The nuclear receptor peroxisome proliferator-activated receptor-gamma (PPARgamma) has important roles in adipogenesis and immune response as well as roles in both lipid and carbohydrate metabolism. Although synthetic agonists for PPARgamma are widely used as insulin sensitizers, the identity of the natural ligand(s) for PPARgamma is still not clear. Suggested natural ligands include 15-deoxy-delta12,14-prostaglandin J2 and oxidized fatty acids such as 9-HODE and 13-HODE. Crystal structures of PPARgamma have revealed the mode of recognition for synthetic compounds. Here we report structures of PPARgamma bound to oxidized fatty acids that are likely to be natural ligands for this receptor. These structures reveal that the receptor can (i) simultaneously bind two fatty acids and (ii) couple covalently with conjugated oxo fatty acids. Thermal stability and gene expression analyses suggest that such covalent ligands are particularly effective activators of PPARgamma and thus may serve as potent and biologically relevant ligands.

SummaryPeroxisome proliferator-activated receptor γ (PPARγ) is a lipid-activated transcription factor regulating lipid metabolism and inflammatory response in macrophages and dendritic cells (DCs). These immune cells exposed to distinct inflammatory milieu show cell type specification as a result of altered gene expression. We demonstrate here a mechanism how inflammatory molecules modulate PPARγ signaling in distinct subsets of cells. Proinflammatory molecules inhibited whereas interleukin-4 (IL-4) stimulated PPARγ activity in macrophages and DCs. Furthermore, IL-4 signaling augmented PPARγ activity through an interaction between PPARγ and signal transducer and activators of transcription 6 (STAT6) on promoters of PPARγ target genes, including FABP4. Thus, STAT6 acts as a facilitating factor for PPARγ by promoting DNA binding and consequently increasing the number of regulated genes and the magnitude of responses. This interaction, underpinning cell type-specific responses, represents a unique way of controlling nuclear receptor signaling by inflammatory molecules in immune cells.

Summary In mammals, dosage compensation between XX and XY individuals occurs through X chromosome inactivation (XCI). The noncoding Xist RNA is expressed and initiates XCI only when more than one X chromosome is present. Current models invoke a dependency on the X-to-autosome ratio (X:A), but molecular factors remain poorly defined. Here, we demonstrate that molecular titration between an X-encoded RNA and an autosomally encoded protein dictates Xist induction. In pre-XCI cells, CTCF protein represses Xist transcription. At the onset of XCI, Jpx RNA is upregulated, binds CTCF, and extricates CTCF from one Xist allele. We demonstrate that CTCF is an RNA-binding protein and is titrated away from the Xist promoter by Jpx RNA. Thus, Jpx activates Xist by evicting CTCF. The functional antagonism via molecular titration reveals a role for long noncoding RNA in epigenetic regulation.

Retinoid X receptor (RXR) belongs to a family of ligandactivated transcription factors that regulate many aspects of metazoan life. A class of nuclear receptors requires RXR as heterodimerization partner for their function. This places RXR in the crossroad of multiple distinct biological pathways. This and the fact that the debate on the endogenous ligand requirement for RXR is not yet settled make RXR still an enigmatic transcription factor. Here, we review some of the biology of RXR. We place RXR into the evolution of nuclear receptors, review structural details and ligands of the receptor. Then processes regulated by RXR are discussed focusing on the developmental roles deduced from studies on knockout animals and metabolic roles in diseases such as diabetes and atherosclerosis deduced from pharmacological studies. Finally, aspects of RXR's involvement in myeloid differentiation and apoptosis are summarized along with issues on RXR's suitability as a therapeutic target.

Summary Although human induced pluripotent stem cells (hiPSCs) have enormous potential in regenerative medicine, their epigenetic variability suggests that some lines may not be suitable for human therapy. There are currently few benchmarks for assessing quality. Here we show that X-inactivation markers can be used to separate hiPSC lines into distinct epigenetic classes and that the classes are phenotypically distinct. Loss of XIST expression is strongly correlated with upregulation of X-linked oncogenes, accelerated growth rate in vitro, and poorer differentiation in vivo. Whereas differences in X-inactivation potential result in epigenetic variability of female hiPSC lines, male hiPSC lines generally resemble each other and do not overexpress the oncogenes. Neither physiological oxygen levels nor HDAC inhibitors offer advantages to culturing female hiPSC lines. We conclude that female hiPSCs may be epigenetically less stable in culture and caution that loss of XIST may result in qualitatively less desirable stem cell lines.

A key issue in the immune system is to generate specific cell types, often with opposing activities. The mechanisms of differentiation and subtype specification of immune cells such as macrophages and dendritic cells are critical to understand the regulatory principles and logic of the immune system. In addition to cytokines and pathogens, it is increasingly appreciated that lipid signaling also has a key role in differentiation and subtype specification. In this review we explore how intracellular lipid signaling via a set of transcription factors regulates cellular differentiation, subtype specification, and immune as well as metabolic homeostasis. We introduce macrophages and dendritic cells and then we focus on a group of transcription factors, nuclear receptors, which regulate gene expression upon receiving lipid signals. The receptors we cover are the ones with a recognized physiological function in these cell types and ones which heterodimerize with the retinoid X receptor. These are as follows: the receptor for a metabolite of vitamin A, retinoic acid: retinoic acid receptor (RAR), the vitamin D receptor (VDR), the fatty acid receptor: peroxisome proliferator-activated receptor γ (PPARγ), the oxysterol receptor liver X receptor (LXR), and their obligate heterodimeric partner, the retinoid X receptor (RXR). We discuss how they can get activated and how ligand is generated and eliminated in these cell types. We also explore how activation of a particular target gene contributes to biological functions and how the regulation of individual target genes adds up to the coordination of gene networks. It appears that RXR heterodimeric nuclear receptors provide these cells with a coordinated and interrelated network of transcriptional regulators for interpreting the lipid milieu and the metabolic changes to bring about gene expression changes leading to subtype and functional specification. We also show that these networks are implicated in various immune diseases and are amenable to therapeutic exploitation.

CTCF is a master regulator that plays important roles in genome architecture and gene expression. How CTCF is recruited in a locus-specific manner is not fully understood. Evidence from epigenetic processes, such as X-chromosome inactivation (XCI), indicates that CTCF associates functionally with RNA. Using genome-wide approaches to investigate the relationship between its RNA interactome and epigenomic landscape, here we report that CTCF binds thousands of transcripts in mouse embryonic stem cells, many in close proximity to CTCF’s genomic binding sites. CTCF is a specific and high-affinity RNA-binding protein (Kd <1 nM). During XCI, CTCF differentially binds the active and inactive X-chromosomes and interacts directly with Tsix, Xite, and Xist RNAs. Tsix and Xite RNAs target CTCF to the X-inactivation center, thereby inducing homologous X-chromosome pairing. Our work elucidates one mechanism by which CTCF is recruited in a locus-specific manner and implicates CTCF-RNA interactions in long-range chromosomal interactions.

ABCG2, a member of the ATP-binding cassette transporters has been identified as a protective pump against endogenous and exogenous toxic agents. ABCG2 was shown to be expressed at high levels in stem cells and variably regulated during cell differentiation. Here we demonstrate that functional ABCG2 is expressed in human monocyte-derived dendritic cells by the activation of a nuclear hormone receptor, PPAR␥. We identified and characterized a 150-base pair long conserved enhancer region, containing three functional PPAR response elements (PPARE), upstream of the human ABCG2 gene. We confirmed the binding of the PPAR␥⅐RXR heterodimer to this enhancer region, suggesting that PPAR␥ directly regulates the transcription of ABCG2. Consistent with these results, elevated expression of ABCG2 mRNA was coupled to enhanced protein production, resulting in increased xenobiotic extrusion capacity via ABCG2 in PPAR␥-activated cells. Furthermore PPAR␥ instructed dendritic cells showed increased Hoechst dye extrusion and resistance to mitoxantrone. Collectively, these results uncovered a mechanism by which up-regulation of functional ABCG2 expression can be achieved via exogenous or endogenous activation of the lipid-activated transcription factor, PPAR␥. The increased expression of the promiscuous ABCG2 transporter can significantly modify the xenobiotic and drug resistance of human myeloid dendritic cells.