Calcium signalling in mammalian cell lines expressing wild type and mutant human \u03b11-Antitrypsin

Nancy T Malintan,Steven D Buckingham,David A Lomas,David B Sattelle

doi:10.1038/s41598-019-53535-1

Nancy T Malintan, Steven D Buckingham + Show 2 more

Open Access

https://doi.org/10.1038/s41598-019-53535-1

Copy DOI

Journal: Scientific Reports	Publication Date: Nov 21, 2019
Citations: 3	License type: open-access

Affiliation: University College London

Abstract

A possible role for calcium signalling in the autosomal dominant form of dementia, familial encephalopathy with neuroserpin inclusion bodies (FENIB), has been proposed, which may point towards a mechanism by which cells could sense and respond to the accumulation of mutant serpin polymers in the endoplasmic reticulum (ER). We therefore explored possible defects in Ca2+-signalling, which may contribute to the pathology associated with another serpinopathy, α1-antitrypsin (AAT) deficiency. Using CHO K1 cell lines stably expressing a wild type human AAT (MAAT) and a disease-causing polymer-forming variant (ZAAT) and the truncated variant (NHK AAT), we measured basal intracellular free Ca2+, its responses to thapsigargin (TG), an ER Ca2+-ATPase blocker, and store-operated Ca2+-entry (SOCE). Our fura2 based Ca2+ measurements detected no differences between these 3 parameters in cell lines expressing MAAT and cell lines expressing ZAAT and NHK AAT mutants. Thus, in our cell-based models of α1-antitrypsin (AAT) deficiency, unlike the case for FENIB, we were unable to detect defects in calcium signalling.

Highlights

A possible role for calcium signalling in the autosomal dominant form of dementia, familial encephalopathy with neuroserpin inclusion bodies (FENIB), has been proposed, which may point towards a mechanism by which cells could sense and respond to the accumulation of mutant serpin polymers in the endoplasmic reticulum (ER)
Quantification by densitometry of the immunoblotting showed that cells expressing wild type AAT (MAAT) yielded significantly lower levels of AAT compared to those expressing the disease variants AAT (ZAAT and NHK AAT) (Fig. 1b)
The higher intracellular levels observed for ZAAT agreed with previously reported studies showing that the ZAAT variant is retained in the endoplasmic reticulum (ER) of the cells as AAT-protein polymers[25,66]

Summary

Introduction

A possible role for calcium signalling in the autosomal dominant form of dementia, familial encephalopathy with neuroserpin inclusion bodies (FENIB), has been proposed, which may point towards a mechanism by which cells could sense and respond to the accumulation of mutant serpin polymers in the endoplasmic reticulum (ER). The ER overload response is triggered[25,34], leading to ER stress Such ER swelling has been detected in both the ZAAT polymer-forming mutant cells and NHK-AAT cells containing a truncated AAT mutant in which the protein is degraded but not in the wild type (MAAT) cells[25]. In 2009 Davies et al suggested www.nature.com/scientificreports a possible role for calcium signalling in a serpinopathy, the autosomal dominant form of dementia familial encephalopathy with neuroserpin inclusion bodies (FENIB)[60]. Studies with thapsigargin (TG), an ER Ca2+-ATPase blocker, pointed to a role for calcium signalling in FENIB60

Methods

Results

Conclusion