Structural basis for allosteric control of the SERCA-Phospholamban membrane complex by Ca2+ and phosphorylation

  1. Daniel K Weber
  2. U Venkateswara Reddy
  3. Songlin Wang
  4. Erik K Larsen
  5. Tata Gopinath
  6. Martin B Gustavsson
  7. Razvan L Cornea
  8. David D Thomas
  9. Alfonso De Simone
  10. Gianluigi Veglia  Is a corresponding author
  1. University of Minnesota, United States
  2. Imperial College London, United Kingdom

Abstract

Phospholamban (PLN) is a mini-membrane protein that directly controls the cardiac Ca2+-transport response to b-adrenergic stimulation, thus modulating cardiac output during the fight-or-flight response. In the sarcoplasmic reticulum membrane, PLN binds to the sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA), keeping this enzyme's function within a narrow physiological window. PLN phosphorylation by cAMP-dependent protein kinase A or increase in Ca2+ concentration reverses the inhibitory effects through an unknown mechanism. Using oriented-sample solid-state NMR spectroscopy and replica-averaged NMR-restrained structural refinement, we reveal that phosphorylation of PLN;s cytoplasmic regulatory domain signals the disruption of several inhibitory contacts at the transmembrane binding interface of the SERCA-PLN complex that are propagated to the enzyme;s active site, augmenting Ca2+ transport. Our findings address long-standing questions about SERCA regulation, epitomizing a signal transduction mechanism operated by posttranslationally modified bitopic membrane proteins.

Data availability

Here are the links/codes for data deposited:BMRB:50718:Monomeric phospholamban in oriented bicelles;50719:Monomeric phosphorylated phospholamban in oriented bicelles;50720:Phospholamban bound to SERCA in oriented bicelles (calcium-free E2 state);50721:Phospholamban bound to SERCA in oriented bicelles (calcium-bound E1 state);50722:Phosphorylated phospholamban bound to SERCA in oriented bicelles(calcium-free E2 state);50723:Phosphorylated phospholamban bound to SERCA in oriented bicelles(calcium-bound E1 state).And the link to DRUM:https://conservancy.umn.edu/handle/11299/218010

Article and author information

Author details

  1. Daniel K Weber

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. U Venkateswara Reddy

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Songlin Wang

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  4. Erik K Larsen

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  5. Tata Gopinath

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Martin B Gustavsson

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. Razvan L Cornea

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. David D Thomas

    Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8822-2040
  9. Alfonso De Simone

    Division of Molecular Biosciences, Imperial College London, London, United Kingdom
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8789-9546
  10. Gianluigi Veglia

    Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, United States
    For correspondence
    vegli001@umn.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2795-6964

Funding

National Institutes of Health (GM064742)

  • Gianluigi Veglia

National Institutes of Health (HL144100)

  • Gianluigi Veglia

National Institutes of Health (HL139065)

  • David D Thomas

National Institutes of Health (AG026160)

  • Razvan L Cornea

European Commission (BioDisOrder - 819644)

  • Alfonso De Simone

American Heart Association (19POST34420009)

  • Daniel K Weber

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

Reviewing Editor

  1. Volker Dötsch, Goethe University, Germany

Publication history

  1. Received: January 4, 2021
  2. Accepted: May 10, 2021
  3. Accepted Manuscript published: May 12, 2021 (version 1)
  4. Version of Record published: June 7, 2021 (version 2)

Copyright

© 2021, Weber 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

  • 1,108
    Page views
  • 185
    Downloads
  • 3
    Citations

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

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. Daniel K Weber
  2. U Venkateswara Reddy
  3. Songlin Wang
  4. Erik K Larsen
  5. Tata Gopinath
  6. Martin B Gustavsson
  7. Razvan L Cornea
  8. David D Thomas
  9. Alfonso De Simone
  10. Gianluigi Veglia
(2021)
Structural basis for allosteric control of the SERCA-Phospholamban membrane complex by Ca2+ and phosphorylation
eLife 10:e66226.
https://doi.org/10.7554/eLife.66226

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Lukas P Feilen et al.
    Research Article

    Cleavage of membrane proteins in the lipid bilayer by intramembrane proteases is crucial for health and disease. Although different lipid environments can potently modulate their activity, how this is linked to their structural dynamics is unclear. Here we show that the carboxy-peptidase-like activity of the archaeal intramembrane protease PSH, a homolog of the Alzheimer's disease-associated presenilin/γ-secretase is impaired in micelles and promoted in a lipid bilayer. Comparative molecular dynamics simulations revealed that important elements for substrate binding such as transmembrane domain 6a of PSH are more labile in micelles and stabilized in the lipid bilayer. Moreover, consistent with an enhanced interaction of PSH with a transition-state analog inhibitor, the bilayer promoted the formation of the enzyme´s catalytic active site geometry. Our data indicate that the lipid environment of an intramembrane protease plays a critical role in structural stabilization and active site arrangement of the enzyme-substrate complex thereby promoting intramembrane proteolysis.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    William J Allen et al.
    Research Article Updated

    Transport of proteins across and into membranes is a fundamental biological process with the vast majority being conducted by the ubiquitous Sec machinery. In bacteria, this is usually achieved when the SecY-complex engages the cytosolic ATPase SecA (secretion) or translating ribosomes (insertion). Great strides have been made towards understanding the mechanism of protein translocation. Yet, important questions remain – notably, the nature of the individual steps that constitute transport, and how the proton-motive force (PMF) across the plasma membrane contributes. Here, we apply a recently developed high-resolution protein transport assay to explore these questions. We find that pre-protein transport is limited primarily by the diffusion of arginine residues across the membrane, particularly in the context of bulky hydrophobic sequences. This specific effect of arginine, caused by its positive charge, is mitigated for lysine which can be deprotonated and transported across the membrane in its neutral form. These observations have interesting implications for the mechanism of protein secretion, suggesting a simple mechanism through which the PMF can aid transport by enabling a 'proton ratchet', wherein re-protonation of exiting lysine residues prevents channel re-entry, biasing transport in the outward direction.