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Ultradian hormone stimulation induces glucocorticoid receptor-mediated pulses of gene transcription

Nature Cell Biology volume 11, pages 1093–1102 (2009)Cite this article

Abstract

Studies on glucocorticoid receptor (GR) action typically assess gene responses by long-term stimulation with synthetic hormones. As corticosteroids are released from adrenal glands in a circadian and high-frequency (ultradian) mode, such treatments may not provide an accurate assessment of physiological hormone action. Here we demonstrate that ultradian hormone stimulation induces cyclic GR-mediated transcriptional regulation, or gene pulsing, both in cultured cells and in animal models. Equilibrium receptor-occupancy of regulatory elements precisely tracks the ligand pulses. Nascent RNA transcripts from GR-regulated genes are released in distinct quanta, demonstrating a profound difference between the transcriptional programs induced by ultradian and constant stimulation. Gene pulsing is driven by rapid GR exchange with response elements and by GR recycling through the chaperone machinery, which promotes GR activation and reactivation in response to the ultradian hormone release, thus coupling promoter activity to the naturally occurring fluctuations in hormone levels. The GR signalling pathway has been optimized for a prompt and timely response to fluctuations in hormone levels, indicating that biologically accurate regulation of gene targets by GR requires an ultradian mode of hormone stimulation.

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Figure 1: GR–MMTV promoter interactions in response to ultradian corticosterone fluctuations.
Figure 2: GR-chaperone cycle.
Figure 3: GFP–GR loading and dynamics at the MMTV promoter array during ultradian stimulation with natural and synthetic glucocorticoids.
Figure 4: Equilibrium association of GR with endogenous GR response elements (GREs).
Figure 5: Cyclic RNA Polymerase II loading and dynamics in response to ultradian corticosterone treatment.
Figure 6: Effects of ultradian corticosterone treatment on transcription of GR-regulated genes in 3134 cells.
Figure 7: Differential transcription outputs of ultradian and constant hormone treatments.
Figure 8: Pulsatile corticosterone treatment of adrenalectomized (ADX) rats and a model for gene pulsing.

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Acknowledgements

We thank T. Karpova and J. McNally for assistance, C. Zeiss Inc. for the opportunity to carry out live cell experiments on a Zeiss Duoscan microscope, D. Shatti for technical assistance with nuclear extract preparation and Y. Kershaw for assistance with the in vivo experiments. This research was supported (in part) by the Intramural Research Program of the NIH, National Cancer Institute, Center for Cancer Research. The animal work was supported by a BBSRC grant (BB/C51297X/1) and a Wellcome Trust Programme Grant (074112/Z/04/Z) to S.L. The qPCR was performed on an Applied Biosystems 7500 System funded by a Wellcome Trust Equipment Grant (075548/Z/04/Z).

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Authors and Affiliations

  1. Laboratory of Receptor Biology and Gene Expression, Center for Cancer Research, National Cancer Institute, NIH, 41 Library Drive, Bethesda, 20892–5055, MD, USA

    Diana A. Stavreva, Malgorzata Wiench, Sam John, Thomas A. Johnson, Ty C. Voss & Gordon L. Hager

  2. Henry Wellcome Laboratories for Integrative Neuroscience and Endocrinology, The Dorothy Hodgkin Building, University of Bristol, Whitson Street, Bristol, BS1 3NY, UK

    Becky L. Conway-Campbell, Mervyn A. McKenna, John R. Pooley & Stafford L. Lightman

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Contributions

D.A.S. designed and performed most of the experiments and, together with G.L.H., wrote the initial draft of the manuscript; M.W. and D.A.S. performed transcription experiments, analysed data and participated in manuscript preparation and data interpretation; M.W. also performed all cell-line ChIP experiments; S.J. and T.A.J. performed confirmatory experiments, designed primers for PCR and participated in data interpretation and manuscript revision; T.V. performed some of the single cell analyses, helped with preparation of movies and participated in manuscript revision; B.C.C. and M.M. designed the live animal corticosterone pulsing experiments; J.P. performed surgery, conducted the live animal timecourse experiments and performed ChIP assays; B.CC. performed the TransAM DNA binding assays and prepared figures and text for the in vivo section; M.M. performed the corticosterone EIA and qPCR; S.L. provided data for the corticosterone profile, designed experiments together with G.L.H and D.A.S., provided valuable insights into data interpretation and participated in manuscript preparation; G.L.H. oversaw the project, designed many of the experiments and, together with D.A.S., prepared most figures, coordinated work among collaborators and participated in all stages of manuscript revision.

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Correspondence to Gordon L. Hager.

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Stavreva, D., Wiench, M., John, S. et al. Ultradian hormone stimulation induces glucocorticoid receptor-mediated pulses of gene transcription. Nat Cell Biol 11, 1093–1102 (2009). https://doi.org/10.1038/ncb1922

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