scholarly journals Target Specificities of DrosophilaEnhancer of split Basic Helix-Loop-Helix Proteins

1999 ◽  
Vol 19 (7) ◽  
pp. 4600-4610 ◽  
Author(s):  
Barbara H. Jennings ◽  
David M. Tyler ◽  
Sarah J. Bray
Keyword(s):  
Dna Sequences ◽  
Target Genes ◽  
Notch Signalling Pathway ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Dna Binding Site ◽  
E Box ◽  
The Individual ◽  
Enhancer Of Split

ABSTRACT Seven Enhancer of split genes in Drosophila melanogaster encode basic-helix-loop-helix transcription factors which are components of the Notch signalling pathway. They are expressed in response to Notch activation and mediate some effects of the pathway by regulating the expression of target genes. Here we have determined that the optimal DNA binding site for the Enhancer of split proteins is a palindromic 12-bp sequence, 5′-TGGCACGTG(C/T)(C/T)A-3′, which contains an E-box core (CACGTG). This site is recognized by all of the individual Enhancer of split basic helix-loop-helix proteins, consistent with their ability to regulate similar target genes in vivo. We demonstrate that the 3 bp flanking the E-box core are intrinsic to DNA recognition by these proteins and that the Enhancer of split and proneural proteins can compete for binding on specific DNA sequences. Furthermore, the regulation conferred on a reporter gene in Drosophila by three closely related sequences demonstrates that even subtle sequence changes within an E box or flanking bases have dramatic consequences on the overall repertoire of proteins that can bind in vivo.

scholarly journals PU.1/Pip and Basic Helix Loop Helix Zipper Transcription Factors Interact With Binding Sites in the CD20 Promoter to Help Confer Lineage- and Stage-Specific Expression of CD20 in B Lymphocytes

Blood ◽  
1997 ◽  
Vol 90 (10) ◽  
pp. 3984-3995 ◽  
Author(s):  
Andreas Himmelmann ◽  
Agostino Riva ◽  
Gaye Lynn Wilson ◽  
Brian P. Lucas ◽  
Claire Thevenin ◽  
...  
Keyword(s):  
B Cell ◽  
Plasma Cells ◽  
Expression Vectors ◽  
Specific Gene ◽  
Cell Stage ◽  
Specific Expression ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Box

Abstract CD20 is a B-lineage–specific gene expressed at the pre–B-cell stage of B-cell development that disappears on differentiation to plasma cells. As such, it serves as an excellent paradigm for the study of lineage and developmental stage-specific gene expression. Using in vivo footprinting we identified two sites in the promoter at −45 and −160 that were occupied only in CD20+ B cells. The −45 site is an E box that binds basic helix-loop-helix-zipper proteins whereas the −160 site is a composite PU.1 and Pip binding site. Transfection studies with reporter constructs and various expression vectors verified the importance of these sites. The composite PU.1 and Pip site likely accounts for both lineage and stage-specific expression of CD20 whereas the CD20 E box binding proteins enhance overall promoter activity and may link the promoter to a distant enhancer.

scholarly journals The Basic Helix-Loop-Helix Transcription Factor NeuroD1 Facilitates Interaction of Sp1 with the Secretin Gene Enhancer

2007 ◽  
Vol 27 (22) ◽  
pp. 7839-7847 ◽  
Author(s):  
Subir K. Ray ◽  
Andrew B. Leiter
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Target Genes ◽  
Zinc Fingers ◽  
Homeodomain Proteins ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Gene Enhancer ◽  
Genes Encoding

ABSTRACT The basic helix-loop-helix transcription factor NeuroD1 is required for late events in neuronal differentiation, for maturation of pancreatic β cells, and for terminal differentiation of enteroendocrine cells expressing the hormone secretin. NeuroD1-null mice demonstrated that this protein is essential for expression of the secretin gene in the murine intestine, and yet it is a relatively weak transcriptional activator by itself. The present study shows that Sp1 and NeuroD1 synergistically activate transcription of the secretin gene. NeuroD1, but not its widely expressed dimerization partner E12, physically interacts with the C-terminal 167 amino acids of Sp1, which include its DNA binding zinc fingers. NeuroD1 stabilizes Sp1 DNA binding to an adjacent Sp1 binding site on the promoter to generate a higher-order DNA-protein complex containing both proteins and facilitates Sp1 occupancy of the secretin promoter in vivo. NeuroD-dependent transcription of the genes encoding the hormones insulin and proopiomelanocortin is potentiated by lineage-specific homeodomain proteins. The stabilization of binding of the widely expressed transcription factor Sp1 to the secretin promoter by NeuroD represents a distinct mechanism from other NeuroD target genes for increasing NeuroD-dependent transcription.

scholarly journals The Notch signalling pathway is required for Enhancer of split bHLH protein expression during neurogenesis in the Drosophila embryo

Development ◽  
1994 ◽  
Vol 120 (12) ◽  
pp. 3537-3548 ◽  
Author(s):  
B. Jennings ◽  
A. Preiss ◽  
C. Delidakis ◽  
S. Bray
Keyword(s):  
Protein Expression ◽  
Notch Signalling ◽  
Signalling Pathway ◽  
Drosophila Embryo ◽  
Protein Accumulation ◽  
Notch Signalling Pathway ◽  
Cell Fate Decisions ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Enhancer Of Split

The Enhancer of split locus is required during many cell-fate decisions in Drosophila, including the segregation of neural precursors in the embryo. We have generated monoclonal antibodies that recognise some of the basic helix-loop-helix proteins encoded by the Enhancer of split locus and have used them to examine expression of Enhancer of split proteins during neurogenesis. The proteins are expressed in a dynamic pattern in the ventral neurogenic region and are confined to those ectodermal cells that surround a neuroblast in the process of delaminating. There is no staining in the neuroblasts themselves. We have also examined the relationship between Enhancer of split protein accumulation and the Notch signalling pathway. Protein expression is abolished in a number of neurogenic mutant backgrounds, including Notch, but is increased as a result of expressing a constitutively active Notch product. We conclude that Notch signalling activity is directly responsible for the accumulation of basic helix-loop-helix proteins encoded by the Enhancer of split locus.

scholarly journals Targeting the Microphthalmia Basic Helix-Loop-Helix–Leucine Zipper Transcription Factor to a Subset of E-Box Elements In Vitro and In Vivo

1998 ◽  
Vol 18 (12) ◽  
pp. 6930-6938 ◽  
Author(s):  
I. Aksan ◽  
C. R. Goding
Keyword(s):  
Transcription Factor ◽  
Leucine Zipper ◽  
Pigment Cells ◽  
Specific Gene ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
P Gene ◽  
E Box

ABSTRACT The development of melanocytes, which are pigment-producing cells responsible for skin, hair, and eye color, is absolutely dependent on the action of the microphthalmia basic helix-loop-helix–leucine zipper (bHLH-LZ) transcription factor (Mi); mice lacking a functional Mi protein are entirely devoid of pigment cells. Mi has been shown to activate transcription of the tyrosinase,TRP-1, TRP-2, and QNR-71 genes through specific E-box elements, most notably the highly conserved M box. We investigated the mechanism which enables Mi to be recruited specifically to a restricted subset of E boxes in target promoters while being prevented from binding E-box elements in other promoters. We show both in vitro and in vivo that the presence of a T residue flanking a CATGTG E box is an essential determinant of the ability of Mi to bind DNA, and we successfully predict that the CATGTG E box from the P gene would not bind Mi. In contrast, no specific requirement for the sequences flanking a CACGTG E box was observed, and no binding to an atypical E box in the c-Kit promoter was detected. The relevance of these observations to the control of melanocyte-specific gene expression was highlighted by the fact that the E-box elements located in thetyrosinase, TRP-1, TRP-2, andQNR-71 promoters without exception possess a 5′ flanking T residue which is entirely conserved between species as diverse as man and turtle. The ability of Mi to discriminate between different E-box motifs provides a mechanism to restrict the repertoire of genes which are likely to be regulated by Mi and provides insight into the ability of bHLH-LZ transcription factors to achieve the specificity required for the precise coordination of transcription during development.

scholarly journals Selective utilization of basic helix-loop-helix-leucine zipper proteins at the immunoglobulin heavy-chain enhancer.

1997 ◽  
Vol 17 (1) ◽  
pp. 18-23 ◽  
Author(s):  
R S Carter ◽  
P Ordentlich ◽  
T Kadesch
Keyword(s):  
Heavy Chain ◽  
Leucine Zipper ◽  
Physiological Role ◽  
Immunoglobulin Heavy Chain ◽  
Dominant Negative ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Box ◽  
Derivatives Of

The microE3 E box within the immunoglobulin heavy-chain (IgH) enhancer binds several proteins of the basic helix-loop-helix-leucine zipper (bHLHzip) class, including TFE3, USF1, and Max. Both TFE3 and USF have been described as transcriptional activators, and so we investigated their possible roles in activating the IgH enhancer in vivo. Although TFE3 activated various enhancer-based reporters, both USF1 and Max effectively inhibited transcription. Inhibition by USF correlated with the lack of a strong activation domain and was the result of the protein neutralizing the microE3 site. The effects of dominant-negative derivatives of TFE3 and USF1 confirmed that TFE3, or a TFE3-like protein, is the primary cellular bHLHzip protein that activates the IgH enhancer. In addition to providing a physiological role for TFE3, our results call into question the traditional view of USF1 as an obligate transcriptional activator.

scholarly journals Calcium Regulation of Myogenesis by Differential Calmodulin Inhibition of Basic Helix-Loop-Helix Transcription Factors

2008 ◽  
Vol 19 (6) ◽  
pp. 2509-2519 ◽  
Author(s):  
Jannek Hauser ◽  
Juha Saarikettu ◽  
Thomas Grundström
Keyword(s):  
Transcription Factors ◽  
Target Genes ◽  
Myogenic Differentiation ◽  
Bhlh Protein ◽  
Channel Blockers ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Protein ◽  
Bhlh Transcription Factors

The members of the MyoD family of basic helix-loop-helix (bHLH) transcription factors are critical regulators of skeletal muscle differentiation that function as heterodimers with ubiquitously expressed E-protein bHLH transcription factors. These heterodimers must compete successfully with homodimers of E12 and other E-proteins to enable myogenesis. Here, we show that E12 mutants resistant to Ca2+-loaded calmodulin (CaM) inhibit MyoD-initiated myogenic conversion of transfected fibroblasts. Ca2+ channel blockers reduce, and Ca2+ stimulation increases, transcription by coexpressed MyoD and wild-type E12 but not CaM-resistant mutant E12. Furthermore, CaM-resistant E12 gives lower MyoD binding and higher E12 binding to a MyoD-responsive promoter in vivo and cannot rescue myogenic differentiation that has been inhibited by siRNA against E12 and E47. Our data support the concept that Ca2+-loaded CaM enables myogenesis by inhibiting DNA binding of E-protein homodimers, thereby promoting occupancy of myogenic bHLH protein/E-protein heterodimers on promoters of myogenic target genes.

scholarly journals Transcription enhancer factor 1 interacts with a basic helix-loop-helix zipper protein, Max, for positive regulation of cardiac alpha-myosin heavy-chain gene expression.

1997 ◽  
Vol 17 (7) ◽  
pp. 3924-3936 ◽  
Author(s):  
M P Gupta ◽  
C S Amin ◽  
M Gupta ◽  
N Hay ◽  
R Zak
Keyword(s):  
Myosin Heavy Chain ◽  
Reporter Gene ◽  
Heavy Chain ◽  
Troponin T ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Box ◽  
Two Factors ◽  
Transcription Enhancer ◽  
Enhancer Factor

The M-CAT binding factor transcription enhancer factor 1 (TEF-1) has been implicated in the regulation of several cardiac and skeletal muscle genes. Previously, we identified an E-box-M-CAT hybrid (EM) motif that is responsible for the basal and cyclic AMP-inducible expression of the rat cardiac alpha-myosin heavy chain (alpha-MHC) gene in cardiac myocytes. In this study, we report that two factors, TEF-1 and a basic helix-loop-helix leucine zipper protein, Max, bind to the alpha-MHC EM motif. We also found that Max was a part of the cardiac troponin T M-CAT-TEF-1 complex even when the DNA template did not contain an apparent E-box binding site. In the protein-protein interaction assay, a stable association of Max with TEF-1 was observed when glutathione S-transferase (GST)-TEF-1 or GST-Max was used to pull down in vitro-translated Max or TEF-1, respectively. In addition, Max was coimmunoprecipitated with TEF-1, thus documenting an in vivo TEF-1-Max interaction. In the transient transcription assay, overexpression of either Max or TEF-1 resulted a mild activation of the alpha-MHC-chloramphenicol acetyltransferase (CAT) reporter gene at lower concentrations and repression of this gene at higher concentrations. However, when Max and TEF-1 expression plasmids were transfected together, the repression mediated by a single expression plasmid was alleviated and a three- to fourfold transactivation of the alpha-MHC-CAT reporter gene was observed. This effect was abolished once the EM motif in the promoter-reporter construct was mutated, thus suggesting that the synergistic transactivation function of the TEF-1-Max heterotypic complex is mediated through binding of the complex to the EM motif. These results demonstrate a novel association between Max and TEF-1 and indicate a positive cooperation between these two factors in alpha-MHC gene regulation.

scholarly journals In vivo transcriptional regulation of N-Myc target genes is controlled by E-box methylation

2005 ◽  
Vol 102 (34) ◽  
pp. 12117-12122 ◽  
Author(s):  
G. Perini ◽  
D. Diolaiti ◽  
A. Porro ◽  
G. Della Valle
Keyword(s):  
Transcriptional Regulation ◽  
Target Genes ◽  
E Box

Scleraxis: a basic helix-loop-helix protein that prefigures skeletal formation during mouse embryogenesis

Development ◽  
1995 ◽  
Vol 121 (4) ◽  
pp. 1099-1110 ◽  
Author(s):  
P. Cserjesi ◽  
D. Brown ◽  
K.L. Ligon ◽  
G.E. Lyons ◽  
N.G. Copeland ◽  
...  
Keyword(s):  
Connective Tissue ◽  
Expression Pattern ◽  
Consensus Sequence ◽  
Cell Types ◽  
Cell Type ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Box ◽  
Cell Type Specific ◽  
Bhlh Proteins

Members of the basic helix-loop-helix (bHLH) family of transcription factors have been shown to regulate growth and differentiation of numerous cell types. Cell-type-specific bHLH proteins typically form heterodimers with ubiquitous bHLH proteins, such as E12, and bind a DNA consensus sequence known as an E-box. We used the yeast two-hybrid system to screen mouse embryo cDNA libraries for cDNAs encoding novel cell-type-specific bHLH proteins that dimerize with E12. One of the cDNAs isolated encoded a novel bHLH protein, called scleraxis. During mouse embryogenesis, scleraxis transcripts were first detected between day 9.5 and 10.5 post coitum (p.c.) in the sclerotome of the somites and in mesenchymal cells in the body wall and limb buds. Subsequently, scleraxis was expressed at high levels within mesenchymal precursors of the axial and appendicular skeleton and in cranial mesenchyme in advance of chondrogenesis; its expression pattern in these cell types foreshadowed the developing skeleton. Prior to formation of the embryonic cartilaginous skeleton, scleraxis expression declined to low levels. As development proceeded, high levels of scleraxis expression became restricted to regions where cartilage and connective tissue formation take place. Scleraxis bound the E-box consensus sequence as a heterodimer with E12 and activated transcription of a reporter gene linked to its DNA-binding site. The expression pattern, DNA-binding properties and transcriptional activity of scleraxis suggest that it is a regulator of gene expression within mesenchymal cell lineages that give rise to cartilage and connective tissue.

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