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J D Baxter

Publications and source records attributed to J D Baxter.

At least 73 records · Page 4Linked to original sources
PubMed1995

The nuclear hormone receptor gene superfamily.

The nuclear hormone receptor gene superfamily encodes structurally related proteins that regulate transcription of target genes. These macromolecules include receptors for steroid and thyroid hormones, vitamins, and other proteins for which no ligands have been found. These receptors have modular domains. The DNA-binding domain directs the receptors to bind specific DNA sequences as monomers, homodimers, or heterodimers. The ligand-binding domain responds to binding of the cognate hormone; this domain and the amino terminal domain interact with other transcription factors. Nuclear receptor-specific actions are derived from a combination of diverse elements, including availability of ligand, receptors, and nonreceptor factors; target-site structure; interactions with other proteins, such as the general transcription factors; and influences of other signaling pathways. These interactions result in ligand-regulated and ligand-independent effects on initiation of transcription of the target genes. Understanding the mechanisms of nuclear receptor action will enhance our knowledge of transcription and hormone influences on disease and facilitate the design of drugs with greater therapeutic value.

Endocrine System Diseases↗
PubMed1994

Paradoxical production of target protein using antisense RNA expression vectors.

We used antisense RNA in a protocol designed to reduce estrogen receptor (ER) content in human breast cancer cells and observed paradoxical increases in ER levels. ER protein activity was measured using a highly sensitive reporter gene assay that relies on the ability of functional ER to bind a consensus estrogen response element (ERE) and drive the production of chloramphenicol acetyl-transferase (CAT). Upon transient transfection of ER-positive cell lines with three different vectors containing the full-length ER cDNA cloned in an antisense orientation, we observed unexpected increases in ER-driven CAT activity. To further investigate this phenomenon, expression from the antisense ER vectors was studied in an ER-negative breast tumor cell line, MDA-MB-453. ER activity was observed in these ER-negative cells upon transient transfection with each of three antisense ER vectors, but not from control vectors. Expression of ER from antisense constructs was 30-100-times less efficient than ER expression from isogenic sense constructs. The paradoxical ER activity was consistent with expected ER behavior in that it exhibited characteristic binding to the natural ligand, 17 beta-estradiol (E2), and it was inhibited by the antiestrogens, 4-hydroxy-tamoxifen (OHT) and ICI 164384 (ICI). Control vectors containing a truncated antisense ER cDNA produced no ER activity. Although the mechanism for this ER expression has not been determined, it appears likely that it is due to transcription off the opposite strand of the antisense construct.(ABSTRACT TRUNCATED AT 250 WORDS)

Breast Neoplasms↗
PubMed1994

Preliminary crystallographic studies of the ligand-binding domain of the thyroid hormone receptor complexed with triiodothyronine.

A truncated, recombinant form of the thyroid hormone receptor, including the hormone binding domain, has been co-crystallized with the hormone T3. The crystals are monoclinic, most likely space group P2, with two molecules per asymmetric unit and cell dimensions a = 63.6 A, b = 80.8 A, c = 100.9 A and beta = 92.1 degrees. The crystals diffract to only medium resolution and decay rapidly in the X-ray beam using laboratory sources. By contrast, high resolution, high-quality data are obtained using synchrotron radiation in conjunction with cryocrystallography.

Binding Sites↗
PubMed1994

Phosphorylation selectively increases triiodothyronine receptor homodimer binding to DNA.

Thyroid hormone receptors (TRs) are ligand-regulated transcription factors that bind to thyroid hormone response elements (TREs) as monomers and homodimers, and as heterodimers with nuclear proteins such as TR auxiliary proteins and retinoid X receptors. Recently, bacterially expressed human TR beta-1 (hTR beta-1) was shown to be phosphorylated in vitro by HeLa cytosolic extract. However, little is known about the consequences of phosphorylation on the nature of TR complexes. Therefore, we studied the effect of phosphorylation on TR binding of TREs. Bacterially expressed hTR beta-1 was phosphorylated in vitro with ATP by HeLa cytosolic extract. The ratio of phosphoserine to phosphothreonine was approximately 5:1. We then analyzed phosphorylated hTR beta-1 binding to several TREs by electrophoretic mobility shift assay. Phosphorylated hTR beta-1 bound better as a homodimer to the TREs than hTR beta-1 incubated with preheated cytosolic extract. Alkaline phosphatase treatment of the phosphorylated hTR beta-1 eliminated the enhanced homodimer binding to DNA. In contrast, phosphorylation did not affect TR/TR auxiliary protein or TR/retinoid X receptor heterodimer binding to DNA. Triiodothyronine decreased both phosphorylated and unphosphorylated hTR beta-1 homodimer binding to several TREs, and the addition of okadaic acid did not alter this triiodothyronine effect. These results indicate that phosphorylation, in addition to ligand binding, modulates TR dimer binding to TREs. As such, it is possible that phosphorylation may also participate in TR-mediated regulation of transcription.

Blotting, Western↗
PubMed1994

AIDS hot lines and information agencies. The consistency of their information.

OBJECTIVES: To determine the consistency of responses given by acquired immunodeficiency syndrome (AIDS) hot lines and AIDS information agencies to a standard set of questions related to human immunodeficiency virus (HIV) infection and to investigate the training of workers in these telephone information agencies. DESIGN: A two-part questionnaire that addressed issues of HIV transmission and testing (part 1) and sexual behavior, diet, alcohol use, and partner notification (part 2). SETTING: Thirty-three AIDS telephone information agencies. PARTICIPANTS: Information specialists at these agencies were contacted on multiple occasions over a 1-year period. INTERVENTION: Initially, 48 calls were made to these agencies. Responses were recorded in the words of the information specialists, categorized, and assessed for consistency. Two additional surveys using the same questionnaire were also performed, examining the variability of responses within an individual agency and the variability of responses from six agencies, three with and three without formal training of their information specialists. RESULTS: The initial survey revealed inconsistencies and inaccuracies in responses from AIDS telephone information agencies particularly in the areas of HIV testing, risk factor assessment, safe-sex recommendations, and life-style changes for infected persons. The second survey revealed variability in responses from six information specialists within a single agency. The third survey revealed that, overall, agencies with and without formal training of workers were equally inconsistent in their responses. CONCLUSIONS: There are identifiable deficiencies in the quality and consistency of information given by some AIDS telephone information agencies. Training procedures of information specialists at these agencies are variable and may be inadequate.

Acquired Immunodeficiency Syndrome↗
PubMed1994

Accuracy and cost-effectiveness of a one-tube, three-color method for obtaining absolute CD4 counts and CD4:CD8 ratios.

An increasing number of assays to determine absolute CD4 counts are being performed as the acquired immunodeficiency syndrome epidemic continues. Simultaneously, there is increasing pressure to contain costs in the clinical laboratory. A rapid, one-tube, three-color method for obtaining CD4 counts has been evaluated for accuracy, compared with a more traditional two-color panel of markers. The data, compiled on 102 patients, including both pediatric and adult human immunodeficiency virus-infected patients, indicate a high degree of correlation between the one-tube method and a traditional panel. This approach should be considered for cost-effective and accurate CD4 determinations.

Adult↗
PubMed1994

Relapsing chickenpox in a young man with non-Hodgkin's lymphoma.

Reinfection of previously "immune" patients with varicella-zoster virus can result in recurrent chickenpox. However, relapses of chickenpox are not well recognized. We describe a young man with lymphoma who rapidly developed multiple, discrete relapses of chickenpox each time therapy with acyclovir was stopped. His cutaneous infection eventually became unremitting, despite continuous treatment with acyclovir. No evidence ever suggested visceral dissemination of varicella. The patient died suddenly; the cause of death was not determined.

Acyclovir↗
PubMed1994

Roles of 3,5,3'-triiodothyronine and deoxyribonucleic acid binding on thyroid hormone receptor complex formation.

Thyroid hormone receptors (TRs) bind to thyroid hormone response elements (TREs) in the promoter region of target genes as monomers, homodimers, and heterodimers with nuclear proteins such as retinoid-X receptors (RXRs). Recently, we observed that T3 decreased TR homodimer, but not TR/RXR heterodimer, binding to TREs, suggesting that the latter complexes may be involved in transcriptional activation of target genes. However, little is known about TR complexes that form in solution. Thus far, there have been only limited studies comparing TR complex formation in solution and on DNA as well as examining the effects of T3 and the putative ligand for RXRs, 9-cis retinoic acid (9-cis RA), on TR complex formation. In this paper, we used a coimmunoprecipitation assay with anti-TR beta 1 antibody and the electrophoretic mobility shift assay under similar buffer and incubation conditions to demonstrate that in the absence of T3, TR beta 1 is present as a monomer in solution and binds primarily as a homodimer to the chicken lysozyme TRE, F2. In the presence of T3, TR beta 1 cannot form a homodimer on F2, but, instead, exists as a liganded monomer in solution. Kinetic studies demonstrated that T3 markedly increased the dissociation rate of TR homodimer from F2. Using similar methods, we observed TR beta 1/RXR alpha heterodimer formation in solution and 10-fold greater formation on F2. Neither T3 nor 9-cis RA significantly affected TR beta 1/RXR alpha heterodimer formation. Taken together, these results suggest that both T3 and TRE binding are important determinants of the formation of specific TR complexes in solution and on DNA.

Base Sequence↗
PubMed1994

Heterodimerization and deoxyribonucleic acid-binding properties of a retinoid X receptor-related factor.

The extent thyroid hormone receptors (TRs) bind to AGGTCA-related motifs as monomers and/or homodimers, and as heterodimers with retinoid X receptors (RXRs) depends on the number, spacing, and orientation of these half-sites. Here we show that recombinant RXR alpha affects TR binding to DNA in diverse ways; it enhances recombinant TR beta 1 binding to four-nucleotide-spaced direct repeat and palindromes but not to inverted palindromes. We also used an endogenous factor termed RXR alpha-RF that cross-reacted with antibodies to RXR alpha and copurified and formed heterodimers on DNA with rat liver TRs (mostly TR beta 1 isoform), supporting the fact that endogenous TRs are commonly heterodimers. RXR alpha-RF formed, like recombinant RXR alpha, heterodimers on DNA with vitamin D and retinoic acid but not estrogen receptors. RXR alpha-RF differed from recombinant RXR alpha in that it provoked enhancement of TR beta 1 binding to DNA irrespective of half-site architecture, was resistant to heating to 50 C, and did not form heterodimers with recombinant TR alpha 2 on four-nucleotide-spaced direct repeat. The overall enhancement of TR-DNA recognition by endogenous RXR alpha-RF, not found in studies with recombinant RXR alpha, might exemplify properties acquired in vivo by endogenous RXRs; this could promote wider DNA recognition by TRs and expand the thyroid hormone transcriptional influence in the cell.

Animals↗
PubMed1993

Positive and negative modulation of Jun action by thyroid hormone receptor at a unique AP1 site.

We have characterized the putative AP1 site in the backbone of pUC plasmids and found unique regulatory effects. The site, which mapped to a 19-bp region around nucleotide 37, conferred transcriptional activation by Jun or Jun/Fos that was boosted up to fivefold by unliganded thyroid hormone receptor (TR). Thyroid hormone changed potentiation of the Jun response by TR into repression. Although the plasmid sequence is a near-perfect consensus AP1 site, the perfect consensus AP1 site from the human collagenase promoter did not show the same effects. Deletion of the ligand binding domain of the TR eliminated the ability of the receptor to boost Jun activity, and deletion, mutation, or changes in specificity of the DNA binding domain eliminated both its ability to potentiate Jun activity and repress with hormone. In vitro Jun/Fos complexes bound the operative plasmid fragment, and the presence of TR interfered very little with Jun/Fos binding activity. Protein interaction studies in the absence of DNA showed that TR bound Jun protein in solution either in the presence or in the absence of hormone. These observations suggest a mechanism for synergy and repression by TR through modulation of Jun activity: positive when TR is unliganded, and negative when hormone is bound. They also suggest that the presence of the plasmid element can confound studies of the regulation of linked promoters.

Animals↗
PubMed1993

Clinical use of BOBCAT: testing reliability and validity of computerized pure-tone audiometry with noise-exposed workers, children and the aged.

An investigation was conducted to determine the feasibility of implementing computerized audiometry in various clinical groups, using the Battery of Basic Computerized Audiometry Tests (BOBCAT). Reliability, validity and speed of execution were assessed as a function of hearing loss in a group of noise-exposed workers. Children and the aged were also included to represent potentially 'more-difficult-to-test' patients due to fluctuating attention, motivation and/or response behavior. Children were aged 7.5-12 years; seniors, 65-80 years. Reliability was assessed by calculating reliability coefficients between air conduction pure-tone thresholds (0.5, 1, 2, 3, 4 and 6 kHz) obtained under two test modalities namely, computerized audiometry and conventional testing performed by a small panel of trained examiners. Both procedures followed ANSI S3.21 1978 standards. Content validity was measured using measures of central tendency and correlations. Coefficients of reliability remained equally high across frequencies regardless of degree of hearing loss and group. As well, group means and correlations between conventional and computerized audiometry indicated that the two methods measured pure-tone hearing sensitivity with the same degree of accuracy; that is, within +/- 0.5 dB. Finally, speed of execution was found to be slower using BOBCAT, in particular with noise-exposed workers. This finding was interpreted as evidence that trained examiners have used shortcuts when they expect consistent and/or unbiased response behavior.

Acoustic Impedance Tests↗
PubMed1992

The effects of glucocorticoid therapy.

The therapeutic benefit of glucocorticoids appears to derive chiefly from their suppression of immunologic and inflammatory responses, but beyond these generalized phenomena, there are a number of tissue-specific effects. Increasing knowledge of these actions is shedding light on such issues as steroid resistance and steroid-induced adrenocortical insufficiency.

Adrenal Insufficiency↗
PubMed1992

Negative thyroid hormone control of human growth hormone gene expression is mediated by 3'-untranslated/3'-flanking DNA.

The intact human growth hormone (hGH) gene is negatively regulated by triiodothyronine (T3) treatment in transfected rat pituitary tumor cells. We now demonstrate that this responsiveness is mediated by a negative thyroid hormone response element (nTRE) localized to the proximal 3'-untranslated/3'-flanking region (3'-UT/FR). This region binds thyroid hormone receptors specifically and with high affinity. nTRE function was promoter-dependent, since it suppressed the activity of a positive TRE in the human chorionic somatomammotropin promoter, partially repressed activity of the herpes simplex virus TK promoter, but did not function with the human actin or Rous sarcoma virus promoters. T3 treatment did not alter transcript termination sites nor selectively affect the stability of transcripts containing the hGH 3'-UT/FR when transcription was blocked by actinomycin D treatment. The function of the nTRE depended on its location in the 3'-UT/FR; it was inactive when positioned down-stream of the simian virus 40 (SV40) 3'-UT/FR, and it acted as a positive TRE when placed upstream of the hGH promoter. These results demonstrate a novel localization of a TRE with unique properties which suggests expanded mechanisms by which thyroid hormone receptors can affect gene expression.

Animals↗
PubMed1992

Thyroid hormone responsiveness in human growth hormone-related genes. Possible correlation with receptor-induced DNA conformational changes.

Triiodothyronine (T3) induces the transcription of the human chorionic somatomammotropin (hCS) promoter transfected into rat pituitary (GC) cells, but does not stimulate the homologous human growth hormone (hGH) promoter. As demonstrated by forward and reverse mutagenesis, this differential T3 responsiveness is due to subtle structural differences in a T3 response element located between nucleotides -64 and -44 of the 5'-flanking DNA of the hGH and hCS promoters. Synthetic hCS(-70/-40) DNA binds thyroid hormone receptors with a 4-fold higher affinity than the corresponding hGH T3 response element, indicating that small differences in receptor binding properties are reflected by major differences in T3 responsiveness. Analysis of circular permutation fragments containing the native hGH and hCS or mutated hCS(-70/-40) sequences demonstrates that the thyroid hormone receptor induces DNA bending. The extent of bending shows a possible correlation with the function of these sequences, suggesting that the receptor-induced changes in DNA conformation may be required for thyroid hormone receptor action.

Animals↗
PubMed1992

Acute syphilitic meningitis in a man with seropositivity for human immunodeficiency virus infection and normal numbers of CD4 T lymphocytes.

Coinfection with the human immunodeficiency virus (HIV) and Treponema pallidum may predispose to accelerated neurosyphilis. The mechanism underlying this interaction is undefined, but usually presumed to result from HIV-induced suppression of cell-mediated immunity as reflected in the CD4 T-lymphocyte count. We report a patient infected with HIV who developed aggressive neurosyphilis despite a CD4 count of 1000/mm3. The CD4 cells constituted 17% of his total lymphocytes. Our case suggests that while severe neurosyphilis can occur in HIV-infected persons with normal absolute CD4 counts, the percentage of CD4 cells may be a better indicator of the risk of neurosyphilis. These observations are relevant to designing treatment strategies for patients coinfected with HIV and T pallidum based on measures of their immunocompetence.

CD4-Positive T-Lymphocytes↗
PubMed1992

The core promoter region of the tumor necrosis factor alpha gene confers phorbol ester responsiveness to upstream transcriptional activators.

Activators of protein kinase C, such as 12-O-tetradecanoylphorbol 13-acetate (TPA), are known to regulate the expression of many genes, including the tumor necrosis factor alpha (TNF) gene, by affecting the level or activity of upstream transcription factors. To investigate the mechanism whereby TPA activates the TNF promoter, a series of 5'-deletion mutants of the human TNF promoter linked to chloramphenicol acetyltransferase was transfected into U937 human promonocytic cells. TPA produced a 7- to 11-fold activation of all TNF promoters tested, even those promoters truncated to contain only the core promoter with no upstream enhancer elements. The proximal TNF promoter containing only 28 nucleotides upstream and 10 nucleotides downstream of the RNA start site confers TPA activation to a variety of unrelated upstream enhancer elements and transcription factors, including Sp1, CTF/NF1, cyclic AMP-response element, GAL-E1a, and GAL-VP16. The level of activation by TPA depends on the TATA box structure, since the TPA response is greater in promoters containing the sequence TATAAA than in those containing TATTAA or TATTTA. These findings suggest that the core promoter region is a target for gene regulation by second-messenger pathways.

Base Sequence↗