Ca ²⁺ -driven PDIA6 phase separation to ensure proinsulin quality control

The endoplasmic reticulum (ER) plays key roles in protein quality control ^1,2 and dynamic Ca ²⁺ storage ^3,4 in eukaryotic cells. However, the protein homeostasis (proteostasis) system that regulates these ER functions is still incompletely characterised. Previous study revealed the importance of oligomerization in the function PDIA1, an ER-resident disulfide isomerase and molecular chaperone, regulates oligomeric states in accordance with client folding ⁵ . This result suggests that at least some of the 20 members of other PDI family may undergo regulated self-assembly in order to optimally function. Here, we show that Ca ²⁺ triggers the phase separation of PDIA6 into liquid-like condensates. In contrast to the condensation mechanism observed for proteins containing low-complexity domains, our results indicate that transient but specific electrostatic interactions occur between the first and the third folded thioredoxin-like domains of PDIA6. We further show that the Ca ²⁺ -driven condensation of PDIA6 recruits PDIA3 and proinsulin, thus increasing their local concentrations. This process results in the 30-fold enhancement of proinsulin folding and in the inhibition of proinsulin aggregation. Our findings shed light on a condensation-driven Ca ²⁺ -mediated proteostasis cascade in the ER by revealing how the efficiency of the protein folding process can be enhanced within quality control granules.