scholarly journals DNA Methylation Contributes to the Differential Expression Levels of Mecp2 in Male Mice Neurons and Astrocytes

2019 ◽  
Vol 20 (8) ◽  
pp. 1845 ◽  
Author(s):  
Vichithra R.B. Liyanage ◽  
Carl O. Olson ◽  
Robby M. Zachariah ◽  
James R. Davie ◽  
Mojgan Rastegar
Keyword(s):  
Dna Methylation ◽  
Current Knowledge ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Autism Spectrum ◽  
Brain Cells ◽  
Specific Expression ◽  
Expression Levels ◽  
Mecp2 Duplication Syndrome ◽  
Cell Type Specific Expression

Methyl CpG binding protein-2 (MeCP2) isoforms (E1 and E2) are important epigenetic regulators in brain cells. Accordingly, MeCP2 loss- or gain-of-function mutation causes neurodevelopmental disorders, including Rett syndrome (RTT), MECP2 duplication syndrome (MDS), and autism spectrum disorders (ASD). Within different types of brain cells, highest MeCP2 levels are detected in neurons and the lowest in astrocytes. However, our current knowledge of Mecp2/MeCP2 regulatory mechanisms remains largely elusive. It appears that there is a sex-dependent effect in X-linked MeCP2-associated disorders, as RTT primarily affects females, whereas MDS is found almost exclusively in males. This suggests that Mecp2 expression levels in brain cells might be sex-dependent. Here, we investigated the sex- and cell type-specific expression of Mecp2 isoforms in male and female primary neurons and astrocytes isolated from the murine forebrain. Previously, we reported that DNA methylation of six Mecp2 regulatory elements correlated with Mecp2 levels in the brain. We now show that in male brain cells, DNA methylation is significantly correlated with the transcript expression of these two isoforms. We show that both Mecp2 isoforms are highly expressed in male neurons compared to male astrocytes, with Mecp2e1 expressed at higher levels than Mecp2e2. Our data indicate that higher DNA methylation at the Mecp2 regulatory element(s) is associated with lower levels of Mecp2 isoforms in male astrocytes compared to male neurons.

scholarly journals Positive and negative elements regulate a melanocyte-specific promoter

1992 ◽  
Vol 12 (8) ◽  
pp. 3653-3662
Author(s):  
P Lowings ◽  
U Yavuzer ◽  
C R Goding
Keyword(s):  
Protein Interactions ◽  
Regulatory Element ◽  
Basal Layer ◽  
Regulatory Elements ◽  
Hair Follicles ◽  
Specific Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Cell Type Specific Expression ◽  
Cell Type Specific

Melanocytes are specialized cells residing in the hair follicles, the eye, and the basal layer of the human epidermis whose primary function is the production of the pigment melanin, giving rise to skin, hair, and eye color. Melanogenesis, a process unique to melanocytes that involves the processing of tyrosine by a number of melanocyte-specific enzymes, including tyrosinase and tyrosinase-related protein 1 (TRP-1), occurs only after differentiation from the melanocyte precursor, the melanoblast. In humans, melanogenesis is inducible by UV irradiation, with melanin being transferred from the melanocyte in the epidermis to the surrounding keratinocytes as protection from UV-induced damage. Excessive exposure to UV, however, is the primary cause of malignant melanoma, an increasingly common and highly aggressive disease. As an initial approach to understanding the regulation of melanocyte differentiation and melanocyte-specific transcription, we have isolated the gene encoding TRP-1 and examined the cis- and trans-acting factors required for cell-type-specific expression. We find that the TRP-1 promoter comprises both positive and negative regulatory elements which confer efficient expression in a TRP-1-expressing, pigmented melanoma cell line but not in NIH 3T3 or JEG3 cells and that a minimal promoter extending between -44 and +107 is sufficient for cell-type-specific expression. Assays for DNA-protein interactions coupled with extensive mutagenesis identified three factors, whose binding correlated with the function of two positive and one negative regulatory element. One of these factors, termed M-box-binding factor 1, binds to an 11-bp motif, the M box, which acts as a positive regulatory element both in TRP-1-expressing and -nonexpressing cell lines, despite being entirely conserved between the melanocyte-specific tyrosinase and TRP-1 promoters. The possible mechanisms underlying melanocyte-specific gene expression are discussed.

scholarly journals Positive and negative elements regulate a melanocyte-specific promoter.

1992 ◽  
Vol 12 (8) ◽  
pp. 3653-3662 ◽  
Author(s):  
P Lowings ◽  
U Yavuzer ◽  
C R Goding
Keyword(s):  
Protein Interactions ◽  
Regulatory Element ◽  
Basal Layer ◽  
Regulatory Elements ◽  
Hair Follicles ◽  
Specific Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Cell Type Specific Expression ◽  
Cell Type Specific

Melanocytes are specialized cells residing in the hair follicles, the eye, and the basal layer of the human epidermis whose primary function is the production of the pigment melanin, giving rise to skin, hair, and eye color. Melanogenesis, a process unique to melanocytes that involves the processing of tyrosine by a number of melanocyte-specific enzymes, including tyrosinase and tyrosinase-related protein 1 (TRP-1), occurs only after differentiation from the melanocyte precursor, the melanoblast. In humans, melanogenesis is inducible by UV irradiation, with melanin being transferred from the melanocyte in the epidermis to the surrounding keratinocytes as protection from UV-induced damage. Excessive exposure to UV, however, is the primary cause of malignant melanoma, an increasingly common and highly aggressive disease. As an initial approach to understanding the regulation of melanocyte differentiation and melanocyte-specific transcription, we have isolated the gene encoding TRP-1 and examined the cis- and trans-acting factors required for cell-type-specific expression. We find that the TRP-1 promoter comprises both positive and negative regulatory elements which confer efficient expression in a TRP-1-expressing, pigmented melanoma cell line but not in NIH 3T3 or JEG3 cells and that a minimal promoter extending between -44 and +107 is sufficient for cell-type-specific expression. Assays for DNA-protein interactions coupled with extensive mutagenesis identified three factors, whose binding correlated with the function of two positive and one negative regulatory element. One of these factors, termed M-box-binding factor 1, binds to an 11-bp motif, the M box, which acts as a positive regulatory element both in TRP-1-expressing and -nonexpressing cell lines, despite being entirely conserved between the melanocyte-specific tyrosinase and TRP-1 promoters. The possible mechanisms underlying melanocyte-specific gene expression are discussed.

scholarly journals The transcriptional tissue specificity of the human proα1(I) collagen gene is determined by a negative cis-regulatory element in the promoter

1992 ◽  
Vol 286 (1) ◽  
pp. 179-185 ◽  
Author(s):  
C P Simkevich ◽  
J P Thompson ◽  
H Poppleton ◽  
R Raghow
Keyword(s):  
Tissue Specificity ◽  
Regulatory Element ◽  
Mesenchymal Cell ◽  
Cell Types ◽  
Regulatory Elements ◽  
Negative Influence ◽  
Collagen Gene ◽  
Specific Expression ◽  
Cis Acting ◽  
First Intron

The transcriptional activity of plasmid pCOL-KT, in which human pro alpha 1 (I) collagen gene upstream sequences up to -804 and most of the first intron (+474 to +1440) drive expression of the chloramphenicol acetyltransferase (CAT) gene [Thompson, Simkevich, Holness, Kang & Raghow (1991) J. Biol. Chem. 266, 2549-2556], was tested in a number of mesenchymal and non-mesenchymal cells. We observed that pCOL-KT was readily expressed in fibroblasts of human (IMR-90 and HFL-1), murine (NIH 3T3) and avian (SL-29) origin and in a human rhabdomyosarcoma cell line (A204), but failed to be expressed in human erythroleukaemia (K562) and rat pheochromocytoma (PC12) cells, indicating that the regulatory elements required for appropriate tissue-specific expression of the human pro alpha 1 (I) collagen gene were present in pCOL-KT. To delineate the nature of cis-acting sequences which determine the tissue specificity of pro alpha 1 (I) collagen gene expression, functional consequences of deletions in the promoter and first intron of pCOL-KT were tested in various cell types by transient expression assays. Cis elements in the promoter-proximal and intronic sequences displayed either a positive or a negative influence depending on the cell type. Thus deletion of fragments using EcoRV (nt -625 to -442 deleted), XbaI (-804 to -331) or SstII (+670 to +1440) resulted in 2-10-fold decreased expression in A204 and HFL-1 cells. The negative influences of deletions in the promoter-proximal sequences was apparently considerably relieved by deleting sequences in the first intron, and the constructs containing the EcoRV/SstII or XbaI/SstII double deletions were expressed to a much greater extent than either of the single deletion constructs. In contrast, the XbaI* deletion (nt -804 to -609), either alone or in combination with the intronic deletion, resulted in very high expression in all cells regardless of their collagen phenotype; the XbaI*/(-SstII) construct, which contained the intronic SstII fragment (+670 to +1440) in the reverse orientation, was not expressed in either mesenchymal or nonmesenchymal cells. Based on these results, we conclude that orientation-dependent interactions between negatively acting 5′-upstream sequences and the first intron determine the mesenchymal cell specificity of human pro alpha 1 (I) collagen gene transcription.

Identification of upstream and intragenic regulatory elements that confer cell-type-restricted and differentiation-specific expression on the muscle creatine kinase gene

1988 ◽  
Vol 8 (7) ◽  
pp. 2896-2909 ◽  
Author(s):  
E A Sternberg ◽  
G Spizz ◽  
W M Perry ◽  
D Vizard ◽  
T Weil ◽  
...  
Keyword(s):  
Transcription Initiation ◽  
Regulatory Element ◽  
Simian Virus 40 ◽  
Muscle Cells ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Simian Virus ◽  
Cell Type ◽  
Specific Expression ◽  
Developing Muscle

Terminal differentiation of skeletal myoblasts is accompanied by induction of a series of tissue-specific gene products, which includes the muscle isoenzyme of creatine kinase (MCK). To begin to define the sequences and signals involved in MCK regulation in developing muscle cells, the mouse MCK gene has been isolated. Sequence analysis of 4,147 bases of DNA surrounding the transcription initiation site revealed several interesting structural features, some of which are common to other muscle-specific genes and to cellular and viral enhancers. To test for sequences required for regulated expression, a region upstream of the MCK gene from -4800 to +1 base pairs, relative to the transcription initiation site, was linked to the coding sequences of the bacterial chloramphenicol acetyltransferase (CAT) gene. Introduction of this MCK-CAT fusion gene into C2 muscle cells resulted in high-level expression of CAT activity in differentiated myotubes and no detectable expression in proliferating undifferentiated myoblasts or in nonmyogenic cell lines. Deletion mutagenesis of sequences between -4800 and the transcription start site showed that the region between -1351 and -1050 was sufficient to confer cell type-specific and developmentally regulated expression on the MCK promoter. This upstream regulatory element functioned independently of position, orientation, or distance from the promoter and therefore exhibited the properties of a classical enhancer. This upstream enhancer also was able to confer muscle-specific regulation on the simian virus 40 promoter, although it exhibited a 3- to 5-fold preference for its own promoter. In contrast to the cell type- and differentiation-specific expression of the upstream enhancer, the MCK promoter was able to function in myoblasts and myotubes and in nonmyogenic cell lines when combined with the simian virus 40 enhancer. An additional positive regulatory element was identified within the first intron of the MCK gene. Like the upstream enhancer, this intragenic element functioned independently of position, orientation, and distance with respect to the MCK promoter and was active in differentiated myotubes but not in myoblasts. These results demonstrate that expression of the MCK gene in developing muscle cells is controlled by complex interactions among multiple upstream and intragenic regulatory elements that are functional only in the appropriate cellular context.

Viral Strategies for Targeting the Central and Peripheral Nervous Systems

2018 ◽  
Vol 41 (1) ◽  
pp. 323-348 ◽  
Author(s):  
Claire N. Bedbrook ◽  
Benjamin E. Deverman ◽  
Viviana Gradinaru
Keyword(s):  
Nervous System ◽  
Regulatory Elements ◽  
Specific Cell ◽  
Cell Type ◽  
Specific Expression ◽  
Recombinant Viruses ◽  
Neuronal Populations ◽  
Neuroscience Research ◽  
Cell Type Specific Expression ◽  
Cell Type Specific

Recombinant viruses allow for targeted transgene expression in specific cell populations throughout the nervous system. The adeno-associated virus (AAV) is among the most commonly used viruses for neuroscience research. Recombinant AAVs (rAAVs) are highly versatile and can package most cargo composed of desired genes within the capsid's ∼5-kb carrying capacity. Numerous regulatory elements and intersectional strategies have been validated in rAAVs to enable cell type–specific expression. rAAVs can be delivered to specific neuronal populations or globally throughout the animal. The AAV capsids have natural cell type or tissue tropism and trafficking that can be modified for increased specificity. Here, we describe recently engineered AAV capsids and associated cargo that have extended the utility of AAVs in targeting molecularly defined neurons throughout the nervous system, which will further facilitate neuronal circuit interrogation and discovery.

scholarly journals Characterization of two atypical promoters and alternate mRNA processing in the mouse Thy-1.2 glycoprotein gene.

1986 ◽  
Vol 6 (8) ◽  
pp. 2923-2931 ◽  
Author(s):  
H A Ingraham ◽  
G A Evans
Keyword(s):  
Genomic Sequence ◽  
Regulatory Elements ◽  
Initiation Site ◽  
Lymphoid Cells ◽  
Gene Encoding ◽  
Specific Expression ◽  
Transcriptional Initiation ◽  
Glycoprotein Gene ◽  
Cell Type Specific Expression ◽  
Flanking Region

The promoter and 5' flanking region of the mouse Thy-1.2 glycoprotein gene were characterized by DNA sequencing, primer extension analysis, and deletion analysis. Transcriptional initiation sites were identified which corresponded to two separate exons upstream of the portion of the gene encoding the Thy-1.2 glycoprotein. We demonstrated that the mouse Thy-1.2 gene was transcribed from two atypical promoters separated by 260 base pairs in the genomic sequence. These promoters contained neither TATAAG nor GGPyCCAATCT homologous sequences but defined a conserved nonamer CTCCCTGCT at -48 from each initiation site. Two Thy-1.2 mRNA species of 1,835 and 1,939 nucleotides, differing in the 5' untranslated region of the mRNA, were thus transcribed from the single Thy-1.2 gene by mRNA splicing to the same downstream exon. Recombinant genomes in which the bacterial chloramphenicol acetyltransferase gene was expressed from either of the two Thy-1.2 promoters demonstrated that each promoter functioned independently and did not direct cell-specific expression in lymphoid cells. The 5' flanking region of the Thy-1.2 gene upstream of -68 could be eliminated without altering cell-type-specific expression. This suggests that regulatory elements responsible for tissue and developmental stage-specific expression of the Thy-1.2 gene are not present in the 5' flanking DNA but may reside downstream of the promoters.

scholarly journals Neurospora crassa 1,3-α-glucan synthase, AGS-1, is required for cell wall biosynthesis during macroconidia development

Microbiology ◽  
2014 ◽  
Vol 160 (8) ◽  
pp. 1618-1627 ◽  
Author(s):  
Ci Fu ◽  
Asuma Tanaka ◽  
Stephen J. Free
Keyword(s):  
Cell Wall ◽  
Neurospora Crassa ◽  
Fluorescent Protein ◽  
Regulatory Elements ◽  
Specific Expression ◽  
Glucan Synthase ◽  
Aerial Hyphae ◽  
Cell Type Specific Expression ◽  
Direct Cell ◽  
Cell Type Specific

The Neurospora crassa genome encodes two 1,3-α-glucan synthases. One of these 1,3-α-glucan synthase genes, ags-1, was shown to be required for the synthesis of 1,3-α-glucan in the aerial hyphae and macroconidia cell walls. 1,3-α-Glucan was found in the conidia cell wall, but was absent from the vegetative hyphae cell wall. Deletion of ags-1 affected conidial development. Δags-1 produced only 5 % as many conidia as the WT and most of the conidia produced by Δags-1 were not viable. The ags-1 upstream regulatory elements were shown to direct cell-type-specific expression of red fluorescent protein in conidia and aerial hyphae. A haemagglutinin-tagged AGS-1 was found to be expressed in aerial hyphae and conidia. The research showed that 1,3-α-glucan is an aerial hyphae and conidia cell wall component, and is required for normal conidial differentiation.

scholarly journals Developmental Dependence on NurRE and EboxNeuro for Expression of Pituitary Proopiomelanocortin

2008 ◽  
Vol 22 (7) ◽  
pp. 1647-1657 ◽  
Author(s):  
Pierre-Luc Lavoie ◽  
Lionel Budry ◽  
Aurélio Balsalobre ◽  
Jacques Drouin
Keyword(s):  
Signaling Pathways ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Portal System ◽  
Orphan Nuclear Receptors ◽  
Related Factors ◽  
Specific Expression ◽  
Pituitary Development ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix

Abstract Cell-specific expression of the pituitary proopiomelanocortin (POMC) gene depends on the combinatorial action of a large number of DNA-binding transcription factors (TFs). These include general and cell-restricted factors, as well as factors that act as effectors of signaling pathways. We have previously defined in the distal POMC promoter a composite regulatory element that contains targets for basic helix-loop-helix TFs conferring cell specificity and for NGFI-B orphan nuclear receptors that are responsive to CRH signaling and to glucocorticoid negative feedback. These factors act on neighboring regulatory elements, the EboxNeuro and NurRE, respectively. Currently, the EboxNeuro is thought to be the target of NeuroD1 during fetal development, but this factor may not account for activity in the adult pituitary; it is also unknown whether the NurRE and NGFI-B-related factors are active before establishment of the hypothalamic-pituitary portal system. In order to assess the importance of these regulatory elements and their cognate TFs throughout pituitary organogenesis and in the adult, we have assessed the activity of mutant POMC promoters in transgenic mice throughout development. These experiments indicate that the EboxNeuro and cognate basic helix-loop-helix factors are required throughout development and in the adult gland, beyond expression of NeuroD1. Similarly, the data reveal sustained importance of the NurRE and its cognate factors throughout pituitary development. These data contrast the sustained dependence throughout development on the same regulatory elements with the highly dynamic patterns of TF expression and the modulation of their activity in response to signaling pathways.

scholarly journals An internal regulatory element controls troponin I gene expression.

1989 ◽  
Vol 9 (4) ◽  
pp. 1397-1405 ◽  
Author(s):  
K E Yutzey ◽  
R L Kline ◽  
S F Konieczny
Keyword(s):  
Transcriptional Activation ◽  
Troponin I ◽  
Protein Gene ◽  
Regulatory Element ◽  
Consensus Sequence ◽  
Regulatory Elements ◽  
Contractile Protein ◽  
Specific Expression ◽  
Cis Acting ◽  
Specific Expression Pattern

During skeletal myogenesis, approximately 20 contractile proteins and related gene products temporally accumulate as the cells fuse to form multinucleated muscle fibers. In most instances, the contractile protein genes are regulated transcriptionally, which suggests that a common molecular mechanism may coordinate the expression of this diverse and evolutionarily unrelated gene set. Recent studies have examined the muscle-specific cis-acting elements associated with numerous contractile protein genes. All of the identified regulatory elements are positioned in the 5'-flanking regions, usually within 1,500 base pairs of the transcription start site. Surprisingly, a DNA consensus sequence that is common to each contractile protein gene has not been identified. In contrast to the results of these earlier studies, we have found that the 5'-flanking region of the quail troponin I (TnI) gene is not sufficient to permit the normal myofiber transcriptional activation of the gene. Instead, the TnI gene utilizes a unique internal regulatory element that is responsible for the correct myofiber-specific expression pattern associated with the TnI gene. This is the first example in which a contractile protein gene has been shown to rely primarily on an internal regulatory element to elicit transcriptional activation during myogenesis. The diversity of regulatory elements associated with the contractile protein genes suggests that the temporal expression of the genes may involve individual cis-trans regulatory components specific for each gene.

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