sequenced. The result has shown that two open reading frames (ORF1 and ORF2) exist on this regulatory region. ORF1, which initiates from 454 bp downstream of the stop codon of the phenol hydroxylase genes, was found to be composed of 501 amino acids. ORF2, whose start codon is overlapped with the stop codon of ORFl, was found to contain 232 amino acids. The comparison of amino acid sequences with other proteins has revealed that ORF1 belongs to the family of NtrC transcriptional activator, whereas ORF2 shares high homology with the family of GntR protein, which is known to be a negative regulator. ORF1 and ORF2 were designated as a putative positive regulator, phlR2 and a negative regulator phlA, respectively. Possible regulatory mechanisms of phenol metabolism in this strain was discussed.

Regulation of Actin Gene Expression During the Differentiation of Naegleria gruberi

Kim, Misook,Lee, Joo-Hun The Microbiological Society of Korea 2001 The journal of microbiology Vol.39 No.1

The regulation of actin gene expression during the differentiation of Naegleria gruberi was examined. Actin mRNA concentration was maximal in amoebae and decreased rapidly after the initiation of differentiation. At 20 min after initiation, the concentration of actin mRNA decreased to 55% of the maximal value. The actin mRNA concentration decreased to the minimum at 80 min (15% of the maximum), and then began to increase slightly at the end of differentiation. This decrease of actin mRNA concentration was regulated by the repression of actin gene transcription based on nuclear run-on transcription experiments. The rates of transcription of actin gene in nuclei prepared at 40 and 80 min after the initiation of differentiation were 50 and 28% of that of nuclei prepared at the beginning of differentiation, respectively. The addition of cycloheximide at the initiation of differentiation inhibited both the rapid decrease in the concentration of actin mRNA and the repression of actin gene transcription. These results suggest that the rapid decrease in the concentration of actin mRNA during the differentiation of N. gruberi is accomplished by the repression of actin gene transcription and this transcriptional regulation requires continuous protein synthesis during the differentiation.

Transcriptional Regulation of the Xbr-1a/Xvent-2 Gene by BMP-4 Signaling during Xenopus Embryonic Development

김재봉(Jae-bong Kim),이효상(Hyo-sang Lee),노동현(Dong-Hyun Roh),황유석(Yoo-Seok Hwang),Ren-He Xu, Hsiang-Fu Kung, Yong-Chul Bae, 박매자(Mae-Ja Park) 대한해부학회 2000 Anatomy & Cell Biology Vol.33 No.5

BMP-4 signaling is mediated through Smad proteins which may translocate to the nucleus to activate transcription. Little is known about how BMP-4 signaling regulates the transcription of its target genes, e.g., Xvent genes. Therefore, we isolated the genomic clone of a BMP-4 responsive homeobox gene, Xbr-1a/Xvent-2. This clone contains a promoter and three exons for the entire coding region. Using the primer extension, we identified the transcription initiation site corresponding to position -64 bp upstream to the ATG codon of the Xvent-2 gene. The promoter was linked to the luciferase reporter gene, and promoter activity determined by luciferase assay. The temporal promoter activity peaked between embryonic stages 13~17, in agreement with its temporal mRNA expression in the whole embryo. Through the serial deletion mutation, the upstream -235 bp of the promoter retains the full transcriptional activity, and is regulated by BMP-4 signaling. The present results suggest that the BMP-4 responsive element is located on the upstream 235 bp of the promoter. BMP-4의 세포 내 신호전달은, Smad 단백질이 세포핵내로 이동하여 표적 유전자의 전사를 자극하는 과정에 의하여, 매개되는 것으로 알려져 있지만, BMP-4가 자신의 표적유전자 중의 하나인, Xvent유전자의 전사를 어떻게 조절하는지는 잘 알려져 있지 않다. 본 실험실에서는 BMP-4 responsive homeobox 유전자인, Xbr-1a/Xvent-2의 genomic clone을 분리하였으며, 이렇게 분리된 clone에는 promoter와 coding region의 전부위에 해당하는 세개의 exon이 들어 있다는 것을 알 수 있었다. 또한 primer extension 실험결과, 전사시작 지점이 Xvent-2 유전자의 ATG 코돈에서 상류쪽 -64 bp이란 것을 알 수 있었다. Promoter 부위를 luciferase reporter 유전자에 연결한 다음, promoter 활성을 luciferase assay를 통하여 측정하였다. Promoter 활성은, 개구리 배자에서의 Xvent-2 유전자의 mRNA 발현양상과 비슷하게, 13~17 배자기에 최고치를 나타내었으며, serial deletion mutation실험을 통하여, promoter 부위의 -235 bp 부위에서 BMP-4/Smad1에 의하여 조절되는, 완전한 전사활성이 관찰되었다. 본 실험의 결과는, Xbr-1a/Xvent-2유전자의 promoter부위에서, 상류쪽 -235 bp 부위에 BMP-4 response element가 존재한 다는 것을 보여주는 것으로 사료된다.

Regulation of polyubiquitin genes to meet cellular ubiquitin requirement

( Seung-woo Han ),( Byung-kwon Jung ),( Kwon-yul Ryu ) 생화학분자생물학회 2021 BMB Reports Vol.54 No.4

Ubiquitin (Ub) is one of the proteins that are highly conserved from yeast to humans. It is an essential core unit of the welldefined post-translational modification, called ubiquitination, which is involved in a variety of biological processes. In metazoans, Ub is encoded by two monoubiquitin genes and two polyubiquitin genes, in which a single Ub is fused to a ribosomal protein or Ub coding units are arranged in tandem repeats. In mice, polyubiquitin genes (Ubb and Ubc) play a pivotal role to meet the requirement of cellular Ub pools during embryonic development. In addition, expression levels of polyubiquitin genes are increased to adapt to environmental stimuli such as oxidative, heat-shock, and proteotoxic stress. Several researchers have reported about the perturbation of Ub pools through genetic alteration or exogenous Ub delivery using diverse model systems. To study Ub pool changes in a physiologically relevant manner, changing Ub pools via the regulation of endogenous polyubiquitin gene expression has recently been introduced. Furthermore, to understand the regulation of polyubiquitin gene expression more precisely, cis-acting elements and trans-acting factors, which are regulatory components of polyubiquitin genes, have been analyzed. In this review, we discuss how the role of polyubiquitin genes has been studied during the past decade, especially focusing on their regulation. [BMB Reports 2021; 54(4): 189-195]