Evidence for the interaction of peroxiredoxin-4 with the store-operated calcium channel activator STIM1 in liver cells

Ka Cheung Tam, Eunus Ali, Jin Hua, Tim Chataway, Greg J. Barritt

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

Ca 2+ entry through store-operated Ca 2+ channels (SOCs) in the plasma membrane (PM) of hepatocytes plays a central role in the hormonal regulation of liver metabolism. SOCs are composed of Orai1, the channel pore protein, and STIM1, the activator protein, and are regulated by hormones and reactive oxygen species (ROS). In addition to Orai1, STIM1 also interacts with several other intracellular proteins. Most previous studies of the cellular functions of Orai1 and STIM1 have employed immortalised cells in culture expressing ectopic proteins tagged with a fluorescent polypeptide such as GFP. Little is known about the intracellular distributions of endogenous Orai1 and STIM1. The aims are to determine the intracellular distribution of endogenous Orai1 and STIM1 in hepatocytes and to identify novel STIM1 binding proteins. Subcellular fractions of rat liver were prepared by homogenisation and differential centrifugation. Orai1 and STIM1 were identified and quantified by western blot. Orai1 was found in the PM (0.03%), heavy (44%) and light (27%) microsomal fractions, and STIM1 in the PM (0.09%), and heavy (85%) and light (13%) microsomal fractions. Immunoprecipitation of STIM1 followed by LC/MS or western blot identified peroxiredoxin-4 (Prx-4) as a potential STIM1 binding protein. Prx-4 was found principally in the heavy microsomal fraction. Knockdown of Prx-4 using siRNA, or inhibition of Prx-4 using conoidin A, did not affect Ca 2+ entry through SOCs but rendered SOCs susceptible to inhibition by H 2 O 2 . It is concluded that, in hepatocytes, a considerable proportion of endogenous Orai1 and STIM1 is located in the rough ER. In the rough ER, STIM1 interacts with Prx-4, and this interaction may contribute to the regulation by ROS of STIM1 and SOCs.

Original languageEnglish
Pages (from-to)14-28
Number of pages15
JournalCell Calcium
Volume74
DOIs
StatePublished - Sep 2018

Bibliographical note

Publisher Copyright:
© 2018 Elsevier Ltd

Funding

This study was supported by grants from the Australian Research Council ( DP140100259 ), National Health and Medical Research Council ( 519115 ), Flinders University , and the Flinders Medical Centre Foundation .

FundersFunder number
Flinders Medical Centre Foundation
Australian Research CouncilDP140100259
Australian Research Council
Australian National Health and Medical Research Council519115
Australian National Health and Medical Research Council
Flinders University

    Keywords

    • Liver
    • Orai1
    • STIM1
    • Store-operated Ca channels
    • Subcellular fractionation
    • peroxiredoxin-4

    ASJC Scopus subject areas

    • Physiology
    • Molecular Biology
    • Cell Biology

    Fingerprint

    Dive into the research topics of 'Evidence for the interaction of peroxiredoxin-4 with the store-operated calcium channel activator STIM1 in liver cells'. Together they form a unique fingerprint.

    Cite this