Protein disulfide isomerase interacts with soluble guanylyl cyclase via a redox-based mechanism and modulates its activity.

Nitric oxide (NO) binds to the receptor, soluble guanylyl cyclase (sGC), stimulating cGMP production. The NO-sGC-cGMP pathway is a key component in the cardiovascular system. Discrepancies in sGC activation and deactivation in vitro vs. in vivo have led to a search for endogenous factors that regulate sGC or assist in cellular localization. In our previous work, which identified Hsp70 as a modulator of sGC, we determined that protein disulfide isomerase (PDI) bound to an sGC-affinity matrix. We here establish and characterize this interaction. Incubation of purified PDI with semi-purified sGC, both reduced and oxidized, resulted in different migration patterns on non-reducing Western blots indicating a redox component to the interaction. In sGC-infected COS-7 cells, transfected Flag-tagged PDI and PDI-CXXS (redox active site “trap-mutant”) pulled down sGC. This PDI-sGC complex was resolved by reductant, confirming a redox interaction. PDI inhibited NO-stimulated sGC activity in COS-7 lysates; however, a PDI redox inactive mutant, PDI-SXXS, did not. Together, these data unveil a novel mechanism of sGC redox modulation via thiol-disulfide exchange. Finally, in smooth muscle cells, endogenous PDI and sGC co-localize by in situ proximity ligation assay, suggesting biological relevance. PDI-dependent redox regulation of sGC NO-sensitivity may provide a secondary control over vascular homeostasis.
Source: BJ Cell - Category: Biochemistry Authors: Tags: BJ Signal Source Type: research