Transcriptional Regulation of AQP-8, a Caenorhabditis elegans Aquaporin Exclusively Expressed in the Excretory System, by the POU Homeobox Transcription Factor CEH-6

Due to the ever changing environmental conditions in soil, regulation of osmotic homeostasis in the soil-dwelling nematode Caenorhabditis elegans is critical. AQP-8 is a C. elegans aquaporin that is expressed in the excretory cell, a renal equivalent tissue, where the protein participates in maintaining water balance. To better understand the regulation of AQP-8, we undertook a promoter analysis to identify the aqp-8 cis-regulatory elements. Using progressive 5′ deletions of upstream sequence, we have mapped an essential regulatory region to roughly 300 bp upstream of the translational start site of aqp-8. Analysis of this region revealed a sequence corresponding to a known DNA functional element (octamer motif), which interacts with POU homeobox transcription factors. Phylogenetic footprinting showed that this site is perfectly conserved in four nematode species. The octamer site's function was further confirmed by deletion analyses, mutagenesis, functional studies, and electrophoretic mobility shift assays. Of the three POU homeobox proteins encoded in the C. elegans genome, CEH-6 is the only member that is expressed in the excretory cell. We show that expression of AQP-8 is regulated by CEH-6 by performing RNA interference experiments. CEH-6's mammalian ortholog, Brn1, is expressed both in the kidney and the central nervous system and binds to the same octamer consensus binding site to drive gene expression. These parallels in transcriptional control between Brn1 and CEH-6 suggest that C. elegans may well be an appropriate model for determining gene-regulatory networks in the developing vertebrate kidney.

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Two different negative regulatory elements control the transcription of T-cell activation gene 3 in activated mast cells

Biochemical Journal ◽

10.1042/bj3230511 ◽

1997 ◽

Vol 323 (2) ◽

pp. 511-519 ◽

Cited By ~ 6

Author(s):

Chad K. OH ◽

Markus NEURATH ◽

Jeong-Je CHO ◽

Tekli SEMERE ◽

Dean D. METCALFE

Keyword(s):

Protein Synthesis ◽

T Cell ◽

Mast Cells ◽

T Cell Activation ◽

Cell Activation ◽

Regulatory Region ◽

Regulatory Elements ◽

Site Directed Mutagenesis ◽

Mobility Shift ◽

Integrin Gene

T-cell activation gene 3 (TCA3) encodes a β-chemokine that is transcriptionally regulated in mast cells; the gene has a functional NF-κB element at positions -194 to -185. The 5´-flanking region of this gene is also known to have a negative regulatory region between -2057 and -1342. To characterize the negative regulatory elements (NREs), this region was sequenced and then digested by HindIII enzyme into two fragments, NRE-1 (-2057 to -1493) and NRE-2 (-1492 to -1342). Both NRE-1 and NRE-2 in the 5´–3´ orientation inhibited chloramphenicol acetyltransferase (CAT)-protein synthesis by a TCA3–CAT construct transfected into mast cells that were then activated. Only NRE-1 inhibited CAT-protein synthesis in the 3´–5´ orientation. Further deletion of the 5´ region of NRE-1 partially abolished the inhibitory activity. Both NRE-1 and NRE-2 inhibited the activity of a CD20–CAT construct independent of cell activation. Electrophoretic mobility shift assays showed DNA–protein complex formation with subsequences (CCCCCATTCT) of NRE-1 (NRE-1a) and (CCATGA) of NRE-2 (NRE-2b). NRE-1a appears to be novel. NRE-2b is identical with a putative silencer motif in the αIIb integrin gene. Site-directed mutagenesis demonstrated that both NRE-1a and NRE-2b are important in the negative regulation of TCA3 promoter activity. In vivo ligation-mediated PCR footprinting of the NRE-2 region revealed protection between -1372 and -1354, which contains NRE-2b. The data thus demonstrate identity of a silencer motif, here termed NRE-2b, in both the αIIb integrin gene and the TCA3, and that this silencer region in mast cells is functional both in vivoand in vitro. Further, evidence is presented that the promoter for TCA3 contains a novel silencer motif, termed NRE-1a, characterized by a CT-rich sequence.

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Regulatory elements in the upstream region of metallothionein gene in tilapia species

Dhaka University Journal of Biological Sciences ◽

10.3329/dujbs.v30i1.51813 ◽

2021 ◽

Vol 30 (1) ◽

pp. 95-103

Author(s):

Mohammad Shamimul Alam ◽

Israt Jahan ◽

Sadniman Rahman ◽

Hawa Jahan ◽

Kaniz Fatema

Keyword(s):

Tissue Specificity ◽

Regulatory Region ◽

Regulatory Elements ◽

Metal Resistance ◽

Genomic Region ◽

Upstream Region ◽

Upstream Sequence ◽

Oreochromis Aureus ◽

Metallothionein Gene ◽

Genomic Regions

Tilapia is a hardy fish which can survive in water bodies polluted with heavy metals. Metal resistance is conferred by higher expression of metallothionein gene (mt) in many organisms. Level, time and tissue-specificity of gene expression is regulated through transcription factor binding sites (TFBS) which may be present in the upstream, downstream, or even in the introns of a gene. So, as a candidate regulatory region, the 5’upstream sequence of mt gene in three tilapia species, Oreochromis aureus, O. niloticus and O. mossambicus was studied. The targeted region was PCR-amplified and then sequenced using a pair of custom-designed primer. A total of only 2.7% variation was found in the sequenced genomic region among the three species. Metal-related TFBS were predicted from these sequences. A total of twenty eight TFBS were found in O. aureus and twenty nine in O. mossambicus and O. niloticus. The number of metalrelated TFBS predicted in the targeted sequence was significantly higher compared to that found in randomly selected other genomic regions of same size from O. niloticus genome. Thus, the results suggest the presence of putative regulatory elements in the targeted upstream region which might have important role in the regulation of mt gene function. Dhaka Univ. J. Biol. Sci. 30(1): 95-103, 2021 (January)

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Transcription factor B cell lineage-specific activator protein regulates the gene for human X-box binding protein 1.

Journal of Experimental Medicine ◽

10.1084/jem.183.2.393 ◽

1996 ◽

Vol 183 (2) ◽

pp. 393-401 ◽

Cited By ~ 123

Author(s):

A M Reimold ◽

P D Ponath ◽

Y S Li ◽

R R Hardy ◽

C S David ◽

...

Keyword(s):

Transcription Factor ◽

B Cell ◽

Cell Lines ◽

Binding Protein ◽

Cell Lineage ◽

Regulatory Elements ◽

Target Sequence ◽

Consensus Binding Site ◽

Mobility Shift ◽

Activator Protein

The transcription factor human X-box binding protein 1 (hXBP-1) is a basic region-leucine zipper protein implicated in the regulation of major histocompatibility complex class II gene expression as well as in exocrine gland and skeletal development. Multiple regulatory elements in the hXBP-1 promoter lie 3' to the transcription start site, including the hX2 site, whose core sequence is an AP-1-like element identical to the hXBP-1 target sequence in the HLA-DRA promoter. One complex identified by electrophoretic mobility shift assay (EMSA), complex 3, was previously shown to protect the hX2 site and more 3' bases. Sequence analysis now shows that this region contains a consensus binding site for transcription factor BSAP (B cell lineage-specific activator protein). Complex 3 and BSAP have identical cell-type specificities, as they are found only in pre-B and mature B cell lines. In EMSAs, BSAP antibody specifically recognized complex 3, and in vitro translated BSAP could bind to an hXBP promoter fragment. Cotransfections using an hXBP-1 reporter construct indicated that BSAP downregulates the hXBP-1 promoter. The highest levels of hXBP-1 mRNA were found when BSAP was not expressed, in pre-Pro-B cells and in plasma cell lines. In addition, hXBP-1 and BSAP levels were inversely correlated along the early stages of B cell development. In the regulation of the hXBP-1 promoter, a strong positive transcriptional influence at the hX2 site is opposed by the downregulatory actions of BSAP.

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A Pair of Highly Conserved Two-Component Systems Participates in the Regulation of the Hypervariable FIR Proteins in Different Legionella Species

Journal of Bacteriology ◽

10.1128/jb.01742-06 ◽

2007 ◽

Vol 189 (9) ◽

pp. 3382-3391 ◽

Cited By ~ 10

Author(s):

Michal Feldman ◽

Gil Segal

Keyword(s):

Binding Site ◽

Legionella Pneumophila ◽

Response Regulator ◽

Regulatory Element ◽

Expression System ◽

Regulatory Region ◽

Regulatory Elements ◽

Mobility Shift ◽

Type Iv ◽

Essential Components

ABSTRACT Legionella pneumophila and other pathogenic Legionella species multiply inside protozoa and human macrophages by using the Icm/Dot type IV secretion system. The IcmQ protein, which possesses pore-forming activity, and IcmR, which functions as its chaperone, are two essential components of this system. It was previously shown that in 29 Legionella species, a large hypervariable-gene family (fir genes) is located upstream from a conserved icmQ gene, but although nonhomologous, the FIR proteins were found to function similarly together with their corresponding IcmQ proteins. Alignment of the regulatory regions of 29 fir genes revealed that they can be divided into three regulatory groups; the first group contains a binding site for the CpxR response regulator, which was previously shown to regulate the L. pneumophila fir gene (icmR); the second group, which includes most of the fir genes, contains the CpxR binding site and an additional regulatory element that was identified here as a PmrA binding site; and the third group contains only the PmrA binding site. Analysis of the regulatory region of two fir genes, which included substitutions in the CpxR and PmrA consensus sequences, a controlled expression system, as well as examination of direct binding with mobility shift assays, revealed that both CpxR and PmrA positively regulate the expression of the fir genes that contain both regulatory elements. The change in the regulation of the fir genes that occurred during the course of evolution might be required for the adaptation of the different Legionella species to their specific environmental hosts.

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Cloning and Functional characterization of seed-specific LEC1A promoter from peanut (Arachis hypogaea L.)

10.1101/2020.11.13.381327 ◽

2020 ◽

Author(s):

Guiying Tang ◽

Pingli Xu ◽

Pengxiang Li ◽

Jieqiong Zhu ◽

Guangxia Chen ◽

...

Keyword(s):

Transcriptional Start Site ◽

Functional Characterization ◽

Regulatory Region ◽

Regulatory Elements ◽

Histochemical Staining ◽

Expression Vectors ◽

Upstream Sequence ◽

Functional Region ◽

Chromosomal Walking ◽

Storage Reserve

AbstractLEAFY COTYLEDON1 (LEC1) is a HAP3 subunit of CCAAT-binding transcription factor, which controls several aspects of embryo and postembryo development, including embryo morphogenesis, storage reserve accumulation and skotomorphogenesis. Herein, using the method of chromosomal walking, a 2707bp upstream sequence from the ATG initiation codon site of AhLEC1A which is a homolog of Arabidopsis LEC1 was isolated in peanut. Its transcriptional start site confirmed by 5’ RACE was located at 82 nt from 5’ upstream of ATG. The bioinformatics analysis revealed that there existed many tissue-specific elements and light responsive motifs in its promoter. To identify the functional region of the AhLEC1A promoter, seven plant expression vectors expressing the GUS (β-glucuronidase) gene, driven by 5’ terminal series deleted fragments of AhLEC1A promoter, were constructed and transformed into Arabidopsis. Results of GUS histochemical staining showed that the regulatory region containing 82bp of 5’ UTR and 2228bp promoter could facilitate GUS to express preferentially in the embryos at different development periods of Arabidopsis. Taken together, it was inferred that the expression of AhLEC1A during seed development of peanut might be controlled positively by several seed-specific regulatory elements, as well as negatively by some other regulatory elements inhibiting its expression in other organs. Moreover, the GUS expression pattern of transgenic seedlings in darkness and in light was relevant to the light-responsive elements scattered in AhLEC1A promoter segment, implying that these light-responsive elements harbored in the AhLEC1A promoter regulate skotomorphogenesis of peanut seeds, and AhLEC1A expression was inhibited after the germinated seedlings were transferred from darkness to light.

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Mapping and analysis of Caenorhabditis elegans transcription factor sequence specificities

eLife ◽

10.7554/elife.06967 ◽

2015 ◽

Vol 4 ◽

Cited By ~ 41

Author(s):

Kamesh Narasimhan ◽

Samuel A Lambert ◽

Ally WH Yang ◽

Jeremy Riddell ◽

Sanie Mnaimneh ◽

...

Keyword(s):

Caenorhabditis Elegans ◽

Dna Binding ◽

Regulatory Elements ◽

Nuclear Hormone Receptor ◽

Binding Motifs ◽

T Box ◽

C Elegans ◽

Dna Binding Domains ◽

Binding Domains ◽

Protein Binding Microarray

Caenorhabditis elegans is a powerful model for studying gene regulation, as it has a compact genome and a wealth of genomic tools. However, identification of regulatory elements has been limited, as DNA-binding motifs are known for only 71 of the estimated 763 sequence-specific transcription factors (TFs). To address this problem, we performed protein binding microarray experiments on representatives of canonical TF families in C. elegans, obtaining motifs for 129 TFs. Additionally, we predict motifs for many TFs that have DNA-binding domains similar to those already characterized, increasing coverage of binding specificities to 292 C. elegans TFs (∼40%). These data highlight the diversification of binding motifs for the nuclear hormone receptor and C2H2 zinc finger families and reveal unexpected diversity of motifs for T-box and DM families. Motif enrichment in promoters of functionally related genes is consistent with known biology and also identifies putative regulatory roles for unstudied TFs.

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Regulatory Elements Required for Development of Caenorhabditis elegans Hermaphrodites Are Conserved in the tra-2 Homologue of C. remanei, a Male/Female Sister Species

Genetics ◽

10.1093/genetics/155.1.105 ◽

2000 ◽

Vol 155 (1) ◽

pp. 105-116 ◽

Cited By ~ 11

Author(s):

Eric S Haag ◽

Judith Kimble

Keyword(s):

Rna Interference ◽

Caenorhabditis Elegans ◽

Sequence Analysis ◽

Mating System ◽

Translational Control ◽

Mating Systems ◽

Regulatory Elements ◽

Sister Species ◽

Its Sequence ◽

C Elegans

Abstract The Caenorhabditis elegans hermaphrodite is essentially a female that produces sperm. In C. elegans, tra-2 promotes female fates and must be repressed to achieve hermaphrodite spermatogenesis. In an effort to learn how mating systems evolve, we have cloned tra-2 from C. remanei, the closest gonochoristic relative of C. elegans. We found its structure to be similar to that of Ce-tra-2 but its sequence to be divergent. RNA interference demonstrates that Cr-tra-2 promotes female fates. Two sites of tra-2 regulation are required for the onset of hermaphrodite spermatogenesis in C. elegans. One, the MX region of TRA-2, is as well conserved in C. remanei as it is in C. briggsae (another male/hermaphrodite species), suggesting that this control is not unique to hermaphrodites. Another, the DRE/TGE element of the tra-2 3′ UTR, was not detected by sequence analysis. However, gel-shift assays demonstrate that a factor in C. remanei can bind specifically to the Cr-tra-2 3′ UTR, suggesting that this translational control is also conserved. We propose that both controls are general and do not constitute a novel “switch” that enables sexual mosaicism in hermaphrodites. However, subtle quantitative or qualitative differences in their employment may underlie differences in mating system seen in Caenorhabditis.

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Cloning and functional characterization of seed-specific LEC1A promoter from peanut (Arachis hypogaea L.)

PLoS ONE ◽

10.1371/journal.pone.0242949 ◽

2021 ◽

Vol 16 (3) ◽

pp. e0242949

Author(s):

Guiying Tang ◽

Pingli Xu ◽

Pengxiang Li ◽

Jieqiong Zhu ◽

Guangxia Chen ◽

...

Keyword(s):

Transcriptional Start Site ◽

Functional Characterization ◽

Regulatory Region ◽

Regulatory Elements ◽

Histochemical Staining ◽

Expression Vectors ◽

Upstream Sequence ◽

Functional Region ◽

Chromosomal Walking ◽

Storage Reserve

LEAFY COTYLEDON1 (LEC1) is a HAP3 subunit of CCAAT-binding transcription factor, which controls several aspects of embryo and postembryo development, including embryo morphogenesis, storage reserve accumulation and skotomorphogenesis. Herein, using the method of chromosomal walking, a 2707bp upstream sequence from the ATG initiation codon site of AhLEC1A which is a homolog of Arabidopsis LEC1 was isolated in peanut. Its transcriptional start site confirmed by 5’ RACE was located at 82 nt from 5’ upstream of ATG. The bioinformatics analysis revealed that there existed many tissue-specific elements and light responsive motifs in its promoter. To identify the functional region of the AhLEC1A promoter, seven plant expression vectors expressing the GUS (β-glucuronidase) gene, driven by 5’ terminal series deleted fragments of AhLEC1A promoter, were constructed and transformed into Arabidopsis. Results of GUS histochemical staining showed that the regulatory region containing 82bp of 5’ UTR and 2228bp promoter could facilitate GUS to express preferentially in the embryos at different development periods of Arabidopsis. Taken together, it was inferred that the expression of AhLEC1A during seed development of peanut might be controlled positively by several seed-specific regulatory elements, as well as negatively by some other regulatory elements inhibiting its expression in other organs. Moreover, the GUS expression pattern of transgenic seedlings in darkness and in light was relevant to the light-responsive elements scattered in AhLEC1A promoter segment, implying that these light-responsive elements harbored in the AhLEC1A promoter regulate skotomorphogenesis of peanut seeds, and AhLEC1A expression was inhibited after the germinated seedlings were transferred from darkness to light.

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Expression Pattern, Regulation, and Biological Role of Runt Domain Transcription Factor, run, in Caenorhabditis elegans

Molecular and Cellular Biology ◽

10.1128/mcb.22.2.547-554.2002 ◽

2002 ◽

Vol 22 (2) ◽

pp. 547-554 ◽

Cited By ~ 40

Author(s):

Seunghee Nam ◽

Yun-Hye Jin ◽

Qing-Lin Li ◽

Kwang-Youl Lee ◽

Goo-Bo Jeong ◽

...

Keyword(s):

Transcription Factor ◽

Caenorhabditis Elegans ◽

Essential Role ◽

Regulatory Elements ◽

Intestinal Cells ◽

Human Leukemia ◽

Runt Domain ◽

C Elegans ◽

Intron Region

ABSTRACT The Caenorhabditis elegans run gene encodes a Runt domain factor. Runx1, Runx2, and Runx3 are the three known mammalian homologs of run. Runx1, which plays an essential role in hematopoiesis, has been identified at the breakpoint of chromosome translocations that are responsible for human leukemia. Runx2 plays an essential role in osteogenesis, and inactivation of one allele of Runx2 is responsible for the human disease cleidocranial dysplasia. To understand the role of run in C. elegans, we used transgenic run::GFP reporter constructs and a double-stranded RNA-mediated interference method. The expression of run was detected as early as the bean stage exclusively in the nuclei of seam hypodermal cells and lasted until the L3 stage. At the larval stage, expression of run was additionally detected in intestinal cells. The regulatory elements responsible for the postembryonic hypodermal seam cells and intestinal cells were separately located within a 7.2-kb-long intron region. This is the first report demonstrating that an intron region is essential for stage-specific and cell type-specific expression of a C. elegans gene. RNA interference analysis targeting the run gene resulted in an early larva-lethal phenotype, with apparent malformation of the hypodermis and intestine. These results suggest that run is involved in the development of a functional hypodermis and gut in C. elegans. The highly conserved role of the Runt domain transcription factor in gut development during evolution from nematodes to mammals is discussed.

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Conservation of glp-1 Regulation and Function in Nematodes

Genetics ◽

10.1093/genetics/157.2.639 ◽

2001 ◽

Vol 157 (2) ◽

pp. 639-654

Author(s):

David Rudel ◽

Judith Kimble

Keyword(s):

Stem Cells ◽

Phylogenetic Analysis ◽

Caenorhabditis Elegans ◽

Sequence Analysis ◽

Regulatory Elements ◽

Germline Stem Cells ◽

Biological Functions ◽

C Elegans ◽

Caenorhabditis Species ◽

And Function

Abstract The Caenorhabditis elegans (Ce) glp-1 gene encodes a Notch-like receptor. We have cloned glp-1 from C. briggsae (Cb) and C. remanei (Cr), two Caenorhabditis species that have diverged from C. elegans by roughly 20–40 million years. By sequence analysis, we find that the Cb-GLP-1 and Cr-GLP-1 proteins have retained the same motif architecture as Ce-GLP-1, including number of domains. In addition, two regions (CC-linker and regions flanking the ANK repeats) are as highly conserved as regions previously recognized as essential for signaling (e.g., ANK repeats). Phylogenetic analysis of glp-1 sequences suggests a C. briggsae/C. remanei clade with C. elegans as a sister taxon. Using RNAi to test biological functions, we find that Ce-glp-1, Cb-glp-1, and Cr-glp-1 are all required for proliferation of germline stem cells and for specifying blastomere fates in the embryo. In addition, certain biological roles of Cb-glp-1, e.g., in the vulva, have diverged from those of Ce-glp-1 and Cr-glp-1, suggesting a change in either regulation or function of the Cb-glp-1 gene during evolution. Finally, the regulation of glp-1 mRNA, previously analyzed for Ce-glp-1, is conserved in Cb-glp-1, and we identify conserved 3′ UTR sequences that may serve as regulatory elements.

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