Disrupting the physical interaction between serotonin transporter and soluble guanylate cyclase produces a fast-acting antidepressant activity

5-HT signaling modulates multiple mental states and is strictly controlled by serotonin transporter (SERT). The conventional antidepressants, selective serotonin reuptake inhibitors (SSRIs) that directly inhibit SERT have many limitations in the treatment of major depressive disorder (MDD). In the present study, we identified a protein-protein complex formed by the physical interaction between SERT and soluble guanylate cyclase (sGC). Augmentation of the SERT-sGC interaction decreased SERT cell surface expression by interfering with its trafficking, thereby reducing 5-HT uptake. The specific SERT-sGC association was mediated by the forth internal loop (IL4) motif with a unique structural folding in SERT and exerted a unilateral modulation of SERT but not sGC. Notably, the SERT-sGC association was responsive to treatment of a variety of susceptibility factors in both cells and animal models. Disrupting the SERT-sGC interaction in the dorsal raphe nucleus (DRN) by the IL4 peptide rapidly normalized 5-HT transmission and produced a rapid-onset antidepressant activity. The study not only revealed the molecular mechanism by which SERT is under the regulatory control by the SERT-sGC association, but also provided insights into a novel target against the SERT-sGC interaction in developing fast-acting agents in the treatment of 5-HT-related psychiatric disorders. • We identified an IL4 motif-mediated physical interaction between serotonin transporter (SERT) and soluble guanylate cyclase (sGC). • SERT-sGC association reduces SERT surface expression by impairing its trafficking. • Chronic stress enhances SERT-sGC interaction in the dorsal raphe nucleus, leading to depressive behaviors. • Disrupting SERT-sGC interaction rapidly normalizes 5-HT transmission and produces fast-onset antidepressant effects.