Expression Characterization of AtPDI11 and Functional Analysis of AtPDI11 D Domain in Oxidative Protein Folding.

Fenggui Fan,Hao Zhang,Yahui Wei,Qian Wei

doi:10.3390/ijms23031409

Abstract

The formation and isomerization of disulfide bonds mediated by protein disulfide isomerase (PDI) in the endoplasmic reticulum (ER) is of fundamental importance in eukaryotes. Canonical PDI structure comprises four domains with the order of a-b-b′-a′. In Arabidopsis thaliana, the PDI-S subgroup contains only one member, AtPDI11, with an a-a′-D organization, which has no orthologs in mammals or yeast. However, the expression pattern of AtPDI11 and the functioning mechanism of AtPDI11 D domain are currently unclear. In this work, we found that PDI-S is evolutionarily conserved between land plants and algal organisms. AtPDI11 is expressed in various tissues and its induction by ER stress is disrupted in bzip28/60 and ire1a/b mutants that are null mutants of key components in the unfolded protein response (UPR) signal transduction pathway, suggesting that the induction of AtPDI11 by ER stress is mediated by the UPR signaling pathway. Furthermore, enzymatic activity assays and genetic evidence showed that the D domain is crucially important for the activities of AtPDI11. Overall, this work will help to further understand the working mechanism of AtPDI11 in catalyzing disulfide formation in plants.

Highlights

Most newly synthesized secretory and membrane proteins are folded in the endoplasmic reticulum (ER), in which an oxidative environment facilitates the formation of intramolecular disulfide bonds [1]
Using deletion mutation and biochemical analysis, we found that AtERO1 can interact with multiple AtPDIs, and AtPDIL members mainly serve as an isomerase, while AtPDI-M/S members are more efficient in accepting oxidizing equivalents from AtERO1 and catalyzing disulfide bond formation
In Arabidopsis, protein disulfide isomerase (PDI)-S subgroup member AtPDI11 was upregulated under different concentrations of DTT and Tm (Figures 2–4)

Summary

Introduction

Most newly synthesized secretory and membrane proteins are folded in the endoplasmic reticulum (ER), in which an oxidative environment facilitates the formation of intramolecular disulfide bonds [1]. The cleaved transcription factors bZIP60 and bZIP28 enter the nucleus to upregulate the expression of UPR related genes, such as binding protein (BIP), calnexin (CNX), and PDI, so as to facilitate protein folding and alleviate ER stress [7,8]. PDI-M subgroup has two members, AtPDI9 and AtPDI10 with a0-a-b domain arrangement, which is required for pollen viability and normal exine formation in plants subjected to heat stress [21]. Using deletion mutation and biochemical analysis, we found that AtERO1 can interact with multiple AtPDIs, and AtPDIL members mainly serve as an isomerase, while AtPDI-M/S members are more efficient in accepting oxidizing equivalents from AtERO1 and catalyzing disulfide bond formation. A similar model of cooperation of GmPDIM and GmPDIL-2 was found in soybeans [26]

Methods

Results

Conclusion