http://precedings.nature.com/users/6232bb9a837bc95c3257f1bdcc6095f7/ Documents posted by Michael Baker Nature Publishing Group en Nature Precedings http://precedings.nature.com/images/header_logo.gif http://precedings.nature.com http://precedings.nature.com/documents/3374/version/1 Steroid hormones regulate many physiological processes in vertebrates, nematodes and arthropods through binding to nuclear receptors (NR), a metazoan-specific family of ligand-activated transcription factors. The main steps controlling the diversification of this family are now well understood. In contrast, the origin and evolution of steroid ligands remain mysterious although this is crucial for understanding the emergence of modern endocrine systems. Using a comparative genomic approach, we analyzed complete metazoan genomes to provide a comprehensive view of the evolution of major enzymatic players implicated in steroidogenesis at the whole Metazoan scale. Our analysis reveals that steroidogenesis has been independently elaborated in the three main Bilaterian lineages, and that steroidogenic cytochrome P450 enzymes descended from those that detoxify xenobiotics. http://precedings.nature.com/documents/3374/version/1 Fri, 26 Jun 2009 15:22:07 UTC Independent elaboration of steroid hormone signaling pathways in Metazoans hdl:10101/npre.2009.3374.1 2009-06-26 Michael E. Baker Nature Precedings 2009-06-26T15:22:07Z Manuscript Cancer Developmental Biology Bioinformatics Earth & Environment Evolutionary Biology http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/3316/version/1 The origins of signaling by vertebrate steroids are not fully understood. An important advance was the report that an estrogen-binding steroid receptor [SR] is present in amphioxus, a basal chordate with a similar body plan as vertebrates. To investigate the evolution of estrogen binding to steroid receptors, we constructed a 3D model of amphioxus SR complexed with estradiol. This 3D model indicates that although the SR is activated by estradiol, some interactions between estradiol and human ERα are not conserved in the SR, which can explain the low affinity of estradiol for the SR. These differences between the SR and ERα in the steroid-binding domain are sufficient to suggest that another steroid is the physiological regulator of the SR. The 3D model predicts that mutation of Glu-346 to Gln will increase the affinity of testosterone for amphioxus SR and elucidate the evolution of steroid binding to nuclear receptors. http://precedings.nature.com/documents/3316/version/1 Fri, 05 Jun 2009 20:00:44 UTC 3D model of amphioxus steroid receptor complexed with estradiol hdl:10101/npre.2009.3316.1 2009-06-05 Michael E. Baker Nature Precedings 2009-06-05T20:00:44Z Manuscript Cancer Developmental Biology Bioinformatics Evolutionary Biology http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/1542/version/2 Background. The origins of steroid-dependent regulation of the vertebrate estrogen receptor (ER) are poorly understood. Genes with statistically significant sequence similarity to vertebrate ERs have been found in lamprey, a basal vertebrate, and amphioxus, a basal chordate. Motif analysis of these sequences provides an opportunity to investigate early events in the evolution of the ER.Results. We used artificial intelligence-based software to construct twelve motifs specific to the estrogen-binding domain of ERα and ERβ in land vertebrates and teleosts. We mapped these ER-specific motifs onto the sequences of lamprey, amphioxus, invertebrate and selected vertebrate ERs and amphioxus, Ciona and human estrogen-related receptor (ERR). We find that lamprey ER contains eleven motifs common to ERs in the training set. In contrast, amphioxus ER contains only six motifs. Various invertebrate ERs contain either six or seven motifs. Unexpectedly, human and amphioxus ERRs contain nine of the twelve motifs, despite extensive sequence divergence during the descent of chordate ERs and ERRs from a common ancestor. We mapped the twelve motifs onto a multiple alignment of human, lamprey and amphioxus ERs, which depicted residues in human ERα that are known to bind estradiol. There is excellent conservation of these key residues in lamprey ER and poor conservation in amphioxus ER. Out of seventeen residues on human ERα that bind estradiol, sixteen and six are identical in lamprey and amphioxus ER, respectively. A phylogenetic tree of ERs and ERRs reveals a long branch for amphioxus ER, which is in agreement with the low sequence and motif similarity between amphioxus ER and other ERs.Conclusions. There are significant differences between B. floridae ER and vertebrate ERs in the steroid-binding domain as measured by motif analysis and percent of amino acids that are known to stabilize estradiol in human ERα. The absence in lamprey ER of motif 10, which maps to the C-terminus half of α-helix 9, may be important in recognition of novel estrogens, such as 15α-hydroxy-estradiol. http://precedings.nature.com/documents/1542/version/2 Mon, 14 Apr 2008 20:09:47 UTC Motif analysis of amphioxus, lamprey and invertebrate estrogen receptors and amphioxus and human estrogen-related receptors: Towards a better understanding of estrogen receptor evolution hdl:10101/npre.2008.1542.2 2009-03-04 Michael E. Baker Nature Precedings 2008-04-14T20:09:47Z Manuscript Cancer Developmental Biology Ecology Evolutionary Biology http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/2823/version/1 BACKGROUND: Endocrine disruptors include plasticizers, pesticides, detergents and pharmaceuticals. Turbot and other flatfish are used to characterize the presence of chemicals in the marine environment. Unfortunately, there are relatively few genes of turbot and other flatfish in GenBank, which limits the use of molecular tools such as microarrays and qRT-PCR to study disruption of endocrine responses in sentinel fish captured by regulatory agencies.OBJECTIVES: A multi-gene cross species microarray was fabricated as a diagnostic tool to screen the effects of environmental chemicals in fish, for which there is minimal genomic information. The array included genes that are involved in the actions of adrenal and sex steroids, thyroid hormone, and xenobiotic responses. This will provide a sensitive tool for screening for the presence of chemicals with adverse effects on endocrine responses in coastal fish species.METHODS: A custom multi-species microarray was used to study gene expression in wild hornyhead turbot, collected from polluted and clean coastal waters and in laboratory male zebrafish following exposure to estradiol and 4-nonylphenol. Gene-specific expression in turbot liver was measured by qRT-PCR and correlated to microarray data.RESULTS: Microarray and qRT-PCR analyses of livers from turbot collected near municipal wastewater discharge areas revealed altered gene expression profiles compared to those from reference areas. CONCLUSIONS: The agreement between the array data and qRT-PCR analyses validates this multi-species microarray. The microarray measurement of gene expression in zebrafish, which are phylogenetically distant from turbot, indicates that this multi-species microarray will be useful for measuring endocrine responses in other fish. http://precedings.nature.com/documents/2823/version/1 Mon, 02 Feb 2009 09:24:52 UTC Analysis of Endocrine Disruption in Southern California Coastal Fish using an Aquatic Multi-Species Microarray hdl:10101/npre.2009.2823.1 2009-02-02 Michael E. Baker Nature Precedings 2009-02-02T09:24:52Z Manuscript Developmental Biology Ecology Genetics & Genomics Earth & Environment http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/2768/version/1 Lamprey, a basal vertebrate, contains orthologs of the estrogen receptor (ER), progesterone receptor and corticoid receptor. A perplexing property of lamprey is that 15α-hydroxy-steroids are active steroids. For example, 15α-hydroxy-estradiol [15α-OH-E2] is the estrogen, instead of estradiol (E2). To investigate how 15α-OH-E2 binds lamprey ER, we constructed a 3D model of the lamprey ER with E2 and 15α-OH-E2. Our 3D model shows that Sδ on Met-409 can form a hydrogen bond with the 15α-hydroxyl on 15α-OH-E2. In human ERα, the corresponding residue Ile-424 has a van der Waals contact with 15α-OH-E2. BLAST analysis of GenBank indicates that among vertebrate ERs, only lamprey ER contains a methionine at this position. Thus, the contact between Sδ on Met-409 and 15α-OH-E2 is unique. Our 3D model of lamprey ER should prove useful in virtual screening of chemical libraries to identify compounds for controlling reproduction in sea lamprey, an environmental pest in the Great Lakes in the USA. http://precedings.nature.com/documents/2768/version/1 Fri, 09 Jan 2009 11:30:48 UTC 3D model of lamprey estrogen receptor with estradiol and 15α-hydroxy-estradiol hdl:10101/npre.2009.2768.1 2009-01-09 Michael E. Baker Nature Precedings 2009-01-09T11:30:48Z Manuscript Developmental Biology Ecology Earth & Environment http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/2430/version/1 Intracellular glucocorticoid reactivation is catalyzed by 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1), which functions predominantly as a reductase in cells expressing hexose-6-phosphate dehydrogenase (H6PDH). We recently showed that the ratios of cortisone to cortisol and 7-keto- to 7-hydroxy-neurosteroids are regulated by 11β-HSD1 and very much depend on co-expression with H6PDH, providing cosubstrate NADPH. Here, we investigated the impact of H6PDH on the modulation of 11β-HSD1-dependent inter-conversion of cortisone and cortisol by inhibitors and alternative substrates. Using HEK-293 cells expressing 11β-HSD1 or co-expressing 11β-HSD1 and H6PDH, we observed significant differences of 11β-HSD1 inhibition by natural and pharmaceutical compounds as well as endogenous hormone metabolites. Furthermore, we show potent and dose-dependent inhibition of 11β-HSD1 by 7-keto-DHEA in differentiated human THP-1 macrophages and in HEK-293 cells over-expressing 11β-HSD1 with or without H6PDH. In contrast, 7-ketocholesterol (7-KC) did not inhibit 11β-HSD1 in HEK-293 cells, even in the presence of H6PDH, but inhibited 11β-HSD1 reductase activity in differentiated THP-1 macrophages (IC50 = 8.1 +/- 0.9 μM). 7-keto-DHEA but not 7-KC inhibited 11β-HSD1 in HEK-293 cell lysates. In conclusion, cellular factors such as H6PDH can significantly modulate the effect of inhibitors and alternative 7-oxygenated substrates on intracellular glucocorticoid availability. http://precedings.nature.com/documents/2430/version/1 Wed, 22 Oct 2008 19:07:23 UTC Hexose-6-phosphate dehydrogenase modulates the effect of inhibitors and alternative substrates of 11β-hydroxysteroid dehydrogenase 1 hdl:10101/npre.2008.2430.1 2008-10-22 Michael E. Baker Nature Precedings 2008-10-22T19:07:23Z Manuscript Molecular Cell Biology Pharmacology Bioinformatics http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/2312/version/1 Background. Organotins are highly toxic and widely distributed environmental chemicals. Dibutyltin (DBT) is used as stabilizer in the production of polyvinyl chloride plastics, and it is also the major metabolite formed from tributyltin (TBT) in vivo. DBT is immunotoxic, however, the responsible targets remain to be defined. Due to the importance of glucocorticoids in immune-modulation, we investigated whether DBT could interfere with glucocorticoid receptor (GR) function.Methodology. We used HEK-293 cells transiently transfected with human GR as well as rat H4IIE hepatoma cells and native human macrophages and human THP-1 macrophages expressing endogenous receptor to study organotin effects on GR function. Docking of organotins was used to investigate the binding mechanism. Principal findings. We found that nanomolar concentrations of DBT, but not other organotins tested, inhibit ligand binding to GR and its transcriptional activity. Docking analysis indicated that DBT inhibits GR activation allosterically by inserting into a site close to the steroid-binding pocket, which disrupts a key interaction between the A-ring of the glucocorticoid and the GR. DBT inhibited glucocorticoid-induced expression of phosphoenolpyruvate carboxykinase (PEPCK) and tyrosine-aminotransferase (TAT) and abolished the glucocorticoid-mediated transrepression of TNF-α-induced NF-κB activity. Moreover, DBT abrogated the glucocorticoid-mediated suppression of interleukin-6 (IL-6) and TNF-α production in lipopolysaccharide (LPS)-stimulated native human macrophages and human THP-1 macrophages.Conclusions. DBT inhibits ligand binding to GR and subsequent activation of the receptor. By blocking GR activation, DBT may disturb metabolic functions and modulation of the immune system, providing an explanation for some of the toxic effects of this organotin. http://precedings.nature.com/documents/2312/version/1 Fri, 19 Sep 2008 18:30:41 UTC Dibutyltin Disrupts Glucocorticoid Receptor Function and Impairs Glucocorticoid-induced Suppression of Cytokine Production hdl:10101/npre.2008.2312.1 2008-09-19 Michael E. Baker Nature Precedings 2008-09-19T18:30:41Z Manuscript Immunology Pharmacology Bioinformatics http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/2170/version/1 Although, as their names imply, estrogen receptors [ERs] and estrogen-related receptors [ERRs] are related transcription factors, their evolutionary relationships to each other are not fully understood. To elucidate the origins and evolution of ERs and ERRs, we searched for their orthologs in the recently sequenced genome of Trichoplax, the simplest known animal, and in the genomes of three lophotrochozoans: Capitella, an annelid worm, Helobdella robusta, a leech, and Lottia gigantea, a snail. BLAST searches found an ERR in Trichoplax, but no ER. BLAST searches also found ERRs in all three lophotrochozoans and invertebrate-like ERs in Capitella and Lottia, but not in Helobdella. Unexpectedly we find that the Capitella ER sequence is closest to ERβ, unlike the other invertebrate ER sequences, which are closest to ERα. Our database searches and phylogenetic analysis indicate that invertebrate ERs evolved in a lophotrochozoan and steroid-binding ERs evolved in a deuterostome. http://precedings.nature.com/documents/2170/version/1 Wed, 13 Aug 2008 09:09:40 UTC Trichoplax, the simplest known animal, contains an estrogen-related receptor but no estrogen receptor: Implications for estrogen receptor evolution hdl:10101/npre.2008.2170.1 2008-08-13 Michael E. Baker Nature Precedings 2008-08-13T09:09:40Z Manuscript Developmental Biology Ecology Evolutionary Biology http://creativecommons.org/licenses/by/3.0/ http://precedings.nature.com/documents/1863/version/1 Although, as their names imply, vertebrate and invertebrate estrogen receptors [ERs] and estrogen-related receptors [ERRs] are related transcription factors, their evolutionary relationships to each other are not fully understood. We searched recently sequenced genome of Trichoplax, the simplest known animal, and genomes from three lophotrochozoans: Capitella, a worm, Helobdella robusta, a leech, and Lottia gigantea, a snail, to elucidate the origins and evolution of ERs and ERRs. BLAST found an ERR in Trichoplax, but no ER. BLAST searches of the lophotrochozaons found ERRs in all three and invertebrate ERs in Capitella and Lottia, but not in Helobdella. These database searches and a phylogenetic analyses indicate that invertebrate ERs arose in a protostome, and vertebrate ERs arose later in deuterostome. http://precedings.nature.com/documents/1863/version/1 Tue, 06 May 2008 22:32:29 UTC Trichoplax, the simplest known animal, contains an estrogen-related receptor: Implications for the evolution of vertebrate and invertebrate estrogen receptors hdl:10101/npre.2008.1863.1 2008-05-06 Michael E. Baker Nature Precedings 2008-05-06T22:32:29Z Manuscript Developmental Biology Ecology Evolutionary Biology http://creativecommons.org/licenses/by/3.0/