Abstract 10706: Transnitrosation From the NO-Receptor Soluble Guanylyl Cyclase to Oxidized Thioredoxin-1 Has a Potential Cardiovascular Protective Effect Against Oxidative Stress

Abstract

Soluble guanylyl cyclase (GC1) is an α/β heterodimer producing cGMP when stimulated by nitric oxide (NO). The NO-GC1-cGMP pathway is essential for cardiovascular homeostasis. Oxidative stress is known to disrupt the NO-GC1-cGMP pathway via oxidation of GC1 cysteines, which makes it unresponsive to NO stimulation. Under these conditions, GC1-α subunit “moonlights” by increasing cellular S-nitrosation via specific transfer of its nitrosothiols to cysteines of other proteins (transnitrosation). One of the SNO-targets identified by Mass Spectrometry (MS) was the oxidized form of Thioredoxin1 (oTrx1), which is unidirectionally transnitrosated by GC1 with αC610 as a SNO-donor and C73 of Trx1 as the SNO-recipient. Because oTrx1 itself drives transnitrosation, we sought and MS-identified SNO-proteins targeted by both GC1 and Trx1, suggesting a SNO-GC1→oTrx1→targets cascade. We investigated the (patho)physiological relevance of GC1-dependent transnitrosation activity by creating a knock-in (KI) mice with replacement of αC610 to a serine (αC610S). Biotin switch assays confirm that, under oxidative stress (Angiotensin II (AngII) treatment), global and Trx1 levels of S-nitrosation (SNO) were drastically reduced in lungs and hearts of KI mice compared to WT but the NO-stimulated GC1 activity was similar between WT and KI. Importantly, telemetry recordings indicated that 2 weeks of AngII treatment led to a more severe increase in BP and a more prolonged QTc in KI mice. The AngII-induced rise in MBP was 27.6 ± 2.5 mmHg in KI vs 21.6 ± 1.2mmHg in WT, while AngII-increase in QTc, reflecting delayed repolarization, was 12.53 ± 2.30ms in KI vs. 8.713 ± 2.72ms for the WT. The elevated BP with AngII treatment correlated with higher increase in cardiac hypertrophy, oxidation (measured by DHE staining) and fibrosis (picric acid staining) in KI mice. The AngII-induced increase in a) hypertrophy in KI was 0.63 ± 0.07 vs. 0.24 ± 0.08 mg/g in WT, b) cardiac oxidation was 0.19 ± 0.07 in KI and negligible in WT, and c) fibrosis was 1.75 (A.U) in KI vs. 1.31 in WT. These more severe cardiovascular dysfunctions in KI mice in response to AngII suggest that transnitrosation by GC1, potentially using oTrx1 as a nitrosothiol relay, is a mechanism of protection against oxidative stress.