scholarly journals The pancreatic islet factor STF-1 binds cooperatively with Pbx to a regulatory element in the somatostatin promoter: importance of the FPWMK motif and of the homeodomain.

1995 ◽  
Vol 15 (12) ◽  
pp. 7091-7097 ◽  
Author(s):  
B Peers ◽  
S Sharma ◽  
T Johnson ◽  
M Kamps ◽  
M Montminy
Keyword(s):  
Target Genes ◽  
Gene Selection ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Cooperative Binding ◽  
Homeodomain Protein ◽  
Homeodomain Proteins ◽  
Target Sites

A number of homeodomain proteins have been shown to regulate cellular development by stimulating the transcription of specific target genes. In contrast to their distinct activities in vivo, however, most homeodomain proteins bind indiscriminately to potential target sites in vitro, suggesting the involvement of cofactors which specify target site selection. One such cofactor, termed extradenticle, has been shown to influence segmental morphogenesis in Drosophila melanogaster by binding cooperatively with certain homeodomain proteins to target regulatory elements. Here we demonstrate that STF-1, an orphan homeodomain protein required for pancreatic development in mammals, binds cooperatively to DNA with Pbx, the mammalian homolog of extradenticle. Cooperative binding with Pbx requires a pentapeptide motif (FPWMK) which is well conserved among a large subset of homeodomain proteins. The FPMWK motif is not sufficient to confer Pbx cooperativity on other homeodomain proteins, however; the N-terminal arm of the STF-1 homeodomain is also essential. As cooperative binding with Pbx occurs on only a subset of potential STF-1 target sites, our results suggest that Pbx may specify target gene selection in the developing pancreas by forming heterodimeric complexes with STF-1.

scholarly journals The human FLT1 regulatory element directs vascular expression and modulates angiogenesis pathways in vitro and in vivo

2021 ◽  
Author(s):  
Julian Stolper ◽  
Holly K. Voges ◽  
Michael See ◽  
Neda Rahmani Mehdiabadi ◽  
Gulrez Chahal ◽  
...  
Keyword(s):  
Transgene Expression ◽  
Regulatory Element ◽  
Embryonic Stem ◽  
Regulatory Elements ◽  
Homeodomain Protein ◽  
Cardiovascular Development ◽  
Vascular Expression

AbstractThere is growing evidence that mutations in non-coding cis-regulatory elements (CREs) disrupt proper development. However, little is known about human CREs that are crucial for cardiovascular development. To address this, we bioinformatically identified cardiovascular CREs based on the occupancy of the CRE by the homeodomain protein NKX2-5 and cardiac chromatin histone modifications. This search defined a highly conserved CRE within the FLT1 locus termed enFLT1. We show that the human enFLT1 is an enhancer capable of driving reporter transgene expression in vivo throughout the developing cardiovascular system of medaka. Deletion of the human enFLT1 enhancer (ΔenFLT1) triggered molecular perturbations in extracellular matrix organisation and blood vessel morphogenesis in vitro in endothelial cells derived from human embryonic stem cells and vascular defects in vivo in medaka. These findings highlight the crucial role of the human FLT1 enhancer and its function as a regulator and buffer of transcriptional regulation in cardiovascular development.

scholarly journals Regulation by Homeoproteins: A Comparison of Deformed-Responsive Elements

Genetics ◽  
2000 ◽  
Vol 156 (2) ◽  
pp. 677-686
Author(s):  
Jeffrey A Pederson ◽  
James W LaFollette ◽  
Cornelius Gross ◽  
Alexey Veraksa ◽  
William McGinnis ◽  
...  
Keyword(s):  
Binding Sites ◽  
Target Genes ◽  
Gene Selection ◽  
Consensus Sequence ◽  
Cooperative Binding ◽  
Homeotic Genes ◽  
Specific Expression ◽  
Enhancer Element

Abstract Homeotic genes of Drosophila melanogaster encode transcription factors that specify segment identity by activating the appropriate set of target genes required to produce segment-specific characteristics. Advances in understanding target gene selection have been hampered by the lack of genes known to be directly regulated by the HOM-C proteins. Here we present evidence that the gene 1.28 is likely to be a direct target of Deformed in the maxillary segment. We identified a 664-bp Deformed Response Element (1.28 DRE) that directs maxillary-specific expression of a reporter gene in transgenic embryos. The 1.28 DRE contains in vitro binding sites for Deformed and DEAF-1. The Deformed binding sites do not have the consensus sequence for cooperative binding with the cofactor Extradenticle, and we do not detect cooperative binding to these sites, though we cannot rule out an independent role for Extradenticle. Removing the four Deformed binding sites renders the 1.28 DRE inactive in vivo, demonstrating that these sites are necessary for activation of this enhancer element, and supporting the proposition that 1.28 is activated by Deformed. We show that the DEAF-1 binding region is not required for enhancer function. Comparisons of the 1.28 DRE with other known Deformed-responsive enhancers indicate that there are multiple ways to construct Deformed Response Elements.

Switching the in vivo specificity of a minimal Hox-responsive element

Development ◽  
1997 ◽  
Vol 124 (10) ◽  
pp. 2007-2014 ◽  
Author(s):  
S.K. Chan ◽  
H.D. Ryoo ◽  
A. Gould ◽  
R. Krumlauf ◽  
R.S. Mann
Keyword(s):  
Binding Sites ◽  
Target Genes ◽  
Expression Patterns ◽  
High Specificity ◽  
Mouse Embryos ◽  
Homeodomain Protein ◽  
Homeodomain Proteins ◽  
Responsive Element

The homeodomain proteins encoded by the Hox complex genes do not bind DNA with high specificity. In vitro, Hox specificity can be increased by binding to DNA cooperatively with the homeodomain protein extradenticle or its vertebrate homologs, the pbx proteins (together, the PBC family). Here we show that a two basepair change in a Hox-PBC binding site switches the Hox-dependent expression pattern generated in vivo, from labial to Deformed. The change in vivo correlates with an altered Hox binding specificity in vitro. Further, we identify similar Deformed-PBC binding sites in the Deformed and Hoxb-4 genes and show that they generate Deformed or Hoxb-4 expression patterns in Drosophila and mouse embryos, respectively. These results suggest a model in which Hox-PBC binding sites play an instructive role in Hox specificity by promoting the formation of different Hox-PBC heterodimers in vivo. Thus, the choice of Hox partner, and therefore Hox target genes, depends on subtle differences between Hox-PBC binding sites.

A Single Pitx1 Binding Site Is Essential for Activity of the LHβ Promoter in Transgenic Mice

2001 ◽  
Vol 15 (5) ◽  
pp. 734-746 ◽  
Author(s):  
Christine C. Quirk ◽  
Kristen L. Lozada ◽  
Ruth A. Keri ◽  
John H. Nilson
Keyword(s):  
Transgenic Mice ◽  
Binding Site ◽  
Cell Lines ◽  
Regulatory Element ◽  
Regulatory Elements ◽  
Vital Role ◽  
Proximal Region ◽  
Expression Vectors ◽  
Homeodomain Protein

Abstract Reproduction depends on regulated expression of the LHβ gene. Tandem copies of regulatory elements that bind early growth response protein 1 (Egr-1) and steroidogenic factor 1 (SF-1) are located in the proximal region of the LHβ promoter and make essential contributions to its activity as well as mediate responsiveness to GnRH. Located between these tandem elements is a single site capable of binding the homeodomain protein Pitx1. From studies that employ overexpression paradigms performed in heterologous cell lines, it appears that Egr-1, SF-1, and Pitx1 interact cooperatively through a mechanism that does not require the binding of Pitx1 to its site. Since the physiological ramifications of these overexpression studies remain unclear, we reassessed the requirement for a Pitx1 element in the promoter of the LHβ gene using homologous cell lines and transgenic mice, both of which obviate the need for overexpression of transcription factors. Our analysis indicated a striking requirement for the Pitx1 regulatory element. When assayed by transient transfection using a gonadotrope-derived cell line (LβT2), an LHβ promoter construct harboring a mutant Pitx1 element displayed attenuated transcriptional activity but retained responsiveness to GnRH. In contrast, analysis of wild-type and mutant expression vectors in transgenic mice indicated that LHβ promoter activity is completely dependent on the presence of a functional Pitx1 binding site. Indeed, the dependence on an intact Pitx1 binding site in transgenic mice is so strict that responsiveness to GnRH is also lost, suggesting that the mutant promoter is inactive. Collectively, our data reinforce the concept that activity of the LHβ promoter is determined, in part, through highly cooperative interactions between SF-1, Egr-1, and Pitx1. While Egr-1 can be regarded as a key downstream effector of GnRH, and Pitx1 as a critical partner that activates SF-1, our data firmly establish that the Pitx1 element plays a vital role in permitting these functions to occur in vivo.

scholarly journals STAT5 and Oct-1 Form a Stable Complex That Modulates Cyclin D1 Expression

2003 ◽  
Vol 23 (24) ◽  
pp. 8934-8945 ◽  
Author(s):  
Sophie Magné ◽  
Sandrine Caron ◽  
Martine Charon ◽  
Marie-Christine Rouyez ◽  
Isabelle Dusanter-Fourt
Keyword(s):  
Cyclin D1 ◽  
Target Genes ◽  
Promoter Sequence ◽  
Stable Complex ◽  
Homeodomain Protein ◽  
Carboxy Terminal Region ◽  
Carboxy Terminal ◽  
Cell Cycle Regulator Cyclin

ABSTRACT Signal transducer and activator of transcription 5 (STAT5) is activated by numerous cytokines that control blood cell development. STAT5 was also shown to actively participate in leukemogenesis. Among the target genes involved in cell growth, STAT5 had been shown to activate cyclin D1 gene expression. We now show that thrombopoietin-dependent activation of the cyclin D1 promoter depends on the integrity of a new bipartite proximal element that specifically binds STAT5A and -B transcription factors. We demonstrate that the stable recruitment of STAT5 to this element in vitro requires the integrity of an adjacent octamer element that constitutively binds the ubiquitous POU homeodomain protein Oct-1. We observe that cytokine-activated STAT5 and Oct-1 form a unique complex with the cyclin D1 promoter sequence. We find that STAT5 interacts with Oct-1 in vivo, following activation by different cytokines in various cellular contexts. This interaction involves a small motif in the carboxy-terminal region of STAT5 which, remarkably, is similar to an Oct-1 POU-interacting motif present in two well-known partners of Oct-1, namely, OBF-1/Bob and SNAP190. Our data offer new insights into the transcriptional regulation of the key cell cycle regulator cyclin D1 and emphasize the active roles of both STAT5 and Oct-1 in this process.

Transducing positional information to the Hox genes: critical interaction of cdx gene products with position-sensitive regulatory elements

Development ◽  
1998 ◽  
Vol 125 (22) ◽  
pp. 4349-4358 ◽  
Author(s):  
J. Charite ◽  
W. de Graaff ◽  
D. Consten ◽  
M.J. Reijnen ◽  
J. Korving ◽  
...  
Keyword(s):  
Binding Sites ◽  
Hox Genes ◽  
Regulatory Element ◽  
Ectopic Expression ◽  
Positional Information ◽  
Regulatory Elements ◽  
Gene Products ◽  
Anteroposterior Patterning

Studies of pattern formation in the vertebrate central nervous system indicate that anteroposterior positional information is generated in the embryo by signalling gradients of an as yet unknown nature. We searched for transcription factors that transduce this information to the Hox genes. Based on the assumption that the activity levels of such factors might vary with position along the anteroposterior axis, we devised an in vivo assay to detect responsiveness of cis-acting sequences to such differentially active factors. We used this assay to analyze a Hoxb8 regulatory element, and detected the most pronounced response in a short stretch of DNA containing a cluster of potential CDX binding sites. We show that differentially expressed DNA binding proteins are present in gastrulating embryos that bind to these sites in vitro, that cdx gene products are among these, and that binding site mutations that abolish binding of these proteins completely destroy the ability of the regulatory element to drive regionally restricted expression in the embryo. Finally, we show that ectopic expression of cdx gene products anteriorizes expression of reporter transgenes driven by this regulatory element, as well as that of the endogenous Hoxb8 gene, in a manner that is consistent with them being essential transducers of positional information. These data suggest that, in contrast to Drosophila Caudal, vertebrate cdx gene products transduce positional information directly to the Hox genes, acting through CDX binding sites in their enhancers. This may represent the ancestral mode of action of caudal homologues, which are involved in anteroposterior patterning in organisms with widely divergent body plans and modes of development.

Regulation of Hox target genes by a DNA bound Homothorax/Hox/Extradenticle complex

Development ◽  
1999 ◽  
Vol 126 (22) ◽  
pp. 5137-5148 ◽  
Author(s):  
H.D. Ryoo ◽  
T. Marty ◽  
F. Casares ◽  
M. Affolter ◽  
R.S. Mann
Keyword(s):  
Nuclear Localization ◽  
Target Genes ◽  
Dominant Negative ◽  
Homeodomain Protein ◽  
Hox Proteins ◽  
Trimeric Complex ◽  
Binding Residue ◽  
Dominant Negative Form

To regulate their target genes, the Hox proteins of Drosophila often bind to DNA as heterodimers with the homeodomain protein Extradenticle (EXD). For EXD to bind DNA, it must be in the nucleus, and its nuclear localization requires a third homeodomain protein, Homothorax (HTH). Here we show that a conserved N-terminal domain of HTH directly binds to EXD in vitro, and is sufficient to induce the nuclear localization of EXD in vivo. However, mutating a key DNA binding residue in the HTH homeodomain abolishes many of its in vivo functions. HTH binds to DNA as part of a HTH/Hox/EXD trimeric complex, and we show that this complex is essential for the activation of a natural Hox target enhancer. Using a dominant negative form of HTH we provide evidence that similar complexes are important for several Hox- and exd-mediated functions in vivo. These data suggest that Hox proteins often function as part of a multiprotein complex, composed of HTH, Hox, and EXD proteins, bound to DNA.

scholarly journals A New Role for Sterol Regulatory Element Binding Protein 1 Transcription Factors in the Regulation of Muscle Mass and Muscle Cell Differentiation

2009 ◽  
Vol 30 (5) ◽  
pp. 1182-1198 ◽  
Author(s):  
Virginie Lecomte ◽  
Emmanuelle Meugnier ◽  
Vanessa Euthine ◽  
Christine Durand ◽  
Damien Freyssenet ◽  
...  
Keyword(s):  
Transcription Factors ◽  
Muscle Mass ◽  
Target Genes ◽  
Binding Protein ◽  
Regulatory Element ◽  
Element Binding Protein ◽  
Sterol Regulatory Element ◽  
Myogenic Program

ABSTRACT The role of the transcription factors sterol regulatory element binding protein 1a (SREBP-1a) and SREBP-1c in the regulation of cholesterol and fatty acid metabolism has been well studied; however, little is known about their specific function in muscle. In the present study, analysis of recent microarray data from muscle cells overexpressing SREBP1 suggested that they may play a role in the regulation of myogenesis. We then demonstrated that SREBP-1a and -1c inhibit myoblast-to-myotube differentiation and also induce in vivo and in vitro muscle atrophy. Furthermore, we have identified the transcriptional repressors BHLHB2 and BHLHB3 as mediators of these effects of SREBP-1a and -1c in muscle. Both repressors are SREBP-1 target genes, and they affect the expression of numerous genes involved in the myogenic program. Our findings identify a new role for SREBP-1 transcription factors in muscle, thus linking the control of muscle mass to metabolic pathways.

scholarly journals Both Pbx1 and E2A-Pbx1 bind the DNA motif ATCAATCAA cooperatively with the products of multiple murine Hox genes, some of which are themselves oncogenes.

1995 ◽  
Vol 15 (7) ◽  
pp. 3786-3795 ◽  
Author(s):  
Q Lu ◽  
P S Knoepfler ◽  
J Scheele ◽  
D D Wright ◽  
M P Kamps
Keyword(s):  
Dna Binding ◽  
Target Genes ◽  
Hox Genes ◽  
Physical Interaction ◽  
Homeodomain Protein ◽  
Structural Development ◽  
Cell Leukemia ◽  
Homeodomain Proteins ◽  
Hox Proteins

E2A-PBX1 is the oncogene produced at the t(1;19) chromosomal breakpoint of pediatric pre-B-cell leukemia. Expression of E2A-Pbx1 induces fibroblast transformation and myeloid and T-cell leukemia in mice and arrests differentiation of granulocyte macrophage colony-stimulating factor-dependent myeloblasts in cultured marrow. Recently, the Drosophila melanogaster protein Exd, which is highly related to Pbx1, was shown to bind DNA cooperatively with the Drosophila homeodomain proteins Ubx and Abd-A. Here, we demonstrate that the normal Pbx1 homeodomain protein, as well as its oncogenic derivative, E2A-Pbx1, binds the DNA sequence ATCAATCAA cooperatively with the murine Hox-A5, Hox-B7, Hox-B8, and Hox-C8 homeodomain proteins, which are themselves known oncoproteins, as well as with the Hox-D4 homeodomain protein. Cooperative binding to ATCAATCAA required the homeodomain-dependent DNA-binding activities of both Pbx1 and the Hox partner. In cotransfection assays, Hox-B8 suppressed transactivation by E2A-Pbx1. These results suggest that (i) Pbx1 may participate in the normal regulation of Hox target gene transcription in vivo and therein contribute to aspects of anterior-posterior patterning and structural development in vertebrates, (ii) that E2A-Pbx1 could abrogate normal differentiation by altering the transcriptional regulation of Hox target genes in conjunction with Hox proteins, and (iii) that the oncogenic mechanism of certain Hox proteins may require their physical interaction with Pbx1 as a cooperating, DNA-binding partner.

scholarly journals Therapeutic Targeting of Nonalcoholic Fatty Liver Disease by Downregulating SREBP-1C Expression via AMPK-KLF10 Axis

2021 ◽  
Vol 8 ◽  
Author(s):  
Yu-Chi Chen ◽  
Rong-Jane Chen ◽  
Szu-Yuan Peng ◽  
Winston C. Y. Yu ◽  
Vincent Hung-Shu Chang
Keyword(s):  
Liver Disease ◽  
Fatty Liver ◽  
Nonalcoholic Fatty Liver Disease ◽  
Fatty Liver Disease ◽  
Target Genes ◽  
Regulatory Element ◽  
Wild Type ◽  
Nonalcoholic Fatty Liver

Krüppel-like factor 10 (KLF10) is a phospho-regulated transcriptional factor involved in many biological processes including lipogenesis; however, the transcriptional regulation on lipogenesis by KLF10 remains largely unclear. Lipogenesis is important in the development of nonalcoholic fatty liver disease (NAFLD) which was known regulated mainly by AMP-activated protein kinase (AMPK) and sterol regulatory element-binding protein (SREBP-1C). Interesting, our previous study using phosphorylated site prediction suggested a regulation of AMPK on KLF10. Therefore, we aimed to study the protein–protein interactions of AMPK on the regulation of KLF10, and to delineate the mechanisms of phosphorylated KLF10 in the regulation of NAFLD through SREBP-1C. We performed in vitro and in vivo assays that identified AMPK phosphorylates KLF10 at Thr189 and subsequently modulates the steady state level of KLF10. Meanwhile, a chromatin immunoprecipitation–chip assay revealed the novel target genes and signaling cascades of corresponding to phosphorylated KLF10. SREBP-1C was identified as a target gene suppressed by phosphorylated KLF10 through promoter binding. We further performed high-fat-diet-induced NAFLD models using hepatic-specific KLF10 knockout mice and wild-type mice and revealed that KLF10 knockout markedly led to more severe NAFLD than that in wild-type mice. Taken together, our findings revealed for the first time that AMPK activates and stabilizes the KLF10 protein via phosphorylation at Thr189, thereby repressing the expression of SREBP-1C and subsequent lipogenesis pathways along with metabolic disorders. We suggested that the targeted manipulation of liver metabolism, particularly through increased KLF10 expression, is a potential alternative solution for treating NAFLD.

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