Junctional diversity of H and L chains allows the coexpression of two mutually exclusive idiotope... more Junctional diversity of H and L chains allows the coexpression of two mutually exclusive idiotopes (IdI104 and IdI558).
ChemInform Abstract Die Phthalsäuredialdehyde (I) werden mit dem Nitroderivat (II) basisch zu den... more ChemInform Abstract Die Phthalsäuredialdehyde (I) werden mit dem Nitroderivat (II) basisch zu den 3-Nitro-2-naphthoesäuren (IIIa) kondensiert und diese wiederum mit NaBH4 zu den entsprechenden Alkoholen (IIIb) reduziert. Einige dieser Alkohole (IIIb) werden mit MnOz zu den Aldehyden (IIIc) oxidiert, oder mit SOCl2 in die 2-Chlormethylderivate (IIId)übergeführt. Ferner wird die Umsetzung des Dialdehyds (IV) mit der Nitroverbindung (II) zu den Anthracenen (V) und (VI) beschrieben (Aush-Angaben fehlen meist).
The Journal of Steroid Biochemistry and Molecular Biology, Nov 1, 1994
Steroid-free glucocorticoid receptors are generally considered to reside in the cytoplasm of cell... more Steroid-free glucocorticoid receptors are generally considered to reside in the cytoplasm of cells. After the binding of steroids, the receptors translocate into the nucleus in a manner that has been proposed to involve microtubules. However, some results with inhibitors of microtubule assembly argue to the contrary. In all of these studies, only the whole cell localization of receptors has been examined; the biological activity of these receptors has not been determined. We now report that steroid-induced gene expression is maintained in the absence of intact microtubules. This argues that microtubules are not required for either the nuclear translocation or biological activity of glucocorticoid receptors.
The involvement of a vicinally spaced dithiol group in steroid binding to the glucocorticoid rece... more The involvement of a vicinally spaced dithiol group in steroid binding to the glucocorticoid receptor has been deduced from experiments with the thiol-specific reagent methyl methanethiolsulfonate and the vicinal dithiol-specific reagent sodium arsenite. The vicinally spaced dithiol appears to reside in the 16-kDa trypsin fragment of the receptor, which is thought to contain 3 cysteines (Cys-640, -656, and -661 of the rat receptor) and binds hormone with an approximately 23-fold lower affinity than does the intact 98-kDa receptor. We now report that the steroid binding specificity of preparations of this 16-kDa fragment and the intact receptor are virtually identical. This finding supports our designation of the 16-kDa fragment as a steroid-binding core domain and validates our continued use of this tryptic fragment in studies of steroid binding. To identify the cysteines which comprise the vicinally spaced dithiol group, and to examine further the role of cysteines in steroid binding, a total of five point mutant receptors were prepared: cysteine-to-serine for each suspected cysteine, cysteine-to-glycine for Cys-656, and the C656,661S double mutant. Unexpectedly, each receptor with a single point mutation still bound steroid. Even the double mutant (C656,661S) bound steroid with wild type affinity. These results suggest that none of these cysteines are directly required either for steroid binding to the glucocorticoid receptor or for heat shock protein 90 association with the receptor. However, the presence of Cys-656 was obligatory for covalent labeling of the receptor by [3H]dexamethasone 21-mesylate. Studies with preparations of the 98 and 16 kDa forms of these mutant receptors revealed both that Cys-656 and -661 comprise the vicinally spaced dithiols reacting with arsenite and that any two of the three thiols could form an intramolecular disulfide after treatment with low concentrations of methyl methanethiolsulfonate. These data, in conjunction with those from experiments on the effects of steric bulk on various receptor functions, support a model for the ligand binding cavity of the receptor that involves all three thiols in a flexible cleft but where thiol-steroid interactions are not essential for binding.
Search by Subject Search using Medical Subject Headings (< b> MeSH</b>), a controlled... more Search by Subject Search using Medical Subject Headings (< b> MeSH</b>), a controlled vocabulary for indexing life sciences content.< br/> Note that some records do not have MeSH. These include Patents and the latest PubMed and PubMed Central records.
Ein 1:4: 1‐Gemisch von (I), (II) und (III) liefert bei 0°C in 95%igem Äthanol das Cyclisierungspr... more Ein 1:4: 1‐Gemisch von (I), (II) und (III) liefert bei 0°C in 95%igem Äthanol das Cyclisierungsprodukt (IV) in ausgezeichneter Ausbeute.
c-Myc, the product of a proto-oncogene Myc originally was identified as a transcriptional factor,... more c-Myc, the product of a proto-oncogene Myc originally was identified as a transcriptional factor, which interacts with MAX protein through the basic helix-loop-helix leucine zipper (bHLHZip) domain and binds to the DNA containing the E-box (CACGTG) to regulate global gene expression. Recently, we and others found that c-Myc is a genome-wide non-linear amplifier of all expressed genes. The purpose of this study is to confirm previous results that c-Myc targets active promoters and enhancers to amplify the gene expression and to define the contributions of different Myc boxes (Mbs) in the regulation of gene expression and to identify the protein partners of Mbs. A Gibson Assembly system was used to create the A to N mutations of conserved amino acids in different Mbs. The wild type (WT) and mutated Myc –Pd4-EGFP fusion genes were transiently co-transfected to U2OS cells with Tk-, GRE-TK-, PR-TK-Luc vectors and analyze the reporter activities with a Dual-Luciferase Reporter Assay System, respectively. Our preliminary data demonstrated that WT-c-Myc does amplify reporter gene activity non-linearly. However, constructs with mutatied MbI and MbII were handicapped for amplification. Interestingly, mutated MbIII-b dramatically enhanced reporter gene amplification especially at higher Myc concentrations. Using reporter constructs with and without E-boxes, we found that this element is not necessary to enable c-Myc 's amplification function. To understand c-Myc's function in vivo, we established several Tet-inducible WT and mutated c-Myc stable cell lines with lenti-virus transduced system. The gene expression profiles of these cell lines are being compared. The screening of the partners of different Mb domains is on the way. Citation Format: Zuqin Nie, Chunhua Guo, Stoney S. Simons, David L. Levens. In vitro and in vivo studies of c-Myc amplifiers function. [abstract]. In: Proceedings of the AACR Special Conference on Myc: From Biology to Therapy; Jan 7-10, 2015; La Jolla, CA. Philadelphia (PA): AACR; Mol Cancer Res 2015;13(10 Suppl):Abstract nr A42.
The role of the glucucorticoid receptor in the expression ofa~fig~ucocortjcoid action has been in... more The role of the glucucorticoid receptor in the expression ofa~fig~ucocortjcoid action has been investigated with a chemically-reactive derivative of three glucocorticoid steroids with differing biological potencies, i.e. the C-21 mesylates of cortisol. dexamethasone and deacylcortivazol. Dexamethasone 21-mesylate (Dex-Mes) was the most useful derivative due to its favorable balance of high receptor affinity and predominantly irreversible antiglucocorticoid activity. A number of criteria have been used to conclude that [3H]Dex-Mes covalently labels glucocorticoid receptors In the steroid-binding cavity. The available data indicate that covalent Dex-Mes-labeled receptors (mol. wt % 98,000) are responsible for the irreversible antiglucocort~~o~d activity while the partial agonist activity of Dex-Mes is due to non-covalent Dex-~~-bound receptors. Further support fcrr this hypothesis comes from the observations that dea~y~~ortiva~ol21mesyfate was a full glucocortitoid and did not afhnity label receptors (and marginahy labeled cytosol proteins) although it was capable of cova~ently-labeling bovine serum albumin. Severaf mechanisms for the expression of irreversible antiglucocorticoid activity by covalent Dex-Mes-labeled receptors were examined and can be eliminated. Covalent receptor-Dex-Mes complexes formed in whole HTC cells were found to have a decreased capacity for nuclear binding. This decreased nuclear-binding capacity could be responsible for the whole-cell irreversible antiglucocarticoid activity of Dex-Mes.
<p>The predictions are derived by examining the formulas for these parameters as shown in &... more <p>The predictions are derived by examining the formulas for these parameters as shown in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1004122#pcbi.1004122.s004" target="_blank">S2 Table</a>. F means Factor, GR means steroid-receptor complex, A means accelerator, D means any type of decelerator, C means competitive decelerator, U means uncompetetive decelerator, N means noncompetitive decelerator, L means linear decelerator, and P means partial decelerator. H is the concentration for the half-maximum of the dose-response parameter. This table represents sufficient conditions and is not complete. Note that two cofactors cannot be of the same type at the same step, GR will not repress gene expression if it is an A before the CLS, and the A<sub>min</sub> graphs allow GR to act anywhere as a D or as an A after the CLS.</p><p>Predicted mechanism based on dose-response parameter plots.</p
The presence of a thiol in the steroid binding cavity of glucocorticoid receptors has recently be... more The presence of a thiol in the steroid binding cavity of glucocorticoid receptors has recently been proved by our affinity labeling of Cys-656 in the steroid binding domain of rat receptors (Simons, s. s., Jr., Pum
Junctional diversity of H and L chains allows the coexpression of two mutually exclusive idiotope... more Junctional diversity of H and L chains allows the coexpression of two mutually exclusive idiotopes (IdI104 and IdI558).
ChemInform Abstract Die Phthalsäuredialdehyde (I) werden mit dem Nitroderivat (II) basisch zu den... more ChemInform Abstract Die Phthalsäuredialdehyde (I) werden mit dem Nitroderivat (II) basisch zu den 3-Nitro-2-naphthoesäuren (IIIa) kondensiert und diese wiederum mit NaBH4 zu den entsprechenden Alkoholen (IIIb) reduziert. Einige dieser Alkohole (IIIb) werden mit MnOz zu den Aldehyden (IIIc) oxidiert, oder mit SOCl2 in die 2-Chlormethylderivate (IIId)übergeführt. Ferner wird die Umsetzung des Dialdehyds (IV) mit der Nitroverbindung (II) zu den Anthracenen (V) und (VI) beschrieben (Aush-Angaben fehlen meist).
The Journal of Steroid Biochemistry and Molecular Biology, Nov 1, 1994
Steroid-free glucocorticoid receptors are generally considered to reside in the cytoplasm of cell... more Steroid-free glucocorticoid receptors are generally considered to reside in the cytoplasm of cells. After the binding of steroids, the receptors translocate into the nucleus in a manner that has been proposed to involve microtubules. However, some results with inhibitors of microtubule assembly argue to the contrary. In all of these studies, only the whole cell localization of receptors has been examined; the biological activity of these receptors has not been determined. We now report that steroid-induced gene expression is maintained in the absence of intact microtubules. This argues that microtubules are not required for either the nuclear translocation or biological activity of glucocorticoid receptors.
The involvement of a vicinally spaced dithiol group in steroid binding to the glucocorticoid rece... more The involvement of a vicinally spaced dithiol group in steroid binding to the glucocorticoid receptor has been deduced from experiments with the thiol-specific reagent methyl methanethiolsulfonate and the vicinal dithiol-specific reagent sodium arsenite. The vicinally spaced dithiol appears to reside in the 16-kDa trypsin fragment of the receptor, which is thought to contain 3 cysteines (Cys-640, -656, and -661 of the rat receptor) and binds hormone with an approximately 23-fold lower affinity than does the intact 98-kDa receptor. We now report that the steroid binding specificity of preparations of this 16-kDa fragment and the intact receptor are virtually identical. This finding supports our designation of the 16-kDa fragment as a steroid-binding core domain and validates our continued use of this tryptic fragment in studies of steroid binding. To identify the cysteines which comprise the vicinally spaced dithiol group, and to examine further the role of cysteines in steroid binding, a total of five point mutant receptors were prepared: cysteine-to-serine for each suspected cysteine, cysteine-to-glycine for Cys-656, and the C656,661S double mutant. Unexpectedly, each receptor with a single point mutation still bound steroid. Even the double mutant (C656,661S) bound steroid with wild type affinity. These results suggest that none of these cysteines are directly required either for steroid binding to the glucocorticoid receptor or for heat shock protein 90 association with the receptor. However, the presence of Cys-656 was obligatory for covalent labeling of the receptor by [3H]dexamethasone 21-mesylate. Studies with preparations of the 98 and 16 kDa forms of these mutant receptors revealed both that Cys-656 and -661 comprise the vicinally spaced dithiols reacting with arsenite and that any two of the three thiols could form an intramolecular disulfide after treatment with low concentrations of methyl methanethiolsulfonate. These data, in conjunction with those from experiments on the effects of steric bulk on various receptor functions, support a model for the ligand binding cavity of the receptor that involves all three thiols in a flexible cleft but where thiol-steroid interactions are not essential for binding.
Search by Subject Search using Medical Subject Headings (< b> MeSH</b>), a controlled... more Search by Subject Search using Medical Subject Headings (< b> MeSH</b>), a controlled vocabulary for indexing life sciences content.< br/> Note that some records do not have MeSH. These include Patents and the latest PubMed and PubMed Central records.
Ein 1:4: 1‐Gemisch von (I), (II) und (III) liefert bei 0°C in 95%igem Äthanol das Cyclisierungspr... more Ein 1:4: 1‐Gemisch von (I), (II) und (III) liefert bei 0°C in 95%igem Äthanol das Cyclisierungsprodukt (IV) in ausgezeichneter Ausbeute.
c-Myc, the product of a proto-oncogene Myc originally was identified as a transcriptional factor,... more c-Myc, the product of a proto-oncogene Myc originally was identified as a transcriptional factor, which interacts with MAX protein through the basic helix-loop-helix leucine zipper (bHLHZip) domain and binds to the DNA containing the E-box (CACGTG) to regulate global gene expression. Recently, we and others found that c-Myc is a genome-wide non-linear amplifier of all expressed genes. The purpose of this study is to confirm previous results that c-Myc targets active promoters and enhancers to amplify the gene expression and to define the contributions of different Myc boxes (Mbs) in the regulation of gene expression and to identify the protein partners of Mbs. A Gibson Assembly system was used to create the A to N mutations of conserved amino acids in different Mbs. The wild type (WT) and mutated Myc –Pd4-EGFP fusion genes were transiently co-transfected to U2OS cells with Tk-, GRE-TK-, PR-TK-Luc vectors and analyze the reporter activities with a Dual-Luciferase Reporter Assay System, respectively. Our preliminary data demonstrated that WT-c-Myc does amplify reporter gene activity non-linearly. However, constructs with mutatied MbI and MbII were handicapped for amplification. Interestingly, mutated MbIII-b dramatically enhanced reporter gene amplification especially at higher Myc concentrations. Using reporter constructs with and without E-boxes, we found that this element is not necessary to enable c-Myc 's amplification function. To understand c-Myc's function in vivo, we established several Tet-inducible WT and mutated c-Myc stable cell lines with lenti-virus transduced system. The gene expression profiles of these cell lines are being compared. The screening of the partners of different Mb domains is on the way. Citation Format: Zuqin Nie, Chunhua Guo, Stoney S. Simons, David L. Levens. In vitro and in vivo studies of c-Myc amplifiers function. [abstract]. In: Proceedings of the AACR Special Conference on Myc: From Biology to Therapy; Jan 7-10, 2015; La Jolla, CA. Philadelphia (PA): AACR; Mol Cancer Res 2015;13(10 Suppl):Abstract nr A42.
The role of the glucucorticoid receptor in the expression ofa~fig~ucocortjcoid action has been in... more The role of the glucucorticoid receptor in the expression ofa~fig~ucocortjcoid action has been investigated with a chemically-reactive derivative of three glucocorticoid steroids with differing biological potencies, i.e. the C-21 mesylates of cortisol. dexamethasone and deacylcortivazol. Dexamethasone 21-mesylate (Dex-Mes) was the most useful derivative due to its favorable balance of high receptor affinity and predominantly irreversible antiglucocorticoid activity. A number of criteria have been used to conclude that [3H]Dex-Mes covalently labels glucocorticoid receptors In the steroid-binding cavity. The available data indicate that covalent Dex-Mes-labeled receptors (mol. wt % 98,000) are responsible for the irreversible antiglucocort~~o~d activity while the partial agonist activity of Dex-Mes is due to non-covalent Dex-~~-bound receptors. Further support fcrr this hypothesis comes from the observations that dea~y~~ortiva~ol21mesyfate was a full glucocortitoid and did not afhnity label receptors (and marginahy labeled cytosol proteins) although it was capable of cova~ently-labeling bovine serum albumin. Severaf mechanisms for the expression of irreversible antiglucocorticoid activity by covalent Dex-Mes-labeled receptors were examined and can be eliminated. Covalent receptor-Dex-Mes complexes formed in whole HTC cells were found to have a decreased capacity for nuclear binding. This decreased nuclear-binding capacity could be responsible for the whole-cell irreversible antiglucocarticoid activity of Dex-Mes.
<p>The predictions are derived by examining the formulas for these parameters as shown in &... more <p>The predictions are derived by examining the formulas for these parameters as shown in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1004122#pcbi.1004122.s004" target="_blank">S2 Table</a>. F means Factor, GR means steroid-receptor complex, A means accelerator, D means any type of decelerator, C means competitive decelerator, U means uncompetetive decelerator, N means noncompetitive decelerator, L means linear decelerator, and P means partial decelerator. H is the concentration for the half-maximum of the dose-response parameter. This table represents sufficient conditions and is not complete. Note that two cofactors cannot be of the same type at the same step, GR will not repress gene expression if it is an A before the CLS, and the A<sub>min</sub> graphs allow GR to act anywhere as a D or as an A after the CLS.</p><p>Predicted mechanism based on dose-response parameter plots.</p
The presence of a thiol in the steroid binding cavity of glucocorticoid receptors has recently be... more The presence of a thiol in the steroid binding cavity of glucocorticoid receptors has recently been proved by our affinity labeling of Cys-656 in the steroid binding domain of rat receptors (Simons, s. s., Jr., Pum
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