Abstract

Fibrolamellar carcinoma (FLC) is a rare liver cancer. FLCs uniquely produce DNAJ-PKAc, a chimeric enzyme consisting of a chaperonin-binding domain fused to the Ca subunit of protein kinase A. Biochemical analyses of clinical samples reveal that a unique property of this fusion enzyme is the ability to recruit heat shock protein 70 (Hsp70). This cellular chaperonin is frequently up-regulated in cancers. Gene-editing of mouse hepatocytes generated disease-relevant AML12DNAJ-PKAc cell lines. Further analyses indicate that the proto-oncogene A-kinase anchoring protein-Lbc is up-regulated in FLC and functions to cluster DNAJ-PKAc/Hsp70 sub-complexes with a RAF-MEK-ERK kinase module. Drug screening reveals Hsp70 and MEK inhibitor combinations that selectively block proliferation of AML12DNAJ-PKAc cells. Phosphoproteomic profiling demonstrates that DNAJ-PKAc biases the signaling landscape toward ERK activation and engages downstream kinase cascades. Thus, the oncogenic action of DNAJ-PKAc involves an acquired scaffolding function that permits recruitment of Hsp70 and mobilization of local ERK signaling.

Data availability

There are no restrictions to the availability of our data. Raw mass spectrometry data has been uploaded to MassIVE, an NIH supported MS data repository (MSV000083167). Data will be made publicly available.

The following data sets were generated

Article and author information

Author details

  1. Rigney E Turnham

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. F Donelson Smith

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8080-7589
  3. Heidi L Kenerson

    Department of Surgery, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Mitchell H Omar

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Martin Golkowski

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Irvin Garcia

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Renay Bauer

    Department of Surgery, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Ho-Tak Lau

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Kevin M Sullivan

    Department of Surgery, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  10. Lorene K Langeberg

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3760-7813
  11. Shao-En Ong

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  12. Kimberly J Riehle

    Department of Surgery, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  13. Raymond S Yeung

    Department of Surgery, University of Washington Medical Center, Seattle, United States
    Competing interests
    The authors declare that no competing interests exist.
  14. John D Scott

    Department of Pharmacology, University of Washington Medical Center, Seattle, United States
    For correspondence
    scottjdw@uw.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0367-8146

Funding

National Institute of Diabetes and Digestive and Kidney Diseases (R01DK119192)

  • John D Scott

Fibrolamellar Cancer Foundation

  • John D Scott

National Cancer Institute (R21CA201867)

  • Kimberly J Riehle

St. Baldrick's Foundation

  • Kimberly J Riehle

National Cancer Institute (R21CA177402)

  • Shao-En Ong

NIH Office of the Director (S10 OD021502)

  • Shao-En Ong

National Institutes of Health (2T32CA080416)

  • Rigney E Turnham

National Institute of Diabetes and Digestive and Kidney Diseases (F32DK121415)

  • Mitchell H Omar

Fibrolamellar Cancer Foundation

  • Raymond S Yeung

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

Reviewing Editor

  1. Roger J Davis, University of Massachusetts Medical School, United States

Version history

  1. Received: December 6, 2018
  2. Accepted: May 5, 2019
  3. Accepted Manuscript published: May 7, 2019 (version 1)
  4. Version of Record published: May 23, 2019 (version 2)

Copyright

© 2019, Turnham 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

  • 2,913
    Page views
  • 334
    Downloads
  • 33
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, Scopus, PubMed Central.

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. Rigney E Turnham
  2. F Donelson Smith
  3. Heidi L Kenerson
  4. Mitchell H Omar
  5. Martin Golkowski
  6. Irvin Garcia
  7. Renay Bauer
  8. Ho-Tak Lau
  9. Kevin M Sullivan
  10. Lorene K Langeberg
  11. Shao-En Ong
  12. Kimberly J Riehle
  13. Raymond S Yeung
  14. John D Scott
(2019)
An acquired scaffolding function of the DNAJ-PKAc fusion contributes to oncogenic signaling in fibrolamellar carcinoma
eLife 8:e44187.
https://doi.org/10.7554/eLife.44187

Share this article

https://doi.org/10.7554/eLife.44187

Further reading

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Kristian Davidsen, Jonathan S Marvin ... Lucas B Sullivan
    Research Article

    Intracellular levels of the amino acid aspartate are responsive to changes in metabolism in mammalian cells and can correspondingly alter cell function, highlighting the need for robust tools to measure aspartate abundance. However, comprehensive understanding of aspartate metabolism has been limited by the throughput, cost, and static nature of the mass spectrometry (MS)-based measurements that are typically employed to measure aspartate levels. To address these issues, we have developed a green fluorescent protein (GFP)-based sensor of aspartate (jAspSnFR3), where the fluorescence intensity corresponds to aspartate concentration. As a purified protein, the sensor has a 20-fold increase in fluorescence upon aspartate saturation, with dose-dependent fluorescence changes covering a physiologically relevant aspartate concentration range and no significant off target binding. Expressed in mammalian cell lines, sensor intensity correlated with aspartate levels measured by MS and could resolve temporal changes in intracellular aspartate from genetic, pharmacological, and nutritional manipulations. These data demonstrate the utility of jAspSnFR3 and highlight the opportunities it provides for temporally resolved and high-throughput applications of variables that affect aspartate levels.

    1. Biochemistry and Chemical Biology
    Chi-Ning Chuang, Hou-Cheng Liu ... Ting-Fang Wang
    Research Article

    Serine(S)/threonine(T)-glutamine(Q) cluster domains (SCDs), polyglutamine (polyQ) tracts and polyglutamine/asparagine (polyQ/N) tracts are Q-rich motifs found in many proteins. SCDs often are intrinsically disordered regions that mediate protein phosphorylation and protein-protein interactions. PolyQ and polyQ/N tracts are structurally flexible sequences that trigger protein aggregation. We report that due to their high percentages of STQ or STQN amino acid content, four SCDs and three prion-causing Q/N-rich motifs of yeast proteins possess autonomous protein expression-enhancing activities. Since these Q-rich motifs can endow proteins with structural and functional plasticity, we suggest that they represent useful toolkits for evolutionary novelty. Comparative Gene Ontology (GO) analyses of the near-complete proteomes of 26 representative model eukaryotes reveal that Q-rich motifs prevail in proteins involved in specialized biological processes, including Saccharomyces cerevisiae RNA-mediated transposition and pseudohyphal growth, Candida albicans filamentous growth, ciliate peptidyl-glutamic acid modification and microtubule-based movement, Tetrahymena thermophila xylan catabolism and meiosis, Dictyostelium discoideum development and sexual cycles, Plasmodium falciparum infection, and the nervous systems of Drosophila melanogaster, Mus musculus and Homo sapiens. We also show that Q-rich-motif proteins are expanded massively in 10 ciliates with reassigned TAAQ and TAGQ codons. Notably, the usage frequency of CAGQ is much lower in ciliates with reassigned TAAQ and TAGQ codons than in organisms with expanded and unstable Q runs (e.g. D. melanogaster and H. sapiens), indicating that the use of noncanonical stop codons in ciliates may have coevolved with codon usage biases to avoid triplet repeat disorders mediated by CAG/GTC replication slippage.