scholarly journals Regulation of the human cardiac/slow-twitch troponin C gene by multiple, cooperative, cell-type-specific, and MyoD-responsive elements.

1993 ◽  
Vol 13 (11) ◽  
pp. 6752-6765 ◽  
Author(s):  
T H Christensen ◽  
H Prentice ◽  
R Gahlmann ◽  
L Kedes
Keyword(s):  
Skeletal Muscle ◽  
Heart Muscle ◽  
Regulatory Elements ◽  
Troponin C ◽  
Cell Type ◽  
Promoter Elements ◽  
Specific Expression ◽  
First Intron ◽  
Cell Type Specific ◽  
Slow Twitch

The cardiac troponin C (cTnC) gene produces identical transcripts in slow-twitch skeletal muscle and in heart muscle (R. Gahlmann, R. Wade, P. Gunning, and L. Kedes, J. Mol. Biol. 201:379-391, 1988). A separate gene encodes the fast-twitch skeletal muscle troponin C and is not expressed in heart muscle. We have used transient transfection to characterize the regulatory elements responsible for skeletal and cardiac cell-type-specific expression of the human cTnC (HcTnC) gene. At least four separate elements cooperate to confer tissue-specific expression of this gene in differentiated myotubes; a basal promoter (between -61 and -13) augments transcription 9-fold, upstream major regulatory sequences (between -68 and -142 and between -1319 and -4500) augment transcription as much as 39-fold, and at least two enhancer-like elements in the first intron (between +58 and +1028 and between +1029 and +1523) independently augment transcription 4- to 5-fold. These enhancers in the first intron increase myotube-specific chloramphenicol acetyltransferase activity when linked to their own promoter elements or to the heterologous simian virus 40 promoter, and the effects are multiplicative rather than additive. Each of the major myotube regulatory regions is capable of responding directly or indirectly to the myogenic determination factor, MyoD.A MyoD expression vector in 10T1/2 cells induced constructs carrying either the upstream HcTnC promoter elements or the first intron of the gene 300- to 500-fold. Expression was inhibited by cotransfection with Id, a negative regulator of basic helix-loop-helix transcription factors. The basal promoter contains five tandem TGGGC repeats that interact with Sp1 or an Sp1-like factor in nuclear extracts. Mutational analysis of this element demonstrated that two of the five repeat sequences were sufficient to support basal level muscle cell-specific transcription. Whereas the basal promoter is also critical for expression in cardiac myocytes, the elements upstream of -67 appear to play little or no role. Major augmentation of expression in cardiomyocytes is also provided by sequences in the first intron, but these are upstream (between +58 and +1028). The downstream segment of the first intron has no enhancer activity in cardiomyocytes. A specific DNA-protein complex is formed by this C2 cell enhancer with extracts from C2 cells but not cardiomyocytes. These observations suggest that tissue-specific expression of the HcTnC gene is cooperatively regulated by the complex interactions of multiple regulatory elements and that different elements are used to regulate expression in myogenic and cardiac cells.

Regulation of the human cardiac/slow-twitch troponin C gene by multiple, cooperative, cell-type-specific, and MyoD-responsive elements

1993 ◽  
Vol 13 (11) ◽  
pp. 6752-6765
Author(s):  
T H Christensen ◽  
H Prentice ◽  
R Gahlmann ◽  
L Kedes
Keyword(s):  
Skeletal Muscle ◽  
Heart Muscle ◽  
Regulatory Elements ◽  
Troponin C ◽  
Cell Type ◽  
Promoter Elements ◽  
Specific Expression ◽  
First Intron ◽  
Cell Type Specific ◽  
Slow Twitch

The cardiac troponin C (cTnC) gene produces identical transcripts in slow-twitch skeletal muscle and in heart muscle (R. Gahlmann, R. Wade, P. Gunning, and L. Kedes, J. Mol. Biol. 201:379-391, 1988). A separate gene encodes the fast-twitch skeletal muscle troponin C and is not expressed in heart muscle. We have used transient transfection to characterize the regulatory elements responsible for skeletal and cardiac cell-type-specific expression of the human cTnC (HcTnC) gene. At least four separate elements cooperate to confer tissue-specific expression of this gene in differentiated myotubes; a basal promoter (between -61 and -13) augments transcription 9-fold, upstream major regulatory sequences (between -68 and -142 and between -1319 and -4500) augment transcription as much as 39-fold, and at least two enhancer-like elements in the first intron (between +58 and +1028 and between +1029 and +1523) independently augment transcription 4- to 5-fold. These enhancers in the first intron increase myotube-specific chloramphenicol acetyltransferase activity when linked to their own promoter elements or to the heterologous simian virus 40 promoter, and the effects are multiplicative rather than additive. Each of the major myotube regulatory regions is capable of responding directly or indirectly to the myogenic determination factor, MyoD.A MyoD expression vector in 10T1/2 cells induced constructs carrying either the upstream HcTnC promoter elements or the first intron of the gene 300- to 500-fold. Expression was inhibited by cotransfection with Id, a negative regulator of basic helix-loop-helix transcription factors. The basal promoter contains five tandem TGGGC repeats that interact with Sp1 or an Sp1-like factor in nuclear extracts. Mutational analysis of this element demonstrated that two of the five repeat sequences were sufficient to support basal level muscle cell-specific transcription. Whereas the basal promoter is also critical for expression in cardiac myocytes, the elements upstream of -67 appear to play little or no role. Major augmentation of expression in cardiomyocytes is also provided by sequences in the first intron, but these are upstream (between +58 and +1028). The downstream segment of the first intron has no enhancer activity in cardiomyocytes. A specific DNA-protein complex is formed by this C2 cell enhancer with extracts from C2 cells but not cardiomyocytes. These observations suggest that tissue-specific expression of the HcTnC gene is cooperatively regulated by the complex interactions of multiple regulatory elements and that different elements are used to regulate expression in myogenic and cardiac cells.

scholarly journals Genetic influences on cell type specific gene expression and splicing during neurogenesis elucidate regulatory mechanisms of brain traits

2020 ◽  
Author(s):  
Nil Aygün ◽  
Angela L. Elwell ◽  
Dan Liang ◽  
Michael J. Lafferty ◽  
Kerry E. Cheek ◽  
...  
Keyword(s):  
Gene Expression ◽  
Regulatory Elements ◽  
Regulatory Mechanisms ◽  
Specific Gene ◽  
Cell Type ◽  
Specific Expression ◽  
Risk Variants ◽  
Allele Specific ◽  
Cell Type Specific ◽  
Regulatory Effects

SummaryInterpretation of the function of non-coding risk loci for neuropsychiatric disorders and brain-relevant traits via gene expression and alternative splicing is mainly performed in bulk post-mortem adult tissue. However, genetic risk loci are enriched in regulatory elements of cells present during neocortical differentiation, and regulatory effects of risk variants may be masked by heterogeneity in bulk tissue. Here, we map e/sQTLs and allele specific expression in primary human neural progenitors (n=85) and their sorted neuronal progeny (n=74). Using colocalization and TWAS, we uncover cell-type specific regulatory mechanisms underlying risk for these traits.

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 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 Two distinct auto-regulatory loops operate at the PU.1 locus in B cells and myeloid cells

Blood ◽  
2011 ◽  
Vol 117 (10) ◽  
pp. 2827-2838 ◽  
Author(s):  
Mathias Leddin ◽  
Chiara Perrod ◽  
Maarten Hoogenkamp ◽  
Saeed Ghani ◽  
Salam Assi ◽  
...  
Keyword(s):  
B Cells ◽  
Activity Patterns ◽  
Regulatory Elements ◽  
Specific Pattern ◽  
Cell Type ◽  
Hematopoietic Progenitors ◽  
Specific Expression ◽  
Targeted Deletion ◽  
Cis Element ◽  
Cell Type Specific

Abstract The transcription factor PU.1 occupies a central role in controlling myeloid and early B-cell development, and its correct lineage-specific expression is critical for the differentiation choice of hematopoietic progenitors. However, little is known of how this tissue-specific pattern is established. We previously identified an upstream regulatory cis element whose targeted deletion in mice decreases PU.1 expression and causes leukemia. We show here that the upstream regulatory cis element alone is insufficient to confer physiologic PU.1 expression in mice but requires the cooperation with other, previously unidentified elements. Using a combination of transgenic studies, global chromatin assays, and detailed molecular analyses we present evidence that PU.1 is regulated by a novel mechanism involving cross talk between different cis elements together with lineage-restricted autoregulation. In this model, PU.1 regulates its expression in B cells and macrophages by differentially associating with cell type–specific transcription factors at one of its cis-regulatory elements to establish differential activity patterns at other elements.

Regulation of the PU.1 gene by distal elements

Blood ◽  
2001 ◽  
Vol 98 (10) ◽  
pp. 2958-2965 ◽  
Author(s):  
Youlin Li ◽  
Yutaka Okuno ◽  
Pu Zhang ◽  
Hanna S. Radomska ◽  
Hui-min Chen ◽  
...  
Keyword(s):  
Cell Lines ◽  
Transcriptional Start Site ◽  
Critical Role ◽  
Myeloid Cell ◽  
Regulatory Elements ◽  
Cell Type ◽  
Specific Expression ◽  
Deoxyribonuclease I ◽  
Cell Type Specific Expression ◽  
Cell Type Specific

Abstract The transcription factor PU.1 (also known as Spi-1) plays a critical role in the development of the myeloid lineages, and myeloid cells derived from PU.1−/− animals are blocked at the earliest stage of myeloid differentiation. Expression of the PU.1 gene is tightly regulated during normal hematopoietic development, and dysregulation of PU.1 expression can lead to erythroleukemia. However, relatively little is known about how the PU.1 gene is regulated in vivo. Here it is shown that myeloid cell type–specific expression of PU.1 in stable cell lines and transgenic animals is conferred by a 91-kilobase (kb) murine genomic DNA fragment that consists of the entire PU.1 gene (20 kb) plus approximately 35 kb of upstream and downstream sequences, respectively. To further map the important transcriptional regulatory elements, deoxyribonuclease I hypersensitive site mapping studies revealed at least 3 clusters in the PU.1 gene. A 3.5-kb fragment containing one of these deoxyribonuclease I hypersensitive sites, located −14 kb 5′ of the transcriptional start site, conferred myeloid cell type–specific expression in stably transfected cell lines, suggesting that within this region is an element important for myeloid specific expression of PU.1. Further analysis of this myeloid-specific regulatory element will provide insight into the regulation of this key transcriptional regulator and may be useful as a tool for targeting expression to the myeloid lineage.

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