The cell-specific activity of the estrogen receptor α may be fine-tuned by phosphorylation-induced structural gymnastics

The estrogen receptor α (ERα) regulates the transcription of target genes by recruiting coregulator proteins through several domains including the two activation functions AF1 and AF2. The contribution of the N-terminally located AF1 activity is particularly important in differentiated cells, and for ERα to integrate inputs from other signaling pathways. However, how the phosphorylation of key residues influences AF1 activity has long remained mysterious, in part because the naturally disordered AF1 domain has resisted a structural characterization. The recent discovery of two coregulators that are specific for a phosphorylated form of AF1 suggests that phosphorylation, possibly in conjunction with the subsequent binding of these coregulators, may enforce a stable structure. The binding of the "pioneer" coregulators might facilitate the subsequent recruitment of yet other coregulators. Different AF1 folds may be enabled by the combinatorial action of posttranslational modifications and coregulator binding thereby fine-tuning ERα activities in a cell- and promoter-specific fashion

Biomarkers of browning of white adipose tissue and their regulation during exercise- and diet-induced weight loss

Background: A hypothesis exists whereby an exercise- or dietary-induced negative energy balance reduces human subcutaneous white adipose tissue (scWAT) mass through the formation of brown-like adipocyte (brite) cells. However, the validity of biomarkers of brite formation has not been robustly evaluated in humans, and clinical data that link brite formation and weight loss are sparse. Objectives: We used rosiglitazone and primary adipocytes to stringently evaluate a set of biomarkers for brite formation and determined whether the expression of biomarker genes in scWAT could explain the change in body composition in response to exercise training combined with calorie restriction in obese and overweight women (n = 79). Design: Gene expression was derived from exon DNA microarrays and preadipocytes from obesity-resistant and -sensitive mice treated with rosiglitazone to generate candidate brite biomarkers from a microarray. These biomarkers were evaluated against data derived from scWAT RNA from obese and overweight women before and after supervised exercise 5 d/wk for 16 wk combined with modest calorie restriction (∼0.84 MJ/d). Results: Forty percent of commonly used brite gene biomarkers exhibited an exon or strain-specific regulation. No biomarkers were positively related to weight loss in human scWAT. Greater weight loss was significantly associated with less uncoupling protein 1 expression (P = 0.006, R(2) = 0.09). In a follow-up global analysis, there were 161 genes that covaried with weight loss that were linked to greater CCAAT/enhancer binding protein α activity (z = 2.0, P = 6.6 × 10(−7)), liver X receptor α/β agonism (z = 2.1, P = 2.8 × 10(−7)), and inhibition of leptin-like signaling (z = −2.6, P = 3.9 × 10(−5)). Conclusion: We identify a subset of robust RNA biomarkers for brite formation and show that calorie-restriction–mediated weight loss in women dynamically remodels scWAT to take on a more-white rather than a more-brown adipocyte phenotype

Local origins impart conserved bone type-related differences in human osteoblast behaviour

Osteogenic behaviour of osteoblasts from trabecular, cortical and subchondral bone were examined to determine any bone type-selective differences in samples from both osteoarthritic (OA) and osteoporotic (OP) patients. Cell growth, differentiation; alkaline phosphatase (TNAP) mRNA and activity, Runt-related transcription factor-2 (RUNX2), SP7-transcription factor (SP7), bone sialoprotein-II (BSP-II), osteocalcin/bone gamma-carboxyglutamate (BGLAP), osteoprotegerin (OPG, TNFRSF11B), receptor activator of nuclear factor-κβ ligand (RANKL, TNFSF11) mRNA levels and proangiogenic vascular endothelial growth factor-A (VEGF-A) mRNA and protein release were assessed in osteoblasts from paired humeral head samples from age-matched, human OA/OP (n = 5/4) patients. Initial outgrowth and increase in cell number were significantly faster (p < 0.01) in subchondral and cortical than trabecular osteoblasts, in OA and OP, and this bone type-related differences were conserved despite consistently faster growth in OA. RUNX2/SP7 levels and TNAP mRNA and protein activity were, however, greater in trabecular than subchondral and cortical osteoblasts in OA and OP. BSP-II levels were significantly greater in trabecular and lowest in cortical osteoblasts in both OA and OP. In contrast, BGLAP levels showed divergent bone type-selective behaviour; highest in osteoblasts from subchondral origins in OA and trabecular origins in OP. We found virtually identical bone type-related differences, however, in TNFRSF11B:TNFSF11 in OA and OP, consistent with greater potential for paracrine effects on osteoclasts in trabecular osteoblasts. Subchondral osteoblasts (OA) exhibited highest VEGF-A mRNA levels and release. Our data indicate that human osteoblasts in trabecular, subchondral and cortical bone have inherent, programmed diversity, with specific bone type-related differences in growth, differentiation and pro-angiogenic potential in vitro

Time-series transcriptional profiling yields new perspectives on susceptibility to murine osteoarthritis.

Chronological age is a powerful epidemiologic risk factor for osteoarthritis (OA), a multifactorial disease that is characterized by articular cartilage (AC) degradation. It is unclear from a molecular perspective how aging interacts with OA to produce this risk to AC integrity. To address this key question, we used in vivo time-course analysis of OA development and murine interstrain variability in natural susceptibility to OA to examine changes in non-OA-prone CBA mice versus OA-prone STR/Ort mice, which develop disease that bears significant histologic resemblance to human OA. Through global transcriptome profiling, we attempted to discover the molecular signature linked with both OA vulnerability and progression. Affymetrix Mouse Gene 1.0 ST Array profiles were generated from AC samples derived from CBA and STR/Ort mice at 3 different ages, corresponding to the stages prior to, at, and late after the natural onset of OA in the STR/Ort mice. We found that the OA in STR/Ort mice exhibited a molecular phenotype resembling human OA, and we pinpointed a central role of NF-κB signaling and the emergence of an immune-related signature in OA cartilage over time. We discovered that, strikingly, young healthy AC has a highly expressed skeletal muscle gene expression program, which is switched off during maturation, but is intriguingly retained in AC during OA development in STR/Ort mice. This study is the first to show that AC chondrocytes share a high-abundance gene-expression program with skeletal muscle. We show that failure to switch this program off, as well as the restoration of this program, is associated with inappropriate expression of NF-κB signaling pathways, skeletal muscle-related genes, and induction and/or progression of OA. Copyright © 2012 by the American College of Rheumatology

Ontogeny and thermogenic role for sternal fat in female sheep

Brown adipose tissue acting through a unique uncoupling protein (UCP1) has a critical role in preventing hypothermia in new-born sheep but is then considered to rapidly disappear during postnatal life. The extent to which the anatomical location of fat influences postnatal development and thermogenic function, particularly following feeding, in adulthood, are not known and were both examined in our study. Changes in gene expression of functionally important pathways (i.e. thermogenesis, development, adipogenesis and metabolism) were compared between sternal and retroperitoneal fat depots together with a representative skeletal muscle over the first month of postnatal life, coincident with the loss of brown fat and accumulation of white fat. In adult sheep, implanted temperature probes were used to characterise the thermogenic response of fat and muscle to feeding and the effects of reduced or increased adiposity. UCP1 was more abundant within sternal than retroperitoneal fat and was only retained in the sternal depot of adults. Distinct differences in the abundance of gene pathway markers were apparent between tissues, with sternal fat exhibiting some similarities with muscle that were not apparent in the retroperitoneal depot. In adults, the post-prandial rise in temperature was greater and more prolonged in sternal than retroperitoneal fat and muscle, a difference that was maintained with altered adiposity. In conclusion, sternal adipose tissue retains UCP1 into adulthood when it shows a greater thermogenic response to feeding than muscle and retroperitoneal fat. Sternal fat may be more amenable to targeted interventions that promote thermogenesis in large mammals

Pax6 Represses Androgen Receptor-Mediated Transactivation by Inhibiting Recruitment of the Coactivator SPBP

The androgen receptor (AR) has a central role in development and maintenance of the male reproductive system and in the etiology of prostate cancer. The transcription factor Pax6 has recently been reported to act as a repressor of AR and to be hypermethylated in prostate cancer cells. SPBP is a transcriptional regulator that previously has been shown to enhance the activity of Pax6. In this study we have identified SPBP to act as a transcriptional coactivator of AR. We also show that Pax6 inhibits SPBP-mediated enhancement of AR activity on the AR target gene probasin promoter, a repression that was partly reversed by increased expression of SPBP. Enhanced expression of Pax6 reduced the amount of SPBP associated with the probasin promoter when assayed by ChIP in HeLa cells. We mapped the interaction between both AR and SPBP, and AR and Pax6 to the DNA-binding domains of the involved proteins. Further binding studies revealed that Pax6 and SPBP compete for binding to AR. These results suggest that Pax6 represses AR activity by displacing and/or inhibiting recruitment of coactivators to AR target promoters. Understanding the mechanism for inhibition of AR coactivators can give rise to molecular targeted drugs for treatment of prostate cancer

Gene-chip studies of adipogenesis-regulated microRNAs in mouse primary adipocytes and human obesity

<p>Abstract</p> <p>Background</p> <p>Adipose tissue abundance relies partly on the factors that regulate adipogenesis, i.e. proliferation and differentiation of adipocytes. While components of the transcriptional program that initiates adipogenesis is well-known, the importance of microRNAs in adipogenesis is less well studied. We thus set out to investigate whether miRNAs would be actively modulated during adipogenesis and obesity.</p> <p>Methods</p> <p>Several models exist to study adipogenesis <it>in vitro</it>, of which the cell line 3T3-L1 is the most well known, albeit not the most physiologically appropriate. Thus, as an alternative, we produced EXIQON microarray of brown and white <it>primary </it>murine adipocytes (prior to and following differentiation) to yield global profiles of miRNAs.</p> <p>Results</p> <p>We found 65 miRNAs regulated during <it>in vitro </it>adipogenesis in primary adipocytes. We evaluated the similarity of our responses to those found in non-primary cell models, through literature data-mining. When comparing primary adipocyte profiles, with those of cell lines reported in the literature, we found a high degree of difference in 'adipogenesis' regulated miRNAs suggesting that the model systems may not be accurately representing adipogenesis. The expression of 10 adipogenesis-regulated miRNAs were studied using real-time qPCR and then we selected 5 miRNAs, that showed robust expression, were profiled in subcutaneous adipose tissue obtained from 20 humans with a range of body mass indices (BMI, range = 21-48, and all samples have U133+2 Affymetrix profiles provided). Of the miRNAs tested, mir-21 was robustly expressed in human adipose tissue and positively correlated with BMI (R2 = 0.49, p < 0.001).</p> <p>Conclusion</p> <p>In conclusion, we provide a preliminary analysis of miRNAs associated with primary cell <it>in vitro </it>adipogenesis and demonstrate that the inflammation-associated miRNA, mir-21 is up-regulated in subcutaneous adipose tissue in human obesity. Further, we provide a novel transcriptomics database of EXIQON and Affymetrix adipocyte profiles to facilitate data mining.</p

Genomics of Signaling Crosstalk of Estrogen Receptor α in Breast Cancer Cells

BACKGROUND: The estrogen receptor alpha (ERalpha) is a ligand-regulated transcription factor. However, a wide variety of other extracellular signals can activate ERalpha in the absence of estrogen. The impact of these alternate modes of activation on gene expression profiles has not been characterized. METHODOLOGY/PRINCIPAL FINDINGS: We show that estrogen, growth factors and cAMP elicit surprisingly distinct ERalpha-dependent transcriptional responses in human MCF7 breast cancer cells. In response to growth factors and cAMP, ERalpha primarily activates and represses genes, respectively. The combined treatments with the anti-estrogen tamoxifen and cAMP or growth factors regulate yet other sets of genes. In many cases, tamoxifen is perverted to an agonist, potentially mimicking what is happening in certain tamoxifen-resistant breast tumors and emphasizing the importance of the cellular signaling environment. Using a computational analysis, we predicted that a Hox protein might be involved in mediating such combinatorial effects, and then confirmed it experimentally. Although both tamoxifen and cAMP block the proliferation of MCF7 cells, their combined application stimulates it, and this can be blocked with a dominant-negative Hox mutant. CONCLUSIONS/SIGNIFICANCE: The activating signal dictates both target gene selection and regulation by ERalpha, and this has consequences on global gene expression patterns that may be relevant to understanding the progression of ERalpha-dependent carcinomas

Experimental evidence reveals the UCP1 genotype changes the oxygen consumption attributed to non-shivering thermogenesis in humans

Humans have spread out all over the world adapting to many different cold environments. Recent worldwide genome analyses and animal experiments have reported dozens of genes associated with cold adaptation. The uncoupling protein 1 (UCP1) gene enhances thermogenesis reaction in a physiological process by blocking ATP (adenosine triphosphate) synthesis on a mitochondrial membrane in brown adipose tissues. To our knowledge, no previous studies have shown an association between variants of the UCP1 gene and physiological phenotypes concerning non-shivering thermogenesis (NST) under the condition of low temperature in humans. We showed that the degree of NST for healthy subjects in an artificial climate chamber is significantly different among UCP1 genotypes. Defining the haplotypes covering the UCP1 region (39.4?kb), we found that the frequency of the haplotype with the highest NST was significantly correlated with latitudes and ambient temperature. Thus, the data in this study provide the first evidence that the UCP1 genotype alters the efficiency of NST in humans, and likely supports the hypothesis that the UCP1 gene has been related to cold adaptation in human evolutionary history

Assessment of solar and wind energy resources in Serbia

Serbia aims to improve the composition of its energy sources and increase the contribution of renewable and sustainable energies to meet its growing electricity production demand. Results of many studies and numerous measurement campaigns suggest that Serbia has a considerable potential for the solar and wind energy utilization for electricity generation. The objective of this article is to present a brief assessment of these solar and wind resources. Meteorological data from a network of meteorological stations covering a period of 30 yr are used to obtain annual energy density distributions from these two climatic sources. These results, summarizing in an abbreviated form comprehensive research efforts culminating in the last decade, are presented in an archival publication for the first time. This assessment indicates that southern and southeastern Serbia are very good regions for the solar energy utilization. Southern Banat, mountain ranges of eastern and southeastern Serbia, and the lower Danube river valley are the most promising regions for the wind power plant constructions. Final objective of this article is to show that the problem of individual intermittency of wind and solar power potentials could be compensated in Serbia by their complementary character