1. Biochemistry and Chemical Biology
  2. Cell Biology
Download icon

Continuous transport of a small fraction of plasma membrane cholesterol to endoplasmic reticulum regulates total cellular cholesterol

  1. Rodney Elwood Infante  Is a corresponding author
  2. Arun Radhakrishnan  Is a corresponding author
  1. University of Texas Southwestern Medical Center, United States
Research Article
  • Cited 49
  • Views 2,494
  • Annotations
Cite this article as: eLife 2017;6:e25466 doi: 10.7554/eLife.25466

Abstract

Cells employ regulated transport mechanisms to ensure that their plasma membranes (PMs) are optimally supplied with cholesterol derived from uptake of low-density lipoproteins (LDL) and synthesis. To date, all inhibitors of cholesterol transport block steps in lysosomes, limiting our understanding of post-lysosomal transport steps. Here, we establish the cholesterol-binding domain 4 of anthrolysin O (ALOD4) as a reversible inhibitor of cholesterol transport from PM to endoplasmic reticulum (ER). Using ALOD4, we: 1) deplete ER cholesterol without altering PM or overall cellular cholesterol levels; 2) demonstrate that LDL-derived cholesterol travels from lysosomes first to PM to meet cholesterol needs, and subsequently from PM to regulatory domains of ER to suppress activation of SREBPs, halting cholesterol uptake and synthesis; and 3) determine that continuous PM-to-ER cholesterol transport allows ER to constantly monitor PM cholesterol levels, and respond rapidly to small declines in cellular cholesterol by activating SREBPs, increasing cholesterol uptake and synthesis.

Article and author information

Author details

  1. Rodney Elwood Infante

    Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, United States
    For correspondence
    rodney.infante@utsouthwestern.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2605-822X
  2. Arun Radhakrishnan

    Department of Molecular Genetics, University of Texas Southwestern Medical Center, Dallas, United States
    For correspondence
    arun.radhakrishnan@utsouthwestern.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7266-7336

Funding

National Institutes of Health (HL20948)

  • Rodney Elwood Infante
  • Arun Radhakrishnan

Welch Foundation (I-1793)

  • Arun Radhakrishnan

American Heart Association (12SDG12040267)

  • Arun Radhakrishnan

National Institutes of Health (T32DK007745)

  • Rodney Elwood Infante

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

Reviewing Editor

  1. Stephen G Young, University of California, Los Angeles, United States

Publication history

  1. Received: January 25, 2017
  2. Accepted: April 16, 2017
  3. Accepted Manuscript published: April 17, 2017 (version 1)
  4. Version of Record published: May 16, 2017 (version 2)

Copyright

© 2017, Infante & Radhakrishnan

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

  • 2,494
    Page views
  • 728
    Downloads
  • 49
    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)

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

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

  1. Further reading

Further reading

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Marie E Sweet et al.
    Research Article

    KdpFABC is an ATP-dependent K+ pump that ensures bacterial survival in K+-deficient environments. Whereas transcriptional activation of kdpFABC expression is well studied, a mechanism for down regulation when K+ levels are restored has not been described. Here we show that KdpFABC is inhibited when cells return to a K+-rich environment. The mechanism of inhibition involves phosphorylation of Ser162 on KdpB, which can be reversed in vitro by treatment with serine phosphatase. Mutating Ser162 to Alanine produces constitutive activity, whereas the phosphomimetic Ser162Asp mutation inactivates the pump. Analyses of the transport cycle show that serine phosphorylation abolishes the K+-dependence of ATP hydrolysis and blocks the catalytic cycle after formation of the aspartyl phosphate intermediate (E1~P). This regulatory mechanism is unique amongst P-type pumps and this study furthers our understanding of how bacteria control potassium homeostasis to maintain cell volume and osmotic potential.

    1. Biochemistry and Chemical Biology
    Aleksandra Bebel et al.
    Research Article

    Mobile genetic elements (MGEs) are a rich source of new enzymes, and conversely, understanding the activities of MGE-encoded proteins can elucidate MGE function. Here we biochemically characterize 3 proteins encoded by a conserved operon carried by the Staphylococcal Cassette Chromosome (SCCmec), an MGE that confers methicillin resistance to Staphylococcus aureus, creating MRSA strains. The first of these proteins, CCPol, is an active A-family DNA polymerase. The middle protein, MP, binds tightly to CCPol and confers upon it the ability to synthesize DNA primers de novo. The CCPol-MP complex is therefore a unique primase-polymerase enzyme unrelated to either known primase family. The third protein, Cch2, is a 3'-to-5' helicase. Cch2 additionally binds specifically to a dsDNA sequence downstream of its gene that is also a preferred initiation site for priming by CCPol-MP. Taken together, our results suggest that this is a functional replication module for SCCmec.