Specific DNA Binding by c-Myb: Evidence for a Double Helix-Turn-Helix-Related Motif

Gabrielsen, Odd S.; Sentenac, André; Fromageot, P.

doi:10.1126/science.1887237

Cited by 140 publications

(123 citation statements)

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“…As for R3, a HTH motif with an unconventional turn was predicted for the R2 repeat [3,6): the second (recognition) helix of the HTH motif corresponds closely to the region of R2 found in the present work to have a flexible structure. Nevertheless, several lines of evidence support the idea of a direct role of the C-terminal part of R2 in DNA interaction.…”

Section: Secondary-structure Identificationmentioning

confidence: 94%

“…The c-Myb R2R3 domain was expressed in Escherichia coli as previously described [6] and the labeled proteins (uniformly IsN or doubly ''N, I3C labeled; selectively 15N labeled Lys, Leu and His residues) were obtained by growing the recombinant strain in different labeled media as summarized in Table 1. R2R3 expression was induced by addition of 0.4 mM isopropyl-thio-B-D-galactoside at A,,,, of 0.9 and 2 h after induction, the cells had accumulated the expected 12-kDa polypeptide.…”

Section: Preparation Of the Samplementioning

confidence: 99%

“…Deletion analysis has shown that only the last two repeats R2 and R3 are necessary for specific DNA binding [4, 51, while studies of the isolated R2R3 protein have demonstrated that these two domains are also sufficient to reconstitute full sequence-specific DNA binding [6]. The direct role of R2 in DNA interaction is also implied by the observation that while both R2 and R3 repeats are required for full sequence-specific DNA-binding activity, the isolated R3 domain is not capable of specific DNA binding (Gabrielsen, 0.…”

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confidence: 99%

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Secondary structure of the DNA‐binding domain of the c‐Myb oncoprotein in solution

Jamin

Gabrielsen

Gilles

et al. 1993

European Journal of Biochemistry

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The DNA-binding domain of the c-Myb oncoprotein contains two repeats, R2 and R3, both of which have been proposed to be related to the helix-turn-helix(HTH) motif. As a first step towards determination of the three-dimensional structure of this domain and of the mode of interaction with the DNA, we have undertaken multidimensional heteronuclear NMR studies using uniformly "N-labeled and 13C, 15N double-labeled R2R3 and, a selectively I5N-enriched sample on all lysine, histidine and leucine residues of R2R3. We present almost complete assignments of the backbone 'H, "N and 'TC" atoms and determine the secondary structure of R2R3 in solution.The R3 repeat is composed of three helices (residues 62-75, 78-85 and 91-100) while for the R2 repeat only two helices are found (residues 10-23 and 28-34). The remaining C-terminal part of the R2 repeat, predicted to be helical and part of the HTH motif, undergoes intermediate conformational exchange processes. Stabilization of this segment might occur upon binding to DNA.The c-myb proto-oncogene is the cellular homologue of the retroviral oncogene transduced by avian myeloblastosis virus and avian leukemia virus E26. It is also the target for retroviral insertion in certain murine myeloid leukemias and avian B-cell lymphomas [I, 21. The gene is preferentially expressed in immature cells of the lymphoid, erythroid and myeloid lineages; its level of expression is linked to the differentiation state of the hematopoietic cells. The myb oncogene has been implicated in the development of tumors in chicken, mice and humans and apparently transforms cells by freezing them in an immature proliferative state. The myb proto-oncogene encodes a transcription factor that seems to regulate genes important for development and growth of hematopoietic cells. Its DNA-binding domain is located near the N-terminus and is composed of three imperfect 51 -52-residue repeats (designated RI, R2 and R3). The most striking feature of the repeats is the presence of the three tryptophans repeat regularly spaced by 18 or 19 amino acids. This tryptophan motif is present in diffeAen.L transcriptional activa- Abbreviation. DANTE, delays alternating with nutation for tailored excitation ; CARP, globally optimized alternating-phase rectangular pulse; HMQC, heteronuclear multiple quantum correlation; HNCA, proton to amide nitrogen to a-carbon correlation ; HN(CO)CA, proton to amide nitrogen (via carbonyl) to a-carbon correlation ; HSQC, heteronuclear single quantum correlation; INEPT, insensitive nuclei enhanced by polarization transfer; NOESY, NOE spectroscopy; HTH, helix-turn-helix; TOCSY, total correlation spectroscopy.tors found in a wide spectrum of eukaryotes, including vertebrates, insects, yeast and higher plants and has been shown to be indispensable for maintaining the DNA-binding activity [l-31. Deletion analysis has shown that only the last two repeats R2 and R3 are necessary for specific DNA binding [4, 51, while studies of the isolated R2R3 protein have demonstrated that these two domains are also...

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Section: Secondary-structure Identificationmentioning

confidence: 94%

Section: Preparation Of the Samplementioning

confidence: 99%

mentioning

confidence: 99%

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Secondary structure of the DNA‐binding domain of the c‐Myb oncoprotein in solution

Jamin

Gabrielsen

Gilles

et al. 1993

European Journal of Biochemistry

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“…Information about the plasmids and antibodies used in this work is outlined in Supplementary Information Protein expression and GST pull-down assay GST and GST-SUMO fusion proteins were expressed in Escherichia coli as previously described (Gabrielsen et al, 1991). GST pull-down was performed with both COS-1 expressed and in vitro translated proteins as described in Supplementary Information.…”

Section: Plasmids and Antibodiesmentioning

confidence: 99%

A functional SUMO-interacting motif in the transactivation domain of c-Myb regulates its myeloid transforming ability

et al. 2010

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“…The minimal DBD of human c-Myb, R 2 R 3 (residues 89-104), designated c-Myb-DBD, was expressed in Escherichia coli and purified as described in [34].…”

Section: Expression Of C-myb Proteinsmentioning

confidence: 99%

The human neuroendocrine thyrotropin-releasing hormone receptor promoter is activated by the haematopoietic transcription factor c-Myb

Matre

Høvring

Fjeldheim

et al. 2003

Biochemical Journal

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Thyrotropin-releasing hormone (TRH) receptor (TRHR) is a G-protein-coupled receptor playing a crucial role in the anterior pituitary where it controls the synthesis and secretion of thyroidstimulating hormone and prolactin. Its widespread presence not only in the central nervous system, but also in peripheral tissues, including thymus, indicates other important, but unknown, functions. One hypothesis is that the neuropeptide TRH could play a role in the immune system. We report here that the human TRHR promoter contains 11 putative response elements for the haematopoietic transcription factor c-Myb and is highly Mybresponsive in transfection assays. Analysis of Myb binding to putative response elements revealed one preferred binding site in intron 1 of the receptor gene. Transfection studies of promoter deletions confirmed that this high-affinity element is necessary for efficient Myb-dependent transactivation of reporter plasmids in CV-1 cells. The Myb-dependent activation of the TRHR promoter was strongly suppressed by expression of a dominant negative Myb-Engrailed fusion. In line with these observations, reverse transcriptase PCR analysis of rat tissues showed that the TRHR gene is expressed both in thymocytes and bone marrow. Furthermore, specific, high-affinity TRH agonist binding to cell-surface receptors was demonstrated in thymocytes and a haematopoietic cell line. Our findings imply a novel functional link between the neuroendocrine and the immune systems at the level of promoter regulation.

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Specific DNA Binding by c-Myb: Evidence for a Double Helix-Turn-Helix-Related Motif

Cited by 140 publications

References 28 publications

Secondary structure of the DNA‐binding domain of the c‐Myb oncoprotein in solution

Secondary structure of the DNA‐binding domain of the c‐Myb oncoprotein in solution

A functional SUMO-interacting motif in the transactivation domain of c-Myb regulates its myeloid transforming ability

The human neuroendocrine thyrotropin-releasing hormone receptor promoter is activated by the haematopoietic transcription factor c-Myb

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