scholarly journals Identification and characterization of a neuroretina-specific enhancer element in the quail Pax-6 (Pax-QNR) gene.

1995 ◽  
Vol 15 (2) ◽  
pp. 892-903 ◽  
Author(s):  
S Plaza ◽  
C Dozier ◽  
M C Langlois ◽  
S Saule
Keyword(s):  
Specific Activity ◽  
Retinal Pigment ◽  
Retinal Pigment Epithelial Cells ◽  
Transcriptional Level ◽  
Regulatory Sequences ◽  
Independent Manner ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Enhancer Element

Using nuclear run-on assays, we showed that the tissue-specific expression of quail Pax-6 (Pax-QNR) P0-initiated mRNAs is due in part to regulation of the gene at the transcriptional level. Regulatory sequences governing neuroretina-specific expression of the P0-initiated mRNAs were investigated. By using reporter-based expression assays, we characterized a region within the Pax-QNR gene, located 7.5 kbp downstream from the P0 promoter, that functions as an enhancer in neuroretina cells but not in nonexpressing P0-initiated mRNA cells (quail embryo cells and quail retinal pigment epithelial cells). This enhancer element functioned in a position- and orientation-independent manner both on the Pax-QNR P0 promoter and the heterologous thymidine kinase promoter. Moreover, this enhancer element exhibited a developmental stage-specific activity during embryonic neuroretina development: in contrast to activity at day E7, the enhancer activity was very weak at day E5. This paralleled the level of expression of P0-initiated mRNAs observed at the same stages. Using footprinting, gel retardation, and Southwestern (DNA-protein) analysis, we demonstrated the existence of four neuroretina-specific nuclear protein-binding sites, involving multiple unknown factors. In addition we showed that the quail enhancer element is structurally and functionally conserved in mice. All of these results strongly suggest that this enhancer element may contribute to the neuroretina-specific transcriptional regulation of the Pax-6 gene in vivo.

scholarly journals Dissection of a Complex Enhancer Element: Maintenance of Keratinocyte Specificity but Loss of Differentiation Specificity

2002 ◽  
Vol 22 (12) ◽  
pp. 4293-4308 ◽  
Author(s):  
Charles K. Kaufman ◽  
Satrajit Sinha ◽  
Diana Bolotin ◽  
Jie Fan ◽  
Elaine Fuchs
Keyword(s):  
Gene Expression ◽  
Specific Activity ◽  
Regulatory Elements ◽  
Specific Gene ◽  
Regulatory Sequence ◽  
Open Chromatin ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Enhancer Element

ABSTRACT In this report, we explored the mechanisms underlying keratinocyte-specific and differentiation-specific gene expression in the skin. We have identified five keratinocyte-specific, open chromatin regions that exist within the 6 kb of 5′ upstream regulatory sequence known to faithfully recapitulate the strong endogenous keratin 5 (K5) promoter and/or enhancer activity. One of these, DNase I-hypersensitive site (HSs) 4, was unique in that it acted independently to drive abundant and keratinocyte-specific reporter gene activity in culture and in transgenic mice, despite the fact that it was not essential for K5 enhancer activity. We have identified evolutionarily conserved regulatory elements and a number of their associated proteins that bind to this compact and complex enhancer element. The 125-bp 3′ half of this element (referred to as 4.2) is by far the smallest known strong enhancer element possessing keratinocyte-specific activity in vivo. Interestingly, its activity is restricted to a subset of progeny of K5-expressing cells located within the sebaceous gland. The other half of HSs 4 (termed 4.1) possesses activity to suppress sebocyte-specific expression and induce expression in the channel (inner root sheath) cells surrounding the hair shaft. Our findings lead us to a view of keratinocyte gene expression which is determined by multiple regulatory modules, many of which contain AP-2 and/or Sp1/Sp3 binding sites for enhancing expression in skin epithelium, but which also harbor one or more unique sites for the binding of factors which determine specificity. Through mixing and matching of these modules, additional levels of specificity are obtained, indicating that both transcriptional repressors and activators govern the specificity.

Segmental expression of the EphA4 (Sek-1) receptor tyrosine kinase in the hindbrain is under direct transcriptional control of Krox-20

Development ◽  
1998 ◽  
Vol 125 (3) ◽  
pp. 443-452 ◽  
Author(s):  
T. Theil ◽  
M. Frain ◽  
P. Gilardi-Hebenstreit ◽  
A. Flenniken ◽  
P. Charnay ◽  
...  
Keyword(s):  
Tyrosine Kinase ◽  
Receptor Tyrosine Kinase ◽  
Binding Sites ◽  
Cell Signalling ◽  
Ectopic Expression ◽  
Regulatory Sequences ◽  
Kinase Gene ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Enhancer Element

Segmentation of the vertebrate hindbrain leads to the formation of a series of rhombomeres (r) with distinct identities. Recent studies have uncovered regulatory links between transcription factors governing this process, but little is known of how these relate to molecules mediating cell-cell signalling. The Eph receptor tyrosine kinase gene EphA4 (Sek-1) is expressed in r3 and r5, and function-blocking experiments suggest that it is involved in restricting intermingling of cells between odd- and even-numbered rhombomeres. We have analysed the cis-acting regulatory sequences of the EphA4 gene in transgenic mice and identified a 470 bp enhancer element that drives specific expression in r3 and r5. Within this element, we have identified eight binding sites for the Krox-20 transcription factor that is also expressed in r3 and r5. Mutation of these binding sites abolishes r3/r5 enhancer activity and ectopic expression of Krox-20 leads to ectopic activation of the enhancer. These data indicate that Krox-20 is a direct transcriptional activator of EphA4. Together with evidence that Krox-20 regulates Hox gene expression, our findings reveal a mechanism by which the identity and movement of cells are coupled such that sharply restricted segmental domains are generated.

Cyanidin-3-glucoside attenuates 4-hydroxynonenal- and visible light-induced retinal damage in vitro and in vivo

2019 ◽  
Vol 10 (5) ◽  
pp. 2871-2880 ◽  
Author(s):  
Yong Wang ◽  
Wentao Qi ◽  
Yazhen Huo ◽  
Ge Song ◽  
Hui Sun ◽  
...  
Keyword(s):  
Retinal Pigment Epithelial ◽  
Retinal Pigment ◽  
Retinal Pigment Epithelial Cells ◽  
Retinal Damage ◽  
Protective Effects ◽  
Pigment Epithelial ◽  
Induced Apoptosis ◽  
Pigment Epithelial Cells

Cyanidin-3-glucoside has efficient protective effects on 4-hydroxynonenal-induced apoptosis, senescence, and angiogenesis in retinal pigment epithelial cells.

scholarly journals Direct repeats bind the EcR/USP receptor and mediate ecdysteroid responses in Drosophila melanogaster.

1996 ◽  
Vol 16 (6) ◽  
pp. 2977-2986 ◽  
Author(s):  
C Antoniewski ◽  
B Mugat ◽  
F Delbac ◽  
J A Lepesant
Keyword(s):  
Fat Body ◽  
Cell Transformation ◽  
Direct Repeat ◽  
Transgenic Animals ◽  
Transcriptional Level ◽  
Regulatory Sequences ◽  
Direct Repeats ◽  
Fbp1 Gene

The steroid hormone 20-hydroxyecdysone plays a key role in the induction and modulation of morphogenetic events throughout Drosophila development. Previous studies have shown that a heterodimeric nuclear receptor composed of the EcR and USP proteins mediates the action of the hormone at the transcriptional through binding to palindromic ecdysteroid mediates the action of the hormone at the transcriptional level through binding to palindromic ecdysteroid response elements (EcREs) such as those present in the promoter of the hsp27 gene or the fat body-specific enhancer of the Fbp1 gene. We show that in addition to palindromic EcREs, the EcR/USP heterodimer can bind in vitro with various affinities to direct repetitions of the motif AGGTCA separated by 1 to 5 nucleotides (DR1 to DR5), which are known to be target sites for vertebrate nuclear receptors. At variance with the receptors, EcR/USP was also found to bind to a DR0 direct repeat with no intervening nucleotide. In cell transformation assays, direct repeats DR0 to DR5 alone can render the minimum viral tk or Drosophila Fbp1 promoter responsive to 20-hydroxyecdysone, as does the palindromic hsp27 EcRE. In a transgenic assay, however, neither the palindromic hsp27 element nor direct repeat DR3 alone can make the Fbp1 minimal promoter responsive to premetamorphic ecdysteroid peaks. In contrast, DR0 and DR3 elements, when substituted for the natural palindromic EcRE in the context of the Fbp1 enhancer, can drive a strong fat body-specific ecdysteroid response in transgenic animals. These results demonstrate that directly repeated EcR/USP binding sites are as effective as palindromic EcREs in vivo. They also provide evidence that additional flanking regulatory sequences are crucially required to potentiate the hormonal response mediated by both types of elements and specify its spatial and temporal pattern.

Overlapping positive and negative regulatory elements determine lens-specific activity of the delta 1-crystallin enhancer

1993 ◽  
Vol 13 (9) ◽  
pp. 5206-5215 ◽  
Author(s):  
Y Kamachi ◽  
H Kondoh
Keyword(s):  
Mutational Analysis ◽  
Specific Activity ◽  
Regulatory Elements ◽  
Positive Element ◽  
Specific Expression ◽  
Enhancer Activity ◽  
Negative Element ◽  
Positive Elements ◽  
Lens Cells ◽  
Specific Regulation

Lens-specific expression of the delta 1-crystallin gene is governed by an enhancer in the third intron, and the 30-bp-long DC5 fragment was found to be responsible for eliciting the lens-specific activity. Mutational analysis of the DC5 fragment identified two contiguous, interdependent positive elements and a negative element which overlaps the 3'-located positive element. Previously identified ubiquitous factors delta EF1 bound to the negative element and repressed the enhancer activity in nonlens cells. Mutation and cotransfection analyses indicated the existence of an activator which counteracts the action of delta EF1 in lens cells, probably through binding site competition. We also found a group of nuclear factors, collectively called delta EF2, which bound to the 5'-located positive element. delta EF2a and -b were the major species in lens cells, whereas delta EF2c and -d predominated in nonlens cells. These delta EF2 proteins probably cooperate with factors bound to the 3'-located element in activation in lens cells and repression in nonlens cells. delta EF2 proteins also bound to a promoter sequence of the gamma F-crystallin gene, suggesting that delta EF2 proteins are involved in lens-specific regulation of various crystallin classes.

A novel regulatory element between the human FGA and FGG genes

2012 ◽  
Vol 108 (09) ◽  
pp. 427-434 ◽  
Author(s):  
Richard J. Fish ◽  
Marguerite Neerman-Arbez
Keyword(s):  
Gene Cluster ◽  
Fluorescent Protein ◽  
Regulatory Element ◽  
Luciferase Reporter ◽  
Transgenic Zebrafish ◽  
Regulatory Sequences ◽  
Gene Promoters ◽  
Cvd Risk ◽  
Enhancer Activity

SummaryHigh circulating fibrinogen levels correlate with cardiovascular disease (CVD) risk. Fibrinogen levels vary between people and also change in response to physiological and environmental stimuli. A modest proportion of the variation in fibrinogen levels can be explained by genotype, inferring that variation in genomic sequences that regulate the fibri-nogen genes (FGA, FGB and FGG) may affect hepatic fibrinogen production and perhaps CVD risk. We previously identified a conserved liver enhancer in the fibrinogen gene cluster (CNC12), between FGB and FGA. Genome-wide Chromatin immunoprecipitation-sequencing (ChIP-seq) demonstrated that transcription factors which bind fibrinogen gene promoters also interact with CNC12, as well as two potential fibrinogen enhancers (PFE), between FGA and FGG. Here we show that one of the PFE sequences has potent hepatocyte enhancer activity. Using a luciferase reporter gene system, we found that PFE2 enhances minimal promoter- and FGA promoter-driven gene expression in hepatoma cells, regardless of its orientation with respect to the promoters. A region within PFE2 bears a short series of conserved nucleotides which maintain enhancer activity without flanking sequence. We also demonstrate that PFE2 is a liver enhancer in vivo, driving enhanced green fluorescent protein expression in transgenic zebrafish larval livers. Our study shows that combining public domain ChIP-seq data with in vitro and in vivo functional tests can identify novel fibrinogen gene cluster regulatory sequences. Variation in such elements could affect fibrinogen production and influence CVD risk.

Regulation of the twist target gene tinman by modular cis-regulatory elements during early mesoderm development

Development ◽  
1997 ◽  
Vol 124 (24) ◽  
pp. 4971-4982 ◽  
Author(s):  
Z. Yin ◽  
X.L. Xu ◽  
M. Frasch
Keyword(s):  
Homeobox Gene ◽  
Regulatory Elements ◽  
Bhlh Protein ◽  
Expression Domain ◽  
Enhancer Activity ◽  
Enhancer Element ◽  
Mesoderm Development ◽  
Visceral Mesoderm

The Drosophila tinman homeobox gene has a major role in early mesoderm patterning and determines the formation of visceral mesoderm, heart progenitors, specific somatic muscle precursors and glia-like mesodermal cells. These functions of tinman are reflected in its dynamic pattern of expression, which is characterized by initial widespread expression in the trunk mesoderm, then refinement to a broad dorsal mesodermal domain, and finally restricted expression in heart progenitors. Here we show that each of these phases of expression is driven by a discrete enhancer element, the first being active in the early mesoderm, the second in the dorsal mesoderm and the third in cardioblasts. We provide evidence that the early-active enhancer element is a direct target of twist, a gene encoding a basic helix-loop-helix (bHLH) protein, which is necessary for tinman activation. This 180 bp enhancer includes three E-box sequences which bind Twist protein in vitro and are essential for enhancer activity in vivo. Ectodermal misexpression of twist causes ectopic activation of this enhancer in ectodermal cells, indicating that twist is the only mesoderm-specific activator of early tinman expression. We further show that the 180 bp enhancer also includes negatively acting sequences. Binding of Even-skipped to these sequences appears to reduce twist-dependent activation in a periodic fashion, thus producing a striped tinman pattern in the early mesoderm. In addition, these sequences prevent activation of tinman by twist in a defined portion of the head mesoderm that gives rise to hemocytes. We find that this repression requires the function of buttonhead, a head-patterning gene, and that buttonhead is necessary for normal activation of the hematopoietic differentiation gene serpent in the same area. Together, our results show that tinman is controlled by an array of discrete enhancer elements that are activated successively by differential genetic inputs, as well as by closely linked activator and repressor binding sites within an early-acting enhancer, which restrict twist activity to specific areas within the twist expression domain.

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