The nuclear receptor family member constitutive activated receptor (CAR) is activated by structurally diverse drugs and environmentally-relevant chemicals leading to transcriptional regulation of genes involved in xenobiotic metabolism and transport. Chronic activation of CAR increases liver cancer incidence in rodents, whereas suppression of CAR can lead to steatosis and insulin insensitivity. Here, analytical methods were developed to screen for chemical treatments in a gene expression compendium that lead to alteration of CAR activity. A gene expression biomarker signature of 83 CAR-dependent genes was identified using microarray profiles from the livers of wild-type and CAR-null mice after exposure to three structurally-diverse CAR activators (CITCO, phenobarbital, TCPOBOP). A rank-based algorithm (Running Fisher’s algorithm (p-value ≤ 10-4)) was used to evaluate the similarity between the CAR biomarker signature and a test set of 28 and 32 comparisons positive or negative, respectively, for CAR activation; the test resulted in a balanced accuracy of 97%. The biomarker signature was used to identify chemicals that activate or suppress CAR in an annotated mouse liver/primary hepatocyte gene expression database of ~1850 comparisons. CAR was activated by 1) activators of the aryl hydrocarbon receptor (AhR) in wild-type but not AhR-null mice, 2) pregnane X receptor (PXR) activators in wild-type and to lesser extents in PXR-null mice, and 3) activators of PPARα in wild-type and PPARα-null mice. CAR was consistently activated by five conazole fungicides and four perfluorinated compounds. Comparison of effects in wild-type and CAR-null mice showed that the fungicide propiconazole increased liver weight and hepatocyte proliferation in a CAR-dependent manner, whereas the perfluorinated compound perfluorooctanoic acid (PFOA) increased these endpoints in a CAR-independent manner. A number of compounds suppressed CAR coincident with increases in markers of inflammation including acetaminophen, concanavalin A, lipopolysaccharide, and 300 nm silica particles. In conclusion, we have shown that a CAR biomarker signature coupled with a rank-based similarity method accurately predicts CAR activation. This analytical approach, when applied to a gene expression compendium, increased the universe of known chemicals that directly or indirectly activate CAR, highlighting the promiscuous nature of CAR activation and signaling through activation of other xenobiotic-activated receptors.

Signal transducer and activator of transcription 5b (STAT5b) is a growth hormone (GH)-activated transcription factor and a master regulator of sexually dimorphic gene expression in the liver. Disruption of the GH hypothalamo-pituitary-liver axis controlling STAT5b activation can lead to metabolic dysregulation, steatosis, and liver cancer. Computational approaches were developed to identify factors that disrupt STAT5b function in a mouse liver gene expression compendium. A biomarker comprised of 144 STAT5b-dependent genes was derived using comparisons between wild-type male and wild-type female mice and between STAT5b-null and wild-type mice. Correlations between the STAT5b biomarker gene set and a test set comprised of expression datasets (biosets) with known effects on STAT5b function were evaluated using a rank-based test (the Running Fisher algorithm). Using a similarity p-value ≤ 10−4, the test achieved a balanced accuracy of 99% and 97% for detection of STAT5b activation or STAT5b suppression, respectively. The STAT5b biomarker gene set was then used to identify factors that activate (masculinize) or suppress (feminize) STAT5b function in an annotated mouse liver and primary hepatocyte gene expression compendium of ~1,850 datasets. Disruption of GH-regulated STAT5b is a common phenomenon in liver in vivo, with 5% and 29% of the male datasets, and 11% and 13% of the female datasets, associated with masculinization or feminization, respectively. As expected, liver STAT5b activation/masculinization occurred at puberty and suppression/feminization occurred during aging and in mutant mice with defects in GH signaling. A total of 70 genes were identified that have effects on STAT5b activation in genetic models in which the gene was inactivated or overexpressed. Other factors that affected liver STAT5b function were shown to include fasting, caloric restriction and infections. Together, these findings identify diverse factors that perturb the hypothalamo-pituitary-liver GH axis and disrupt GH-dependent STAT5b activation in mouse liver.