MyoD and myogenin act on the chicken myosin light-chain 1 gene as distinct transcriptional factors

1993 ◽  
Vol 13 (11) ◽  
pp. 7153-7162
Author(s):  
A Asakura ◽  
A Fujisawa-Sehara ◽  
T Komiya ◽  
Y Nabeshima ◽  
Y Nabeshima
Keyword(s):  
Light Chain ◽  
Dna Sequences ◽  
Regulatory Region ◽  
Target Sequence ◽  
Sequence Specificity ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Nonmuscle Cells ◽  
Flanking Sequences ◽  
E Boxes

Expression of MyoD, myogenin, MRF4, and Myf-5 converts nonmuscle cells to muscle cells. In an attempt to analyze the roles of these factors, we have investigated their effects on transcription driven by the promoter of the chicken myosin alkaline light-chain (MLC1) gene. The activation by CMD1 or c-myogenin (chicken MyoD or myogenin, respectively) was dependent on the existence of a muscle-specific regulatory region located from positions -2096 to -1743. Its distal half, containing a pair of E boxes (CANNTG), had been previously characterized as an enhancer responsive to CMD1 but not to c-myogenin. In this study, we report the identification of another enhancer in the muscle-specific regulatory region which is preferentially responsive to c-myogenin. Deletion and mutation analyses indicated that this enhancer requires a single E box and its flanking sequences. Furthermore, analysis of chimeric proteins of CMD1 and c-myogenin indicated that regions outside the basic helix-loop-helix domain of c-myogenin are involved in the specificity of the enhancer. These results show that CMD1 and c-myogenin act on the MLC1 gene by recognizing different upstream DNA sequences and that direct or indirect interactions between the regions outside the basic helix-loop-helix domain and flanking sequences of E boxes are involved in the target sequence specificity.

scholarly journals MyoD and myogenin act on the chicken myosin light-chain 1 gene as distinct transcriptional factors.

1993 ◽  
Vol 13 (11) ◽  
pp. 7153-7162 ◽  
Author(s):  
A Asakura ◽  
A Fujisawa-Sehara ◽  
T Komiya ◽  
Y Nabeshima ◽  
Y Nabeshima
Keyword(s):  
Light Chain ◽  
Dna Sequences ◽  
Regulatory Region ◽  
Target Sequence ◽  
Sequence Specificity ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Nonmuscle Cells ◽  
Flanking Sequences ◽  
E Boxes

Expression of MyoD, myogenin, MRF4, and Myf-5 converts nonmuscle cells to muscle cells. In an attempt to analyze the roles of these factors, we have investigated their effects on transcription driven by the promoter of the chicken myosin alkaline light-chain (MLC1) gene. The activation by CMD1 or c-myogenin (chicken MyoD or myogenin, respectively) was dependent on the existence of a muscle-specific regulatory region located from positions -2096 to -1743. Its distal half, containing a pair of E boxes (CANNTG), had been previously characterized as an enhancer responsive to CMD1 but not to c-myogenin. In this study, we report the identification of another enhancer in the muscle-specific regulatory region which is preferentially responsive to c-myogenin. Deletion and mutation analyses indicated that this enhancer requires a single E box and its flanking sequences. Furthermore, analysis of chimeric proteins of CMD1 and c-myogenin indicated that regions outside the basic helix-loop-helix domain of c-myogenin are involved in the specificity of the enhancer. These results show that CMD1 and c-myogenin act on the MLC1 gene by recognizing different upstream DNA sequences and that direct or indirect interactions between the regions outside the basic helix-loop-helix domain and flanking sequences of E boxes are involved in the target sequence specificity.

scholarly journals The Drosophila Basic Helix-Loop-Helix Protein DIMMED Directly Activates PHM, a Gene Encoding a Neuropeptide-Amidating Enzyme

2007 ◽  
Vol 28 (1) ◽  
pp. 410-421 ◽  
Author(s):  
Dongkook Park ◽  
Orie T. Shafer ◽  
Stacie P. Shepherd ◽  
Hyunsuk Suh ◽  
Jennifer S. Trigg ◽  
...  
Keyword(s):  
Cell Fate ◽  
Transcriptional Activation ◽  
Regulatory Region ◽  
Total Activity ◽  
Bhlh Protein ◽  
Hek 293 ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Boxes

ABSTRACT The basic helix-loop-helix (bHLH) protein DIMMED (DIMM) supports the differentiation of secretory properties in numerous peptidergic cells of Drosophila melanogaster. DIMM is coexpressed with diverse amidated neuropeptides and with the amidating enzyme peptidylglycine α-hydroxylating monooxygenase (PHM) in approximately 300 cells of the late embryo. Here we confirm that DIMM has transcription factor activity in transfected HEK 293 cells and that the PHM gene is a direct target. The mammalian DIMM orthologue MIST1 also transactivated the PHM gene. DIMM activity was dependent on the basic region of the protein and on the sequences of three E-box sites within PHM's first intron; the sites make different contributions to the total activity. These data suggest a model whereby the three E boxes interact cooperatively and independently to produce high PHM transcriptional activation. This DIMM-controlled PHM regulatory region displayed similar properties in vivo. Spatially, its expression mirrored that of the DIMM protein, and its activity was largely dependent on dimm. Further, in vivo expression was highly dependent on the sequences of the same three E boxes. This study supports the hypothesis that DIMM is a master regulator of a peptidergic cell fate in Drosophila and provides a detailed transcriptional mechanism of DIMM action on a defined target gene.

scholarly journals Differences between MyoD DNA binding and activation site requirements revealed by functional random sequence selection.

1996 ◽  
Vol 16 (7) ◽  
pp. 3893-3900 ◽  
Author(s):  
J Huang ◽  
T K Blackwell ◽  
L Kedes ◽  
H Weintraub
Keyword(s):  
Dna Binding ◽  
Binding Site ◽  
Dna Sequences ◽  
Transcriptional Activation ◽  
Random Sequence ◽  
In Vitro Binding ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
Vitro Binding

A method has been developed for selecting functional enhancer/promoter sites from random DNA sequences in higher eukaryotic cells. Of sequences that were thus selected for transcriptional activation by the muscle-specific basic helix-loop-helix protein MyoD, only a subset are similar to the preferred in vitro binding consensus, and in the same promoter context an optimal in vitro binding site was inactive. Other sequences with full transcriptional activity instead exhibit sequence preferences that, remarkably, are generally either identical or very similar to those found in naturally occurring muscle-specific promoters. This first systematic examination of the relation between DNA binding and transcriptional activation by basic helix-loop-helix proteins indicates that binding per se is necessary but not sufficient for transcriptional activation by MyoD and implies a requirement for other DNA sequence-dependent interactions or conformations at its binding site.

Characterization of transposable element-associated mutations that alter yeast alcohol dehydrogenase II expression

1983 ◽  
Vol 3 (1) ◽  
pp. 20-31
Author(s):  
V M Williamson ◽  
D Cox ◽  
E T Young ◽  
D W Russell ◽  
M Smith
Keyword(s):  
Alcohol Dehydrogenase ◽  
Dna Sequences ◽  
Target Sequence ◽  
Direct Repeats ◽  
Wild Type ◽  
Coding Region ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Delta Sequence ◽  
Flanking Sequences

Seven cis-dominant, constitutively expressed mutations of the normally glucose-repressible isozyme of alcohol dehydrogenase (ADHII) from the yeast Saccharomyces cerevisiae are caused by insertion of transposable elements from the Ty1 family in front of the ADHII structural gene (ADR2) (V. M. Williamson, E. T. Young, and M. Ciriacy, Cell 23:605-614, 1981). We cloned ADR2 with its associated Ty1 element from five S. cerevisiae strains carrying these mutations. Comparison of the Ty1 elements by heteroduplex studies and restriction enzyme analyses indicated that four were very similar; the fifth, although the same size as the others (about 5.6 kilobases), differed by the presence of two large substitutions of approximately 1 and 2 kilobases. The DNA sequences of the terminal direct repeats (deltas) were very homologous but not identical and were similar to previously reported Ty1 element direct repeats. We determined the 5'-flanking sequences of the ADR2 gene isolated from a wild-type strain and from five Ty1-associated mutations. The 5-base pair target sequence at the site of Ty1 insertion was present at both ends of each Ty1 element. The sites of insertion of the elements were all different and occurred from 125 to 210 base pairs in front of the coding region of ADR2. The 5' end of the major transcript as determined by S1 mapping was the same in wild-type cells and in Ty1-associated constitutive mutants and was approximately 54 base pairs upstream from the coding region. ADR2 transcripts were not detected when a solo delta sequence was present in the 5'-flanking region of this gene.

scholarly journals Multiple Basic Helix-Loop-Helix Proteins Regulate Expression of the ENO1 Gene of Saccharomyces cerevisiae

2007 ◽  
Vol 6 (5) ◽  
pp. 786-796 ◽  
Author(s):  
Meng Chen ◽  
John M. Lopes
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Dominant Negative ◽  
Bhlh Protein ◽  
Dominant Negative Mutant ◽  
Eukaryotic Transcription ◽  
Control Of Gene Expression ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Boxes ◽  
Bhlh Proteins

ABSTRACT The basic helix-loop-helix (bHLH) eukaryotic transcription factors have the ability to form multiple dimer combinations. This property, together with limited DNA-binding specificity for the E box (CANNTG), makes them ideally suited for combinatorial control of gene expression. We tested the ability of all nine Saccharomyces cerevisiae bHLH proteins to regulate the enolase-encoding gene ENO1. ENO1 was known to be activated by the bHLH protein Sgc1p. Here we show that expression of an ENO1-lacZ reporter was also regulated by the other eight bHLH proteins, namely, Ino2p, Ino4p, Cbf1p, Rtg1p, Rtg3p, Pho4p, Hms1p, and Ygr290wp. ENO1-lacZ expression was also repressed by growth in inositol-choline-containing medium. Epistatic analysis and chromatin immunoprecipitation experiments showed that regulation by Sgc1p, Ino2p, Ino4p, and Cbf1p and repression by inositol-choline required three distal E boxes, E1, E2, and E3. The pattern of bHLH binding to the three E boxes and experiments with two dominant-negative mutant alleles of INO4 and INO2 support the model that bHLH dimer selection affects ENO1-lacZ expression. These results support the general model that bHLH proteins can coordinate different biological pathways via multiple mechanisms.

scholarly journals Expression of the gene for Dec2, a basic helix–loop–helix transcription factor, is regulated by a molecular clock system

2004 ◽  
Vol 382 (1) ◽  
pp. 43-50 ◽  
Author(s):  
Hidenori HAMAGUCHI ◽  
Katsumi FUJIMOTO ◽  
Takeshi KAWAMOTO ◽  
Mitsuhide NOSHIRO ◽  
Koji MAEMURA ◽  
...  
Keyword(s):  
Negative Feedback ◽  
Feedback Loop ◽  
Molecular Mechanisms ◽  
Regulatory Protein ◽  
Circadian Oscillation ◽  
Negative Feedback Loop ◽  
Mrna Levels ◽  
Basic Helix Loop Helix ◽  
Helix Loop Helix ◽  
E Boxes

Dec2, a member of the basic helix–loop–helix superfamily, is a recently confirmed regulatory protein for the clockwork system. Transcripts of Dec2, as well as those of its related gene Dec1, exhibit a striking circadian oscillation in the suprachiasmatic nucleus, and Dec2 inhibits transcription from the Per1 promoter induced by Clock/Bmal1 [Honma, Kawamoto, Takagi, Fujimoto, Sato, Noshiro, Kato and Honma (2002) Nature (London) 419, 841–844]. It is known that mammalian circadian rhythms are controlled by molecular clockwork systems based on negative-feedback loop(s), but the molecular mechanisms for the circadian regulation of Dec2 gene expression have not been clarified. We show here that transcription of the Dec2 gene is regulated by several clock molecules and a negative-feedback loop. Luciferase and gel retardation assays showed that expression of Dec2 was negatively regulated by binding of Dec2 or Dec1 to two CACGTG E-boxes in the Dec2 promoter. Forced expression of Clock/Bmal1 and Clock/Bmal2 markedly increased Dec2 mRNA levels, and up-regulated the transcription of the Dec2 gene through the CACGTG E-boxes. Like Dec, Cry and Per also suppressed Clock/Bmal-induced transcription from the Dec2 promoter. Moreover, the circadian expression of Dec2 transcripts was abolished in the kidney of Clock/Clock mutant mice. These findings suggest that the Clock/Bmal heterodimer enhances Dec2 transcription via the CACGTG E-boxes, whereas the induced transcription is suppressed by Dec2, which therefore must contribute to its own rhythmic expression. In addition, Cry and Per may also modulate Dec2 transcription.

scholarly journals Characterization of transposable element-associated mutations that alter yeast alcohol dehydrogenase II expression.

1983 ◽  
Vol 3 (1) ◽  
pp. 20-31 ◽  
Author(s):  
V M Williamson ◽  
D Cox ◽  
E T Young ◽  
D W Russell ◽  
M Smith
Keyword(s):  
Alcohol Dehydrogenase ◽  
Dna Sequences ◽  
Target Sequence ◽  
Direct Repeats ◽  
Wild Type ◽  
Coding Region ◽  
Base Pairs ◽  
Yeast Saccharomyces Cerevisiae ◽  
Delta Sequence ◽  
Flanking Sequences

Seven cis-dominant, constitutively expressed mutations of the normally glucose-repressible isozyme of alcohol dehydrogenase (ADHII) from the yeast Saccharomyces cerevisiae are caused by insertion of transposable elements from the Ty1 family in front of the ADHII structural gene (ADR2) (V. M. Williamson, E. T. Young, and M. Ciriacy, Cell 23:605-614, 1981). We cloned ADR2 with its associated Ty1 element from five S. cerevisiae strains carrying these mutations. Comparison of the Ty1 elements by heteroduplex studies and restriction enzyme analyses indicated that four were very similar; the fifth, although the same size as the others (about 5.6 kilobases), differed by the presence of two large substitutions of approximately 1 and 2 kilobases. The DNA sequences of the terminal direct repeats (deltas) were very homologous but not identical and were similar to previously reported Ty1 element direct repeats. We determined the 5'-flanking sequences of the ADR2 gene isolated from a wild-type strain and from five Ty1-associated mutations. The 5-base pair target sequence at the site of Ty1 insertion was present at both ends of each Ty1 element. The sites of insertion of the elements were all different and occurred from 125 to 210 base pairs in front of the coding region of ADR2. The 5' end of the major transcript as determined by S1 mapping was the same in wild-type cells and in Ty1-associated constitutive mutants and was approximately 54 base pairs upstream from the coding region. ADR2 transcripts were not detected when a solo delta sequence was present in the 5'-flanking region of this gene.

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