Abstract
Protein disulfide isomerases (PDIs) catalyze the formation, breakage, and rearrangement of disulfide bonds to properly fold nascent polypeptides within the endoplasmic reticulum (ER). Classical animal and yeast PDIs possess two catalytic thioredoxin-like domains (a, a′) and two non-catalytic domains (b, b′), in the order a-b-b′-a′. The model plant, Arabidopsis thaliana, encodes 12 PDI-like proteins, six of which possess the classical PDI domain arrangement (AtPDI1 through AtPDI6). Three additional AtPDIs (AtPDI9, AtPDI10, AtPDI11) possess two thioredoxin domains, but without intervening b-b′ domains. C-terminal green fluorescent protein (GFP) fusions to each of the nine dual-thioredoxin PDI homologs localized predominantly to the ER lumen when transiently expressed in protoplasts. Additionally, expression of AtPDI9:GFP-KDEL and AtPDI10: GFP-KDDL was associated with the formation of ER bodies. AtPDI9, AtPDI10, and AtPDI11 mediated the oxidative folding of alkaline phosphatase when heterologously expressed in the Escherichia coli protein folding mutant, dsbA−. However, only three classical AtPDIs (AtPDI2, AtPDI5, AtPDI6) functionally complemented dsbA−. Interestingly, chemical inducers of the ER unfolded protein response were previously shown to upregulate most of the AtPDIs that complemented dsbA−. The results indicate that Arabidopsis PDIs differ in their localization and protein folding activities to fulfill distinct molecular functions in the ER.
Highlights
Synthesized proteins entering the secretory pathway are folded into their native structures in the endoplasmic reticulum (ER)
We focused the analysis on the Arabidopsis orthologs of the mammalian protein disulfide isomerases (PDIs), PDIA1 and PDIA6
One notable difference between Homo sapiens PDIA1 (HsPDIA1) and its Arabidopsis homologs is the presence of a short, highly acidic region (17 Asp/Glu residues over a 24-a.a. interval) located near the C-terminus of HsPDIA1
Summary
Synthesized proteins entering the secretory pathway are folded into their native structures in the endoplasmic reticulum (ER). In addition to its function as a foldase, PDIA1 exhibits chaperone-like activity [9], and can aid the refolding of non-disulfide-bonded proteins in vitro [10,11]. The genome of the model plant, Arabidopsis thaliana, encodes at least 12 PDI-related proteins, including six PDIA1 orthologs (AtPDI1, AtPDI2, AtPDI3, AtPDI4, AtPDI5, AtPDI6) and two PDIA6 orthologs (AtPDI9, AtPDI10). PDIA6 orthologs have a subcellular localization pattern that is distinct from PDIA1 orthologs, and that overexpression of the PDIA6 orthologs in leaf protoplasts was associated with the formation of ER bodies Both Arabidopsis PDIA6 orthologs, and only three of the six PDIA1 orthologs, were able to functionally complement the Escherichia coli oxidative folding mutant, dsbA−. Our results provide further insight into the functional evolution of PDIs in plants
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