Zn2+-dependent functional switching of ERp18, an ER-resident thioredoxin-like protein.

ERp18 is an endoplasmic reticulum (ER)-resident thioredoxin (Trx) family protein, similar to cytosolic Trx1. The Trx-like domain occupies a major portion of the whole ERp18 structure, which is postulated to be an ER paralog of cytosolic Trx1. Here, we elucidate that zinc ion (Zn2+) binds ERp18 through its catalytic motif, triggering oligomerization of ERp18 from a monomer to a trimer. While the monomeric ERp18 has disulfide oxidoreductase activity, the trimeric ERp18 acquires scavenger activity for hydrogen peroxide (H2O2) in the ER. Depletion of ERp18 thus causes the accumulation of H2O2, which is produced during the oxidative folding of nascent polypeptides in the ER. ERp18 knockdown in C. elegans without Prx4 and GPx7/8, both of which are also known to have H2O2 scavenging activity in the ER, shortened the lifespan, suggesting that ERp18 may form a primitive and essential H2O2 scavenging system for the maintenance of redox homeostasis in the ER.

The oxidative folding of nascent polypeptides provides electrons for reductive reactions in the ER.

The endoplasmic reticulum (ER) maintains an oxidative redox environment that is advantageous for the oxidative folding of nascent polypeptides entering the ER. Reductive reactions within the ER are also crucial for maintaining ER homeostasis. However, the mechanism by which electrons are supplied for the reductase activity within the ER remains unknown. Here, we identify ER oxidoreductin-1α (Ero1α) as an electron donor for ERdj5, an ER-resident disulfide reductase. During oxidative folding, Ero1α catalyzes disulfide formation in nascent polypeptides through protein disulfide isomerase (PDI) and then transfers the electrons to molecular oxygen via flavin adenine dinucleotide (FAD), ultimately yielding hydrogen peroxide (H2O2). Besides this canonical electron pathway, we reveal that ERdj5 accepts electrons from specific cysteine pairs in Ero1α, demonstrating that the oxidative folding of nascent polypeptides provides electrons for reductive reactions in the ER. Moreover, this electron transfer pathway also contributes to maintaining ER homeostasis by reducing H2O2 production in the ER.

Redox-assisted regulation of Ca2+ homeostasis in the endoplasmic reticulum by disulfide reductase ERdj5.

Calcium ion (Ca2+) is an important second messenger that regulates numerous cellular functions. Intracellular Ca2+ concentration ([Ca2+]i) is strictly controlled by Ca2+ channels and pumps on the endoplasmic reticulum (ER) and plasma membranes. The ER calcium pump, sarco/endoplasmic reticulum calcium ATPase (SERCA), imports Ca2+ from the cytosol into the ER in an ATPase activity-dependent manner. The activity of SERCA2b, the ubiquitous isoform of SERCA, is negatively regulated by disulfide bond formation between two luminal cysteines. Here, we show that ERdj5, a mammalian ER disulfide reductase, which we reported to be involved in the ER-associated degradation of misfolded proteins, activates the pump function of SERCA2b by reducing its luminal disulfide bond. Notably, ERdj5 activated SERCA2b at a lower ER luminal [Ca2+] ([Ca2+]ER), whereas a higher [Ca2+]ER induced ERdj5 to form oligomers that were no longer able to interact with the pump, suggesting [Ca2+]ER-dependent regulation. Binding Ig protein, an ER-resident molecular chaperone, exerted a regulatory role in the oligomerization by binding to the J domain of ERdj5. These results identify ERdj5 as one of the master regulators of ER calcium homeostasis and thus shed light on the importance of cross talk among redox, Ca2+, and protein homeostasis in the ER.