scholarly journals Interallelic Complementation at the Mouse Mitf Locus

Genetics ◽  
2003 ◽  
Vol 163 (1) ◽  
pp. 267-276
Author(s):  
Eiríkur Steingrímsson ◽  
Heinz Arnheiter ◽  
Jón Hallsteinn Hallsson ◽  
M Lynn Lamoreux ◽  
Neal G Copeland ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Coat Color ◽  
Retinal Pigment ◽  
Cell Types ◽  
Binding Domain ◽  
Retinal Pigment Epithelial Cells ◽  
Dna Binding Domain ◽  
Allelic Series ◽  
Interallelic Complementation

Abstract Mutations at the mouse microphthalmia locus (Mitf) affect the development of different cell types, including melanocytes, retinal pigment epithelial cells of the eye, and osteoclasts. The MITF protein is a member of the MYC supergene family of basic-helix-loop-helix-leucine-zipper (bHLHZip) transcription factors and is known to regulate the expression of cell-specific target genes by binding DNA as homodimer or as heterodimer with related proteins. The many mutations isolated at the locus have different effects on the phenotype and can be arranged in an allelic series in which the phenotypes range from near normal to white microphthalmic animals with osteopetrosis. Previous investigations have shown that certain combinations of Mitf alleles complement each other, resulting in a phenotype more normal than that of each homozygote alone. Here we analyze this interallelic complementation in detail and show that it is limited to one particular allele, MitfMi-white (MitfMi-wh), a mutation affecting the DNA-binding domain. Both loss- and gain-of-function mutations are complemented, as are other Mitf mutations affecting the DNA-binding domain. Furthermore, this behavior is not restricted to particular cell types: Both eye development and coat color phenotypes are complemented. Our analysis suggests that MitfMi-wh-associated interallelic complementation is due to the unique biochemical nature of this mutation.

scholarly journals The Biological Impact of the Human Master Regulator p53 Can Be Altered by Mutations That Change the Spectrum and Expression of Its Target Genes

2006 ◽  
Vol 26 (6) ◽  
pp. 2297-2308 ◽  
Author(s):  
Daniel Menendez ◽  
Alberto Inga ◽  
Michael A. Resnick
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Tumor Development ◽  
Human Tumor ◽  
Human Cells ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Missense Mutations ◽  
The Relationship ◽  
Biological Outcomes

ABSTRACT Human tumor suppressor p53 is a sequence-specific master regulatory transcription factor that targets response elements (REs) in many genes. p53 missense mutations in the DNA-binding domain are often cancer associated. As shown with systems based on the yeast Saccharomyces cerevisiae, p53 mutants can alter the spectra and intensities of transactivation from individual REs. We address directly in human cells the relationship between changes in the p53 master regulatory network and biological outcomes. Expression of integrated, tightly regulated DNA-binding domain p53 mutants resulted in many patterns of apoptosis and survival following UV or ionizing radiation, or spontaneously. These patterns reflected changes in the spectra and activities of target genes, as demonstrated for P21, MDM2, BAX, and MSH2. Thus, as originally proposed for “master genes of diversity,” p53 mutations in human cells can differentially influence target gene transactivation, resulting in a variety of biological consequences which, in turn, might be expected to influence tumor development and therapeutic efficacy.

scholarly journals The carboxy-terminal transactivation domain of Oct-4 acquires cell specificity through the POU domain.

1997 ◽  
Vol 17 (1) ◽  
pp. 154-162 ◽  
Author(s):  
A Brehm ◽  
K Ohbo ◽  
H Schöler
Keyword(s):  
Dna Binding ◽  
Cell Lines ◽  
Regulatory Mechanism ◽  
Cell Types ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Cell Type ◽  
Pou Domain ◽  
Cell Type Specific ◽  
Carboxy Terminal

The POU transcription factor Oct-4 is expressed in totipotent and pluripotent cells of the early mouse embryo and the germ cell lineage. Transactivation capacities of regions flanking the DNA binding domain of Oct-4 were analyzed in undifferentiated and differentiated cell lines. The amino- and carboxy-terminal regions (N domain and C domain) fused to the Gal4 DNA binding domain both functioned as transactivation domains in all cell lines tested. However, the C domain failed to activate transcription in some cell lines in the context of the native protein. The underlying regulatory mechanism appears to involve the POU domain of Oct-4 and can discriminate between different POU domains, since constructs in which the C domain was instead fused to the POU domain of Pit-1 were again equally active in all cell lines. These results indicate that the C domain is subject to cell-type-specific regulation mediated by the Oct-4 POU domain. Phosphopeptide analysis revealed that the cell-type-specific difference of C-domain activity correlates with a difference in Oct-4 phosphorylation status. Since Oct-4 is expressed in a variety of distinct cell types during murine embryogenesis, these results suggest an additional regulatory mechanism for determining Oct-4 function in rapidly changing cell types during development.

Structure of the DNA-binding domain of the response regulator SaeR fromStaphylococcus aureus

2015 ◽  
Vol 71 (8) ◽  
pp. 1768-1776 ◽  
Author(s):  
Xiaojiao Fan ◽  
Xu Zhang ◽  
Yuwei Zhu ◽  
Liwen Niu ◽  
Maikun Teng ◽  
...  
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Response Regulator ◽  
Regulatory System ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Promoter Regions ◽  
Winged Helix

The SaeR/S two-component regulatory system is essential for controlling the expression of many virulence factors inStaphylococcus aureus. SaeR, a member of the OmpR/PhoB family, is a response regulator with an N-terminal regulatory domain and a C-terminal DNA-binding domain. In order to elucidate how SaeR binds to the promoter regions of target genes, the crystal structure of the DNA-binding domain of SaeR (SaeRDBD) was solved at 2.5 Å resolution. The structure reveals that SaeRDBDexists as a monomer and has the canonical winged helix–turn–helix module. EMSA experiments suggested that full-length SaeR can bind to the P1 promoter and that the binding affinity is higher than that of its C-terminal DNA-binding domain. Five key residues on the winged helix–turn–helix module were verified to be important for binding to the P1 promoterin vitroand for the physiological function of SaeRin vivo.

scholarly journals An Engineered PAX3-KRAB Transcriptional Repressor Inhibits the Malignant Phenotype of Alveolar Rhabdomyosarcoma Cells Harboring the Endogenous PAX3-FKHR Oncogene

2000 ◽  
Vol 20 (14) ◽  
pp. 5019-5031 ◽  
Author(s):  
William J. Fredericks ◽  
Kasirajan Ayyanathan ◽  
Meenhard Herlyn ◽  
Josh R. Friedman ◽  
Frank J. Rauscher
Keyword(s):  
Dna Binding ◽  
Dna Sequences ◽  
Transcriptional Repression ◽  
Target Genes ◽  
Binding Domain ◽  
Stable Expression ◽  
Alveolar Rhabdomyosarcoma ◽  
Dna Binding Domain ◽  
Malignant Phenotype ◽  
Activation Domain

ABSTRACT The t(2;13) chromosomal translocation in alveolar rhabdomyosarcoma tumors (ARMS) creates an oncogenic transcriptional activator by fusion of PAX3 DNA binding motifs to a COOH-terminal activation domain derived from the FKHR gene. The dominant oncogenic potential of the PAX3-FKHR fusion protein is dependent on the FKHR activation domain. We have fused the KRAB repression module to the PAX3 DNA binding domain as a strategy to suppress the activity of the PAX3-FKHR oncogene. The PAX3-KRAB protein bound PAX3 target DNA sequences and repressed PAX3-dependent reporter plasmids. Stable expression of the PAX3-KRAB protein in ARMS cell lines resulted in loss of the ability of the cells to grow in low-serum or soft agar and to form tumors in SCID mice. Stable expression of a PAX3-KRAB mutant, which lacks repression function, or a KRAB protein alone, lacking a PAX3 DNA binding domain, failed to suppress the ARMS malignant phenotype. These data suggest that the PAX3-KRAB repressor functions as a DNA-binding-dependent suppressor of the transformed phenotype of ARMS cells, probably via competition with the endogenous PAX3-FKHR oncogene and repression of target genes required for ARMS tumorigenesis. The engineered repressor approach that directs a transcriptional repression domain to target genes deregulated by the PAX3-FKHR oncogene may be a useful strategy to identify the target genes critical for ARMS tumorigenesis.

The COUP-TFII variant lacking a DNA-binding domain inhibits the activation of the Cyp7a1 promoter through physical interaction with COUP-TFII

2013 ◽  
Vol 452 (2) ◽  
pp. 345-357 ◽  
Author(s):  
Tomoko Yamazaki ◽  
Jun-ichi Suehiro ◽  
Hideki Miyazaki ◽  
Takashi Minami ◽  
Tatsuhiki Kodama ◽  
...  
Keyword(s):  
Dna Binding ◽  
Transcriptional Control ◽  
Target Genes ◽  
Binding Domain ◽  
Physical Interaction ◽  
Dna Binding Domain ◽  
Factor Ii ◽  
Huh7 Cells ◽  
Direct Binding ◽  
Cytochrome P450 Family

The COUP-TFII (chicken ovalbumin upstream promoter-transcription factor II) nuclear receptor, which is composed of a DNA-binding domain and a ligand-binding domain, exerts pleiotropic effects on development and cell differentiation by regulating the transcription of its target genes, including Cyp7a1 (cytochrome P450, family 7, subfamily a, polypeptide 1), which plays important roles in catabolism of cholesterol in the liver. Although multiple variants of COUP-TFII exist, their roles in the regulation of Cyp7a1 expression have not been elucidated. In the present study, we investigated the roles of COUP-TFII-V2 (variant 2), which lacks a DNA-binding domain, in the regulation of the transcriptional control of the Cyp7a1 gene by COUP-TFII in hepatocellular carcinoma cells. We found that COUP-TFII-V2 was significantly expressed in Huh7 cells, in which Cyp7a1 was not expressed. Furthermore, knockdown of COUP-TFII-V2 enhanced endogenous Cyp7a1 expression in Huh7 cells. Although COUP-TFII activates the Cyp7a1 promoter through direct binding to DNA, this activation was affected by COUP-TFII-V2, which physically interacted with COUP-TFII and inhibited its DNA-binding ability. Chromatin immunoprecipitation assays showed that COUP-TFII-V2 inhibited the binding of endogenous COUP-TFII to the intact Cyp7a1 promoter. The results of the present study suggest that COUP-TFII-V2 negatively regulates the function of COUP-TFII by inhibiting its binding to DNA to decrease Cyp7a1 expression.

scholarly journals B-Cell Coactivator OBF-1 Exhibits Unusual Transcriptional Properties and Functions in a DNA-Bound Oct-1-Dependent Fashion

1999 ◽  
Vol 19 (6) ◽  
pp. 4247-4254 ◽  
Author(s):  
Andrea Krapp ◽  
Michel Strubin
Keyword(s):  
Dna Binding ◽  
B Cell ◽  
Target Genes ◽  
Regulatory Elements ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Activation Domain ◽  
Activation Domains ◽  
Dna Binding Domains ◽  
Binding Domains

ABSTRACT Eukaryotic transcriptional activators generally comprise both a DNA-binding domain that recognizes specific cis-regulatory elements in the target genes and an activation domain which is essential for transcriptional stimulation. Activation domains typically behave as structurally and functionally autonomous modules that retain their intrinsic activities when directed to a promoter by a variety of heterologous DNA-binding domains. Here we report that OBF-1, a B-cell-specific coactivator for transcription factor Oct-1, challenges this traditional view in that it contains an atypical activation domain that exhibits two unexpected functional properties when tested in the yeast Saccharomyces cerevisiae. First, OBF-1 by itself has essentially no intrinsic activation potential, yet it strongly synergizes with other activation domains such as VP16 and Gal4. Second, OBF-1 exerts its effect in association with DNA-bound Oct-1 but is inactive when attached to a heterologous DNA-binding domain. These findings suggest that activation by OBF-1 is not obtained by simple recruitment of the coactivator to the promoter but requires interaction with DNA-bound Oct-1 to stimulate a step distinct from those regulated by classical activation domains.

scholarly journals Modulating the selective utilization of carbon sources by engineering the 3rd and 4th helices of the DNA-binding domain of catabolite control protein A (CcpA) in Bacillus licheniformis

2021 ◽  
Author(s):  
Yupeng Zhang ◽  
Youran Li ◽  
Fengxu Xiao ◽  
Hanrong Wang ◽  
Liang Zhang ◽  
...  
Keyword(s):  
Dna Binding ◽  
Bacillus Licheniformis ◽  
Target Genes ◽  
Protein A ◽  
Carbon Sources ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Catabolite Control Protein A ◽  
Control Protein ◽  
Catabolite Control

The gram-positive bacterium Bacillus licheniformis exhibits obvious selective utilization on carbon sources. This process is mainly governed by the global regulator catabolite control protein A (CcpA), which can recognize and bind to multiple target genes widely distributed in metabolic pathways. Although the DNA-binding domain of CcpA has been predicted, the influence of key amino acids on target gene recognition and binding remains elusive. In this study, the impact of Lys31, Ile42 and Leu56 on in vitro protein-DNA interactions and in vivo carbon source selective utilization was investigated. The results showed that alanine substitution of Lys31 and Ile42, located within the 3rd helices of the DNA-binding domain, significantly weakened the binding strength between CcpA and target genes. These mutations also lead to alleviated repression of xylose utilization in the presence of glucose. On the other hand, the Leu56Arg mutant in the 4th helices exhibited enhanced binding affinity compared with that of the wild-type one. When this mutant was used to replace the native one in B. licheniformis cells, the selective utilization of glucose over xylose increased. The above research results are helpful for a deep understanding of how microorganisms can flexibly sense and adapt to changes in the external environment. Additionally, they can provide important theoretical basis for the rational design of biomass utilization and environmental adaptability of B. licheniformiscell factories.

scholarly journals Sox10 Is an Active Nucleocytoplasmic Shuttle Protein, and Shuttling Is Crucial for Sox10-Mediated Transactivation

2002 ◽  
Vol 22 (16) ◽  
pp. 5826-5834 ◽  
Author(s):  
Stephan Rehberg ◽  
Peter Lischka ◽  
Gabi Glaser ◽  
Thomas Stamminger ◽  
Michael Wegner ◽  
...  
Keyword(s):  
Dna Binding ◽  
Nuclear Export ◽  
Target Genes ◽  
Nuclear Export Signal ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Nucleocytoplasmic Shuttling ◽  
Nucleocytoplasmic Shuttle ◽  
Nuclear Localization Signals ◽  
Export Signal

ABSTRACT Sox10 belongs to a family of transcription regulators characterized by a DNA-binding domain known as the HMG box. It plays fundamental roles in neural crest development, peripheral gliogenesis, and terminal differentiation of oligodendrocytes. In accord with its function as transcription factor, Sox10 contains two nuclear localization signals and is most frequently detected in the nucleus. In this study, we report that Sox10 is an active nucleocytoplasmic shuttle protein, competent of both entering and exiting the nucleus. We identified a functional Rev-type nuclear export signal within the DNA-binding domain of Sox10. Mutational inactivation of this nuclear export signal or treatment of cells with the CRM1-specific export inhibitor leptomycin B inhibited nuclear export and consequently nucleocytoplasmic shuttling of Sox10. Importantly, the inhibition of the nuclear export of Sox10 led to decreased transactivation of transfected reporters and endogenous target genes, arguing that continuous nucleocytoplasmic shuttling is essential for the function of Sox10. To our knowledge this is the first time that nuclear export has been reported and shown to be functionally relevant for any Sox protein.

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