ObjectivePancreatic ductal adenocarcinoma (PDA) has among the highest stromal fractions of any cancer and this has complicated attempts at expression-based molecular classification. The goal of this work is to profile purified samples of human PDA epithelium and stroma and examine their respective contributions to gene expression in bulk PDA samples.DesignWe used laser capture microdissection (LCM) and RNA sequencing to profile the expression of 60 matched pairs of human PDA malignant epithelium and stroma samples. We then used these data to train a computational model that allowed us to infer tissue composition and generate virtual compartment-specific expression profiles from bulk gene expression cohorts.ResultsOur analysis found significant variation in the tissue composition of pancreatic tumours from different public cohorts. Computational removal of stromal gene expression resulted in the reclassification of some tumours, reconciling functional differences between different cohorts. Furthermore, we established a novel classification signature from a total of 110 purified human PDA stroma samples, finding two groups that differ in the extracellular matrix-associated and immune-associated processes. Lastly, a systematic evaluation of cross-compartment subtypes spanning four patient cohorts indicated partial dependence between epithelial and stromal molecular subtypes.ConclusionOur findings add clarity to the nature and number of molecular subtypes in PDA, expand our understanding of global transcriptional programmes in the stroma and harmonise the results of molecular subtyping efforts across independent cohorts.

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The cycle of gallbladder filling and emptying controls the flow of bile into the intestine for digestion. Here we show that fibroblast growth factor-15, a hormone made by the distal small intestine in response to bile acids, is required for gallbladder filling. These studies demonstrate that gallbladder filling is actively regulated by an endocrine pathway and suggest a postprandial timing mechanism that controls gallbladder motility.

Liver X receptors (LXRα and LXRβ) are important regulators of cholesterol and lipid metabolism, and their activation has been shown to inhibit cardiovascular disease and reduce atherosclerosis in animal models. Small molecule agonists of LXR activity are therefore of great therapeutic interest. However, the finding that such agonists also promote hepatic lipogenesis has led to the idea that hepatic LXR activity is undesirable from a therapeutic perspective. To investigate whether this might be true, we performed gene targeting to selectively delete LXRα in hepatocytes. Liver-specific deletion of LXRα in mice substantially decreased reverse cholesterol transport, cholesterol catabolism, and cholesterol excretion, revealing the essential importance of hepatic LXRα for whole body cholesterol homeostasis. Additionally, in a pro-atherogenic background, liverspecific deletion of LXRα increased atherosclerosis, uncovering an important function for hepatic LXR activity in limiting cardiovascular disease. Nevertheless, synthetic LXR agonists still elicited anti-atherogenic activity in the absence of hepatic LXRα, indicating that the ability of agonists to reduce cardiovascular disease did not require an increase in cholesterol excretion. Furthermore, when non-atherogenic mice were treated with synthetic LXR agonists, liver-specific deletion of LXRα eliminated the detrimental effect of increased plasma triglycerides, while the beneficial effect of increased plasma HDL was unaltered. In sum, these observations suggest that therapeutic strategies that bypass the liver or limit the activation of hepatic LXRs should still be beneficial for the treatment of cardiovascular disease.

Functional interactions between factors bound at multiple sites on DNA often lead to a synergistic or more-than-additive transcriptional response. We previously defined a class of peptide sequences termed synergy control motifs (SC motifs) that function in multiple regulators by selectively inhibiting synergistic activity driven from multiple but not single response elements. By studying the prototypic SC motifs of the glucocorticoid receptor, we show that SC motifs inhibit transcription per se both in cis and in trans, and that a requirement for multiple contacts with DNA renders them selective for compound response elements. Notably, SC motifs are sites for SUMOylation, and the degree of modification correlates strongly with the extent of synergy control. Recruiting SUMO to the promoter either independently or as a fusion to the glucocorticoid receptor is sufficient to recapitulate the in trans and in cis inhibition by SC motifs without apparent changes in subcellular localization. Moreover, we find that the core ubiquitin fold domain of SUMO is sufficient for inhibition and that, independently of their potential for polySUMO chain formation, SUMO-2 and SUMO-3 are more effective inhibitors than SUMO-1. E ukaryotic transcriptional control is highly combinatorial, and the mosaic of response elements in the regulatory elements of a given gene nucleates the assembly of multiprotein complexes where various forms of functional interactions take place (1). One of the most prevalent is the more-than-additive or synergistic response resulting from the recruitment of an activator to multiple copies of a recognition site (compound response element). Despite their importance, the mechanisms that control synergistic effects are poorly understood (2).We have identified a short regulatory motif embedded in a number of sequence-specific regulators that is both necessary and sufficient to limit their transcriptional synergy (3). Disruption of these conserved synergy control (SC) motifs selectively enhances synergistic activation at compound response elements without altering the activity driven from a single site. Structure͞ function analysis of nine SC motifs from the glucocorticoid (GR), mineralocorticoid, and androgen receptors as well as from ETS-1 (3) and C͞EBP␣ (4) revealed a common core sequence (I͞V-K-X-E) and the presence of proline residues within 0-3 aa from either or both ends of the core. Such sequences occur in conserved regions of many factors and, in some cases, these regions have negative regulatory functions (e.g., SP3, SREBP, c-Myb, and C͞EBP). Moreover, a human mutation (P390S) in one of the SC motifs of the androgen receptor is associated with impaired spermatogenesis (5).A clue to the critical role of the Lys residues in SC motifs came from the identification of the consensus motif (⌿-K-x-E͞D) for the posttranslational modification by SUMO. Conjugation of this ubiquitin-like protein follows an analogous pathway to that of ubiquitination and requires dedicated E1 activating (SAE1͞ SAE2) and E2 conjugating ...