Bone Morphogenetic Protein 9 Is a Mechanistic Biomarker of Portopulmonary Hypertension.

Rationale: BMP9 (bone morphogenetic protein 9) is a circulating endothelial quiescence factor with protective effects in pulmonary arterial hypertension (PAH). Loss-of-function mutations in BMP9, its receptors, and downstream effectors have been reported in heritable PAH.

Objectives: To determine how an acquired deficiency of BMP9 signaling might contribute to PAH.

HD CAGnome: a search tool for huntingtin CAG repeat length-correlated genes.

Background: The length of the huntingtin (HTT) CAG repeat is strongly correlated with both age at onset of Huntington's disease (HD) symptoms and age at death of HD patients. Dichotomous analysis comparing HD to controls is widely used to study the effects of HTT CAG repeat expansion. However, a potentially more powerful approach is a continuous analysis strategy that takes advantage of all of the different CAG lengths, to capture effects that are expected to be critical to HD pathogenesis.

Expressomal approach for comprehensive analysis and visualization of ligand sensitivities of xenoestrogen responsive genes.

Although biological effects of endocrine disrupting chemicals (EDCs) are often observed at unexpectedly low doses with occasional nonmonotonic dose-response characteristics, transcriptome-wide profiles of sensitivities or dose-dependent behaviors of the EDC responsive genes have remained unexplored. Here, we describe expressome analysis for the comprehensive examination of dose-dependent gene responses and its applications to characterize estrogen responsive genes in MCF-7 cells. Transcriptomes of MCF-7 cells exposed to varying concentrations of representative natural and xenobiotic estrogens for 48 h were determined by microarray and used for computational calculation of interpolated approximations of estimated transcriptomes for 300 doses uniformly distributed in log space for each chemical.

Dominant effects of the Huntington's disease HTT CAG repeat length are captured in gene-expression data sets by a continuous analysis mathematical modeling strategy.

In Huntington's disease (HD), the size of the expanded HTT CAG repeat mutation is the primary driver of the processes that determine age at onset of motor symptoms. However, correlation of cellular biochemical parameters also extends across the normal repeat range, supporting the view that the CAG repeat represents a functional polymorphism with dominant effects determined by the longer allele. A central challenge to defining the functional consequences of this single polymorphism is the difficulty of distinguishing its subtle effects from the multitude of other sources of biological variation.

Fulvestrant-induced cell death and proteasomal degradation of estrogen receptor α protein in MCF-7 cells require the CSK c-Src tyrosine kinase.

Fulvestrant is a representative pure antiestrogen and a Selective Estrogen Receptor Down-regulator (SERD). In contrast to the Selective Estrogen Receptor Modulators (SERMs) such as 4-hydroxytamoxifen that bind to estrogen receptor α (ERα) as antagonists or partial agonists, fulvestrant causes proteasomal degradation of ERα protein, shutting down the estrogen signaling to induce proliferation arrest and apoptosis of estrogen-dependent breast cancer cells. We performed genome-wide RNAi knockdown screenings for protein kinases required for fulvestrant-induced apoptosis of the MCF-7 estrogen-dependent human breast caner cells and identified the c-Src tyrosine kinase (CSK), a negative regulator of the oncoprotein c-Src and related protein tyrosine kinases, as one of the necessary molecules.

Dose-related estrogen effects on gene expression in fetal mouse prostate mesenchymal cells.

Developmental exposure of mouse fetuses to estrogens results in dose-dependent permanent effects on prostate morphology and function. Fetal prostatic mesenchyme cells express estrogen receptor alpha (ERα) and androgen receptors and convert stimuli from circulating estrogens and androgens into paracrine signaling to regulate epithelial cell proliferation and differentiation. To obtain mechanistic insight into the role of different doses of estradiol (E2) in regulating mesenchymal cells, we examined E2-induced transcriptomal changes in primary cultures of fetal mouse prostate mesenchymal cells.

Methods for study of neuronal morphogenesis: ex vivo RNAi electroporation in embryonic murine cerebral cortex.

The cerebral cortex directs higher cognitive functions. This six layered structure is generated in an inside-first, outside-last manner, in which the first born neurons remain closer to the ventricle while the last born neurons migrate past the first born neurons towards the surface of the brain. In addition to neuronal migration, a key process for normal cortical function is the regulation of neuronal morphogenesis.

Masculine epigenetic sex marks of the CYP19A1/aromatase promoter in genetically male chicken embryonic gonads are resistant to estrogen-induced phenotypic sex conversion.

Sex of birds is genetically determined through inheritance of the ZW sex chromosomes (ZZ males and ZW females). Although the mechanisms of avian sex determination remains unknown, the genetic sex is experimentally reversible by in ovo exposure to exogenous estrogens (ZZ-male feminization) or aromatase inhibitors (ZW-female masculinization). Expression of various testis- and ovary-specific marker genes during the normal and reversed gonadal sex differentiation in chicken embryos has been extensively studied, but the roles of sex-specific epigenetic marks in sex differentiation are unknown.

Differential effects of the Huntington's disease CAG mutation in striatum and cerebellum are quantitative not qualitative.

Huntington's disease (HD) involves marked early neurodegeneration in the striatum, whereas the cerebellum is relatively spared despite the ubiquitous expression of full-length mutant huntingtin, implying that inherent tissue-specific differences determine susceptibility to the HD CAG mutation. To understand this tissue specificity, we compared early mutant huntingtin-induced gene expression changes in striatum to those in cerebellum in young Hdh CAG knock-in mice, prior to onset of evident pathological alterations. Endogenous levels of full-length mutant huntingtin caused qualitatively similar, but quantitatively different gene expression changes in the two brain regions.

HD CAG-correlated gene expression changes support a simple dominant gain of function.

Huntington's disease is initiated by the expression of a CAG repeat-encoded polyglutamine region in full-length huntingtin, with dominant effects that vary continuously with CAG size. The mechanism could involve a simple gain of function or a more complex gain of function coupled to a loss of function (e.g.

Genome-wide analysis of YY2 versus YY1 target genes.

Yin Yang 1 (YY1) is a critical transcription factor controlling cell proliferation, development and DNA damage responses. Retrotranspositions have independently generated additional YY family members in multiple species. Although Drosophila YY1 [pleiohomeotic (Pho)] and its homolog [pleiohomeotic-like (Phol)] redundantly control homeotic gene expression, the regulatory contributions of YY1-homologs have not yet been examined in other species.

Antiestrogen-resistant subclones of MCF-7 human breast cancer cells are derived from a common monoclonal drug-resistant progenitor.

Emergence of antiestrogen-resistant cells in MCF-7 cells during suppression of estrogen signaling is a widely accepted model of acquired breast cancer resistance to endocrine therapy. To obtain insight into the genomic basis of endocrine therapy resistance, we characterized MCF-7 monoclonal sublines that survived 21-day exposure to tamoxifen (T-series sublines) or fulvestrant (F-series sublines) and sublines unselected by drugs (U-series). All T/F-sublines were resistant to the cytocidal effects of both tamoxifen and fulvestrant.

Regulation of expression of BIK proapoptotic protein in human breast cancer cells: p53-dependent induction of BIK mRNA by fulvestrant and proteasomal degradation of BIK protein.

Induction of mRNA for BIK proapoptotic protein by doxorubicin or gamma-irradiation requires the DNA-binding transcription factor activity of p53. In MCF7 cells, pure antiestrogen fulvestrant also induces BIK mRNA and apoptosis. Here, we provide evidence that, in contrast to doxorubicin or gamma-irradiation, fulvestrant induction of BIK mRNA is not a direct effect of the transcriptional activity of p53, although p53 is necessary for this induction.

Importance of dosage standardization for interpreting transcriptomal signature profiles: evidence from studies of xenoestrogens.

To obtain insights into similarities and differences in the biological actions of related drugs or toxic agents, their transcriptomal signature profiles (TSPs) have been examined in a large number of studies. However, many such reports did not provide proper justification for the dosage criteria of each agent. Using a well characterized cell culture model of estrogen-dependent proliferation of MCF7 human breast cancer cells, we demonstrate how different approaches to dosage standardization exert critical influences on TSPs, leading to different and even conflicting conclusions.

The Bik BH3-only protein is induced in estrogen-starved and antiestrogen-exposed breast cancer cells and provokes apoptosis.

Evidence has been accumulating that some estrogen-dependent human breast cancers require estrogen for not only proliferation but also survival. To obtain insights into the molecular mechanisms of apoptosis of breast cancer cells subjected to estrogen starvation or exposed to antiestrogens, we characterized changes in the gene expression profile of MCF-7/BUS human breast cancer cells and revealed a strong induction of Bik, a member of the BH3-only proapoptotic proteins. The Bik mRNA transcript and protein were strongly induced by estrogen starvation or exposure to fulvestrant, a pure antiestrogen that competes with the natural estrogens for binding to the estrogen receptors.

Global analysis of ligand sensitivity of estrogen inducible and suppressible genes in MCF7/BUS breast cancer cells by DNA microarray.

To obtain comprehensive information on 17beta-estradiol (E2) sensitivity of genes that are inducible or suppressible by this hormone, we designed a method that determines ligand sensitivities of large numbers of genes by using DNA microarray and a set of simple Perl computer scripts implementing the standard metric statistics. We used it to characterize effects of low (0-100 pM) concentrations of E2 on the transcriptome profile of MCF7/BUS human breast cancer cells, whose E2 dose-dependent growth curve saturated with 100 pM E2. Evaluation of changes in mRNA expression for all genes covered by the DNA microarray indicated that, at a very low concentration (10 pM), E2 suppressed approximately 3-5 times larger numbers of genes than it induced, whereas at higher concentrations (30-100 pM) it induced approximately 1.

Cloning of mouse Cited4, a member of the CITED family p300/CBP-binding transcriptional coactivators: induced expression in mammary epithelial cells.

The CITED family proteins bind to CBP/p300 transcriptional integrators through their conserved C-terminal acidic domain and function as coactivators. The 21-kDa mouse Cited4 protein, a novel member of the CITED family, interacted with CBP/p300 as well as isoforms of the TFAP2 transcription factor, coactivating TFAP2-dependent transcription. The cited4 gene consisted of only a single exon located on chromosome 4 at 56.