cAMP signaling regulates DNA hydroxymethylation by augmenting the intracellular labile ferrous iron pool

  1. Vladimir Camarena
  2. David W Sant
  3. Tyler C Huff
  4. Sushimita Mustafi
  5. Ryan K Muir
  6. Allegra T Aron
  7. Christopher J Chang
  8. Adam R Renslo
  9. Paula Monje
  10. Gaofeng Wang  Is a corresponding author
  1. University of Miami, United States
  2. University of California, San Francisco, United States
  3. University of California, Berkeley, United States

Abstract

It is widely accepted that cAMP regulates gene transcription principally by activating the protein kinase A (PKA)-targeted transcription factors. Here, we show that cAMP enhances the generation of 5-hydroxymethylcytosine (5hmC) in multiple cell types. 5hmC is converted from 5-methylcytosine (5mC) by Tet methylcytosine dioxygenases, for which Fe(II) is an essential cofactor. The promotion of 5hmC was mediated by a prompt increase of the intracellular labile Fe(II) pool (LIP). cAMP enhanced the acidification of endosomes for Fe(II) release to the LIP likely through RapGEF2. The effect of cAMP on Fe(II) and 5hmC was confirmed by adenylate cyclase activators, phosphodiesterase inhibitors, and most notably by stimulation of G protein-coupled receptors (GPCR). The transcriptomic changes caused by cAMP occurred in concert with 5hmC elevation in differentially transcribed genes. Collectively, these data show a previously unrecognized regulation of gene transcription by GPCR-cAMP signaling through augmentation of the intracellular labile Fe(II) pool and DNA hydroxymethylation.

Article and author information

Author details

  1. Vladimir Camarena

    Department of Human Genetics, University of Miami, Miami, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9466-6863
  2. David W Sant

    Department of Human Genetics, University of Miami, Miami, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Tyler C Huff

    Department of Human Genetics, University of Miami, Miami, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Sushimita Mustafi

    Department of Human Genetics, University of Miami, Miami, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Ryan K Muir

    Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Allegra T Aron

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Christopher J Chang

    Department of Chemistry, University of California, Berkeley, Berkeley, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Adam R Renslo

    Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Paula Monje

    Department of Neurological Surgery, University of Miami, Miami, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Gaofeng Wang

    Department of Human Genetics, University of Miami, Miami, United States
    For correspondence
    gwang@med.miami.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8202-8282

Funding

National Institute of Neurological Disorders and Stroke (R21CA191668)

  • Gaofeng Wang

National Cancer Institute (R01NS089525)

  • Gaofeng Wang

National Institute of General Medical Sciences (GM079465)

  • Christopher J Chang

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Copyright

© 2017, Camarena et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 2,516
    views
  • 438
    downloads
  • 33
    citations

Views, downloads and citations are aggregated across all versions of this paper published by eLife.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Vladimir Camarena
  2. David W Sant
  3. Tyler C Huff
  4. Sushimita Mustafi
  5. Ryan K Muir
  6. Allegra T Aron
  7. Christopher J Chang
  8. Adam R Renslo
  9. Paula Monje
  10. Gaofeng Wang
(2017)
cAMP signaling regulates DNA hydroxymethylation by augmenting the intracellular labile ferrous iron pool
eLife 6:e29750.
https://doi.org/10.7554/eLife.29750

Share this article

https://doi.org/10.7554/eLife.29750

Further reading

    1. Cell Biology
    2. Evolutionary Biology
    Paul Richard J Yulo, Nicolas Desprat ... Heather L Hendrickson
    Research Article

    Maintenance of rod-shape in bacterial cells depends on the actin-like protein MreB. Deletion of mreB from Pseudomonas fluorescens SBW25 results in viable spherical cells of variable volume and reduced fitness. Using a combination of time-resolved microscopy and biochemical assay of peptidoglycan synthesis, we show that reduced fitness is a consequence of perturbed cell size homeostasis that arises primarily from differential growth of daughter cells. A 1000-generation selection experiment resulted in rapid restoration of fitness with derived cells retaining spherical shape. Mutations in the peptidoglycan synthesis protein Pbp1A were identified as the main route for evolutionary rescue with genetic reconstructions demonstrating causality. Compensatory pbp1A mutations that targeted transpeptidase activity enhanced homogeneity of cell wall synthesis on lateral surfaces and restored cell size homeostasis. Mechanistic explanations require enhanced understanding of why deletion of mreB causes heterogeneity in cell wall synthesis. We conclude by presenting two testable hypotheses, one of which posits that heterogeneity stems from non-functional cell wall synthesis machinery, while the second posits that the machinery is functional, albeit stalled. Overall, our data provide support for the second hypothesis and draw attention to the importance of balance between transpeptidase and glycosyltransferase functions of peptidoglycan building enzymes for cell shape determination.

    1. Cell Biology
    2. Developmental Biology
    Pavan K Nayak, Arul Subramanian, Thomas F Schilling
    Research Article

    Mechanical forces play a critical role in tendon development and function, influencing cell behavior through mechanotransduction signaling pathways and subsequent extracellular matrix (ECM) remodeling. Here we investigate the molecular mechanisms by which tenocytes in developing zebrafish embryos respond to muscle contraction forces during the onset of swimming and cranial muscle activity. Using genome-wide bulk RNA sequencing of FAC-sorted tenocytes we identify novel tenocyte markers and genes involved in tendon mechanotransduction. Embryonic tendons show dramatic changes in expression of matrix remodeling associated 5b (mxra5b), matrilin1 (matn1), and the transcription factor kruppel-like factor 2a (klf2a), as muscles start to contract. Using embryos paralyzed either by loss of muscle contractility or neuromuscular stimulation we confirm that muscle contractile forces influence the spatial and temporal expression patterns of all three genes. Quantification of these gene expression changes across tenocytes at multiple tendon entheses and myotendinous junctions reveals that their responses depend on force intensity, duration and tissue stiffness. These force-dependent feedback mechanisms in tendons, particularly in the ECM, have important implications for improved treatments of tendon injuries and atrophy.