Shaheen Khan

Aryl hydrocarbon receptor agonists directly activate estrogen receptor α in MCF-7 breast cancer cells

Abstract The aryl hydrocarbon receptor (AhR) binds with high affinity to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and related halogenated aromatics, but also binds with lower affinity to structurally diverse exogenous and endogenous chemicals. One study reported that 3-methylcholanthrene (3MC) activated the estrogen receptor (ER) through the AhR, which acts as co-regulatory protein, whereas a recent report showed that 3MC directly bound and activated ERα. This study also shows that the AhR agonists benzo[a]pyrene, 3,3′,4,4′-tetrachlorobiphenyl, chrysin, 6-methyl-1,3,8-trichlorodibenzofuran, and 3,3′-diindolylmethane also induce ERα-dependent transactivation. Moreover, in chromatin immunoprecipitation assays, these compounds induce binding of AhR and ERα to the CYP1A1 and pS2 gene promoters, which is consistent with their activities as both selective AhR modulators (SAhRMs) and selective ER modulators (SERMs).

Activation of nerve growth factor-induced Bα by methylene-substituted diindolylmethanes in bladder cancer cells induces apoptosis and inhibits tumor growth

Nerve growth factor-induced B (NGFI-B) genes are orphan nuclear receptors, and NGFI-Bα (Nur77, TR3) is overexpressed in bladder tumors and bladder cancer cells compared with nontumorous bladder tissue. 1,1-Bis(3′-indolyl)-1-(p-methoxyphenyl)-methane (DIM-C-pPhOCH3) and 1,1-bis(3′-indolyl)-1-(p-phenyl)methane have previously been identified as activators of Nur77, and both compounds inhibited growth and induced apoptosis of UC-5 and KU7 bladder cancer cells. The proapoptotic effects of methylene-substituted diindolylmethanes (C-DIMs) were unaffected by cotreatment with leptomycin B and were dependent on nuclear Nur77, and RNA interference with a small inhibitory RNA for Nur77 (iNur77) demonstrated that C-DIM-induced activation of apoptosis was Nur77-dependent. Microarray analysis of DIM-C-pPhOCH3-induced genes in UC-5 bladder cancer cells showed that this compound induced multiple Nur77-dependent proapoptotic or growth inhibitory genes including tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), cystathionase, p21, p8, and sestrin-2. DIM-C-pPhOCH3 (25 mg/kg/d) also induced apoptosis and inhibited tumor growth in athymic nude mice bearing KU7 cells as xenografts, demonstrating that Nur77-active C-DIMs exhibit potential for bladder cancer chemotherapy by targeting Nur77, which is overexpressed in this tumor type.

Regulatory polymorphisms modulate the expression of HLA class II molecules and promote autoimmunity

Targeted sequencing of sixteen SLE risk loci among 1349 Caucasian cases and controls produced a comprehensive dataset of the variations causing susceptibility to systemic lupus erythematosus (SLE). Two independent disease association signals in the HLA-D region identified two regulatory regions containing 3562 polymorphisms that modified thirty-seven transcription factor binding sites. These extensive functional variations are a new and potent facet of HLA polymorphism. Variations modifying the consensus binding motifs of IRF4 and CTCF in the XL9 regulatory complex modified the transcription of HLA-DRB1, HLA-DQA1 and HLA-DQB1 in a chromosome-specific manner, resulting in a 2.5-fold increase in the surface expression of HLA-DR and DQ molecules on dendritic cells with SLE risk genotypes, which increases to over 4-fold after stimulation. Similar analyses of fifteen other SLE risk loci identified 1206 functional variants tightly linked with disease-associated SNPs and demonstrated that common disease alleles contain multiple causal variants modulating multiple immune system genes. DOI: http://dx.doi.org/10.7554/eLife.12089.001

DNA polymerase-α regulates the activation of type I interferons through cytosolic RNA:DNA synthesis

Aberrant nucleic acids generated during viral replication are the main trigger for antiviral immunity, and mutations that disrupt nucleic acid metabolism can lead to autoinflammatory disorders. Here we investigated the etiology of X-linked reticulate pigmentary disorder (XLPDR), a primary immunodeficiency with autoinflammatory features. We discovered that XLPDR is caused by an intronic mutation that disrupts the expression of POLA1, which encodes the catalytic subunit of DNA polymerase-α. Unexpectedly, POLA1 deficiency resulted in increased production of type I interferons. This enzyme is necessary for the synthesis of RNA:DNA primers during DNA replication and, strikingly, we found that POLA1 is also required for the synthesis of cytosolic RNA:DNA, which directly modulates interferon activation. Together this work identifies POLA1 as a critical regulator of the type I interferon response.

Erratum: Personalized immunomonitoring uncovers molecular networks that stratify lupus patients ((Cell (2016) 165 (551-565))

Romain Banchereau, Seunghee Hong, Brandi Cantarel, Nicole Baldwin, Jeanine Baisch, Michelle Edens, Alma-Martina Cepika, Peter Acs, Jacob Turner, Esperanza Anguiano, Parvathi Vinod, Shaheen Khan, Gerlinde Obermoser, Derek Blankenship, Edward Wakeland, Lorien Nassi, Alisa Gotte, Marilynn Punaro, Yong-Jun Liu, Jacques Banchereau, Jose Rossello-Urgell, Tracey Wright, and Virginia Pascual* *Correspondence: virginia.pascual@bswhealth.org http://dx.doi.org/10.1016/j.cell.2016.05.057

Ligand structure-dependent activation of estrogen receptor α/Sp by estrogens and xenoestrogens

This study investigated the effects of E2, diethylstilbestrol (DES), antiestrogens, the phytoestrogen resveratrol, and the xenoestrogens octylphenol (OP), nonylphenol (NP), endosulfan, kepone, 2,3,4,5-tetrachlorobiphenyl-4-ol (HO-PCB-Cl(4)), bisphenol-A (BPA), and 2,2-bis-(p-hydroxyphenyl)-1,1,1-trichloroethane (HPTE) on induction of luciferase activity in breast cancer cells transfected with a construct (pSp1(3)) containing three tandem GC-rich Sp binding sites linked to luciferase and wild-type or variant ERalpha. The results showed that induction of luciferase activity was highly structure-dependent in both MCF-7 and MDA-MB-231 cells. Moreover, RNA interference assays using small inhibitory RNAs for Sp1, Sp3 and Sp4 also demonstrated structure-dependent differences in activation of ERalpha/Sp1, ERalpha/Sp3 and ERalpha/Sp4. These results demonstrate for the first time that various structural classes of ER ligands differentially activate wild-type and variant ERalpha/Sp-dependent transactivation, selectively use different Sp proteins, and exhibit selective ER modulator (SERM)-like activity.

Role of specificity protein transcription factors in estrogen-induced gene expression in MCF-7 breast cancer cells

Deletion analysis of several 17beta-estradiol (E(2))-responsive genes have identified GC-rich sites that are associated with hormone-induced transactivation in MCF-7 breast cancer cells. However, the role of individual specificity proteins (Sps) in mediating hormone-induced gene expression has not been unequivocally determined. In transient transfection studies using E(2)-responsive GC-rich promoters from the E(2)F1, carbamoylphosphate synthetase/aspartate transcarbamylase/dihydroorotase (CAD), and retinoic acid receptor alpha (RAR alpha) genes, RNA interference using small inhibitory RNAs for Sp1 (iSp1), Sp3 (iSp3), and Sp4 (iSp4) decreased both basal and E(2)-induced transactivation. The contributions of individual Sp proteins to basal and E(2)-induced activity were promoter dependent. iSp1, iSp3, and iSp4 also significantly inhibited hormonal induction of E(2)F1, CAD, and RAR alpha mRNA levels; however, the enhanced inhibitory effects of the latter two small inhibitory RNAs suggest that Sp3 and Sp4 play a major role in estrogen receptor alpha/Sp-mediated gene expression in MCF-7 cells.

Cutting Edge: Inhibiting TBK1 by Compound II Ameliorates Autoimmune Disease in Mice.

TANK-binding kinase 1 (TBK1) is a serine/threonine protein kinase that plays a crucial role in innate immunity. Enhanced TBK1 function is associated with autoimmune diseases and cancer, implicating the potential benefit of therapeutically targeting TBK1. In this article, we examined a recently identified TBK1 inhibitor Compound II on treating autoimmune diseases. We found that Compound II is a potent and specific inhibitor of TBK1-mediated IFN response. Compound II inhibited polyinosinic-polycytidylic acid–induced immune activation in vitro and in vivo. Compound II treatment also ameliorated autoimmune disease phenotypes of Trex1−/− mice, increased mouse survival, and dampened the IFN gene signature in TREX1 mutant patient lymphoblasts. In addition, we found that TBK1 gene expression is elevated in systemic lupus erythematosus patient cells, and systemic lupus erythematosus cells with high IFN signature responded well to Compound II treatment. Together, our findings provided critical experimental evidence for inhibiting TBK1 with Compound II as an effective treatment for TREX1-associated autoimmune diseases and potentially other interferonopathies.

A genome-scale screen reveals context-dependent ovarian cancer sensitivity to miRNA overexpression

Large‐scale molecular annotation of epithelial ovarian cancer (EOC) indicates remarkable heterogeneity in the etiology of that disease. This diversity presents a significant obstacle against intervention target discovery. However, inactivation of miRNA biogenesis is commonly associated with advanced disease. Thus, restoration of miRNA activity may represent a common vulnerability among diverse EOC oncogenotypes. To test this, we employed genome‐scale, gain‐of‐function, miRNA mimic toxicity screens in a large, diverse spectrum of EOC cell lines. We found that all cell lines responded to at least some miRNA mimics, but that the nature of the miRNA mimics provoking a response was highly selective within the panel. These selective toxicity profiles were leveraged to define modes of action and molecular response indicators for miRNA mimics with tumor‐suppressive characteristics in vivo. A mechanistic principle emerging from this analysis was sensitivity of EOC to miRNA‐mediated release of cell fate specification programs, loss of which may be a prerequisite for development of this disease.

Fatty Acid Amide Hydrolase Regulates Peripheral B Cell Receptor Revision, Polyreactivity, and B1 Cells in Lupus.

C57BL/6 mice bearing the Sle2z lupus-susceptibility congenic interval on chromosome 4 display high titers of polyclonal autoantibodies with generalized B cell hyperactivity, hallmarks of systemic lupus erythematosus. In B6.Sle2zHELIg.sHEL BCR-transgenic mice, Sle2z did not breach central tolerance, but it led to heightened expression of endogenous Ig H and L chains in splenic B cells, upregulation of RAG, and serological polyreactivity, suggestive of excessive receptor revision. Fatty acid amide hydrolase (FAAH), a gene in the minimal subcongenic interval generated through recombinant mapping, was found to be upregulated in Sle2z B cells by microarray analysis, Western blot, and functional assays. Pharmacological inhibition of FAAH reversed the increase in receptor revision, RAG expression, and polyreactive autoantibodies in lupus-prone mice. These studies indicate that increased peripheral BCR revision, or selective peripheral expansion of BCR-revised B cells, may lead to systemic autoimmunity and that FAAH is a lupus-susceptibility gene that might regulate this process.