• Open Access

Endoplasmic reticulum polymers impair luminal protein mobility and sensitize to cellular stress in alpha1-antitrypsin deficiency*

Authors

  • Adriana Ordóñez,

    1. Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, UK
    Search for more papers by this author
  • Erik L. Snapp,

    1. Department Anatomy and Structural Biology, Albert Einstein College of Medicine of Yeshiva University, Bronx, NY
    Search for more papers by this author
  • Lu Tan,

    1. Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, UK
    Search for more papers by this author
  • Elena Miranda,

    1. Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, UK
    2. Dipartimento di Biologia e Biotecnologie “Charles Darwin” e Istituto Pasteur–Fondazione Cenci Bolognetti, Università di Roma “La Sapienza,” Rome, Italy
    Search for more papers by this author
  • Stefan J. Marciniak,

    Corresponding author
    1. Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, UK
    • Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, Cambridge, CB2 0XY, UK
    Search for more papers by this author
    • fax: +44 (0)1223 336827

  • David A. Lomas

    1. Department of Medicine, University of Cambridge, Cambridge Institute for Medical Research, Cambridge, UK
    Search for more papers by this author
    • *

      Joint senior authors.


  • Potential conflict of interest: Nothing to report.

Abstract

Point mutants of alpha1-antitrypsin (α1AT) form ordered polymers that are retained as inclusions within the endoplasmic reticulum (ER) of hepatocytes in association with neonatal hepatitis, cirrhosis, and hepatocellular carcinoma. These inclusions cause cell damage and predispose to ER stress in the absence of the classical unfolded protein response (UPR). The pathophysiology underlying this ER stress was explored by generating cell models that conditionally express wild-type (WT) α1AT, two mutants that cause polymer-mediated inclusions and liver disease (E342K [the Z allele] and H334D) and a truncated mutant (Null Hong Kong; NHK) that induces classical ER stress and is removed by ER-associated degradation. Expression of the polymeric mutants resulted in gross changes in the ER luminal environment that recapitulated the changes observed in liver sections from individuals with PI*ZZ α1AT deficiency. In contrast, expression of NHK α1AT caused electron lucent dilatation and expansion of the ER throughout the cell. Photobleaching microscopy in live cells demonstrated a decrease in the mobility of soluble luminal proteins in cells that express E342K and H334D α1AT, when compared to those that express WT and NHK α1AT (0.34 ± 0.05, 0.22 ± 0.03, 2.83 ± 0.30, and 2.84 ± 0.55 μm2/s, respectively). There was no effect on protein mobility within ER membranes, indicating that cisternal connectivity was not disrupted. Polymer expression alone was insufficient to induce the UPR, but the resulting protein overload rendered cells hypersensitive to ER stress induced by either tunicamycin or glucose depletion. Conclusion: Changes in protein diffusion provide an explanation for the cellular consequences of ER protein overload in mutants that cause inclusion body formation and α1AT deficiency. (HEPATOLOGY 2013)

Ancillary