Adrenocorticotropic Hormone-Mediated Signaling Cascades Coordinate a Cyclic Pattern of Steroidogenic Factor 1-Dependent Transcriptional Activation

Department of Molecular and Integrative Pysiology, University of Michigan Medical School, Ann Arbor, Michigan 48109-0678, USA.
Molecular Endocrinology (Impact Factor: 4.02). 02/2006; 20(1):147-66. DOI: 10.1210/me.2005-0215
Source: PubMed


Steroidogenic factor 1 (SF-1) is an orphan nuclear receptor that has emerged as a critical mediator of endocrine function at multiple levels of the hypothalamic-pituitary-steroidogenic axis. Within the adrenal cortex, ACTH-dependent transcriptional responses, including transcriptional activation of several key steroidogenic enzymes within the steroid biosynthetic pathway, are largely dependent upon SF-1 action. The absence of a bona fide endogenous eukaryotic ligand for SF-1 suggests that signaling pathway activation downstream of the melanocortin 2 receptor (Mc2r) modulates this transcriptional response. We have used the chromatin immunoprecipitation assay to examine the temporal formation of ACTH-dependent transcription complexes on the Mc2r gene promoter. In parallel, ACTH-dependent signaling events were examined in an attempt to correlate transcriptional events with the upstream activation of signaling pathways. Our results demonstrate that ACTH-dependent signaling cascades modulate the temporal dynamics of SF-1-dependent complex assembly on the Mc2r promoter. Strikingly, the pattern of SF-1 recruitment and the subsequent attainment of active rounds of transcription support a kinetic model of SF-1 transcriptional activation, a model originally established in the context of ligand-dependent transcription by several classical nuclear hormone receptors. An assessment of the major ACTH-dependent signaling pathways highlights pivotal roles for the MAPK as well as the cAMP-dependent protein kinase A pathway in the entrainment of SF-1-mediated transcriptional events. In addition, the current study demonstrates that specific enzymatic activities are capable of regulating distinct facets of a highly ordered transcriptional response.

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    • "Mechanistically, SF-1 binds as a monomer to specific response elements in the promoters of its target genes. Bound SF-1 recruits either corepressor complexes, which put the gene in a silent state, or coactivator complexes, which activate transcription by altering histone modifications and recruiting the general transcription machinery including RNA polymerase II [14], [15], [16]. Structural studies have shown that SF-1 has a ligand-binding pocket that can accommodate phospholipids [17], [18], [19] and the search for a natural ligand is ongoing. "
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    ABSTRACT: The orphan nuclear receptor Steroidogenic Factor-1 (SF-1, NR5A1) is a critical regulator of development and homeostasis of the adrenal cortex and gonads. We recently showed that a complex containing E3 ubiquitin ligase RNF31 and the known SF-1 corepressor DAX-1 (NR0B1) interacts with SF-1 on target promoters and represses transcription of steroidogenic acute regulatory protein (StAR) and aromatase (CYP19) genes. To further evaluate the role of SF-1 in the adrenal cortex and the involvement of RNF31 in SF-1-dependent pathways, we performed genome-wide gene-expression analysis of adrenocortical NCI-H295R cells where SF-1 or RNF31 had been knocked down using RNA interference. We find RNF31 to be deeply connected to cholesterol metabolism and steroid hormone synthesis, strengthening its role as an SF-1 coregulator. We also find intriguing evidence of negative crosstalk between SF-1 and both transforming growth factor (TGF) β and Wnt/β-catenin signaling. This crosstalk could be of importance for adrenogonadal development, maintenance of adrenocortical progenitor cells and the development of adrenocortical carcinoma. Finally, the SF-1 gene profile can be used to distinguish malignant from benign adrenocortical tumors, a finding that implicates SF-1 in the development of malignant adrenocortical carcinoma.
    PLoS ONE 03/2012; 7(3):e32080. DOI:10.1371/journal.pone.0032080 · 3.23 Impact Factor
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    • "The contribution of these pathways to adrenocortical homeostasis has been reviewed recently (Kim et al., 2009). Additionally, studies into post-translational modifications of Sf1, such as phosphorylation (Campbell et al., 2008;Hammer et al., 1999;Lewis et al., 2008;Sewer and Waterman, 2002;Winnay and Hammer, 2006) and SUMOylation (Chen et al., 2004;Komatsu et al., 2004;Lee et al., 2005;Yang et al., 2009), may provide essential clues into the regulation of adrenal stem/progenitor cells through investigation of this critical mediator of adrenocortical specification. As new data become available, the mechanisms by which this network of signaling cascades and transcriptional regulators control adrenocortical stem/progenitor cell maintenance and differentiation should allow for a more complete understanding of adrenocortical physiology and disease. "
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    ABSTRACT: The origins of our understanding of the cellular and molecular mechanisms by which signaling pathways and downstream transcription factors coordinate the specification of adrenocortical cells within the adrenal gland have arisen from studies on the role of Sf1 in steroidogenesis and adrenal development initiated 20 years ago in the laboratory of Dr. Keith Parker. Adrenocortical stem/progenitor cells have been predicted to be undifferentiated and quiescent cells that remain at the periphery of the cortex until needed to replenish the organ, at which time they undergo proliferation and terminal differentiation. Identification of these stem/progenitor cells has only recently been explored. Recent efforts have examined signaling molecules, including Wnt, Shh, and Dax1, which may coordinate intricate lineage and signaling relationships between the adrenal capsule (stem cell niche) and underlying cortex (progenitor cell pool) to maintain organ homeostasis in the adrenal gland.
    Molecular and Cellular Endocrinology 11/2010; 336(1-2):206-12. DOI:10.1016/j.mce.2010.11.012 · 4.41 Impact Factor
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    • "Samples were then incubated overnight at 4 C in the presence of 1 μg of the indicated IgG from Millipore Corp. (Billerica, MA); immune complexes were recovered by absorption onto protein A-or protein G-agarose beads. The beads were washed extensively, immune complexes were extracted using 1% sodium dodecyl sulfate and 0.1 M NaHCO 3 and the extracted complexes were heated for 4 h at 65 C to reverse cross-links (Winnay and Hammer, 2006). The resultant DNA fragments were purified by resin absorption and elution using a DNA purification kit from Qiagen Inc. and analyzed by quantitative PCR as described above using oligonucleotide primer pairs listed in Appendix A. In each experiment, samples were corrected for total DNA input and normalized to the signal obtained with a control IgG. "
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    ABSTRACT: The contribution of steroidogenic factor 1 (SF-1) to the gene expression profile of Y1 mouse adrenocortical cells was evaluated using short hairpin RNAs to knockdown SF-1. The reduced level of SF-1 RNA was associated with global changes that affected the accumulation of more than 2000 transcripts. Among the down-regulated transcripts were several with functions in steroidogenesis that were affected to different degrees--i.e., Mc2r>Scarb1>Star≥Hsd3b1>Cyp11b1. For Star and Cyp11b1, the different levels of expression correlated with the amount of residual SF-1 bound to the proximal promoter regions. The knockdown of SF-1 did not affect the accumulation of Cyp11a1 transcripts even though the amount of SF-1 bound to the proximal promoter of the gene was reduced to background levels. Our results indicate that transcripts with functions in steroidogenesis vary in their dependence on SF-1 for constitutive expression. On a more global scale, SF-1 knockdown affects the accumulation of a large number of transcripts, most of which are not recognizably involved in steroid hormone biosynthesis.
    Molecular and Cellular Endocrinology 11/2010; 336(1-2):85-91. DOI:10.1016/j.mce.2010.11.024 · 4.41 Impact Factor
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