Protein S-sulfhydration: an emerging regulatory layer with therapeutic potential in metabolic dysfunction-associated steatotic liver disease

Metabolic dysfunction-associated steatotic liver disease (MASLD) has emerged as the most prevalent chronic liver disorder worldwide and represents a key hepatic manifestation of systemic metabolic imbalance (1-3). Driven by excess energy intake, adipocyte dysfunction, and insulin resistance, the prevalence of MASLD continues to rise in parallel with the growing burden of metabolic syndrome, posing a substantial challenge to public health systems (4,5). Characterized by excessive hepatic lipid accumulation, MASLD encompasses a broad disease spectrum ranging from simple steatosis to steatohepatitis, fibrosis, and cirrhosis (6,7). The molecular mechanisms that coordinate hepatic lipid metabolism under fluctuating metabolic and redox conditions remain incompletely understood. In particular, regulatory layers beyond transcriptional control and metabolite availability, including post-translational and redox-sensitive mechanisms, are increasingly appreciated as critical determinants of disease susceptibility and progression (8).

Post-translational modifications (PTMs) provide rapid and reversible means to fine-tune protein function in response to cellular cues, thereby enabling dynamic adaptation of cellular signaling networks to metabolic and environmental changes. Among these, protein S-sulfhydration—a cysteine-based modification mediated by hydrogen sulfide (H₂S)—has recently gained attention as a redox-sensitive regulatory mechanism with broad implications for metabolic homeostasis (9,10). Unlike many irreversible oxidative modifications, S-sulfhydration is reversible and context-dependent, allowing it to act as a finely tunable regulatory switch in redox signaling. Accumulating evidence indicates that protein S-sulfhydration modulates enzyme activity, protein-protein interactions, subcellular localization, as well as transcriptional and signaling functions, thereby exerting pleiotropic effects on cellular metabolism (11,12). As the biochemical donor of this modification, H₂S is endogenously generated primarily through conserved enzymatic pathways involving cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE), and 3-mercaptopyruvate sulfurtransferase (3-MST), which together constitute the major enzymatic sources of H₂S in mammalian cells (13). These enzymes exhibit tissue-specific expression patterns and metabolic regulation, thereby linking sulfur amino acid metabolism to cellular redox status. Importantly, all three enzymes are expressed in the liver, reflecting its central role in systemic H₂S homeostasis. As a major hub for sulfur amino acid metabolism, the liver possesses substantial capacity for H₂S production and turnover, enabling precise spatial and temporal control of protein S-sulfhydration. Accordingly, the liver serves not only as a target of H₂S signaling but also as a key regulatory organ in systemic redox homeostasis. Dysregulation of hepatic H₂S metabolism may therefore contribute to metabolic disorders characterized by redox imbalance, including MASLD.

Against this background, emerging evidence identifies disruption of H₂S-mediated protein S-sulfhydration as a key pathogenic mechanism linking redox imbalance to hepatic lipid metabolic dysregulation in MASLD. In the liver, H₂S signaling suppresses de novo lipogenesis through S-sulfhydration of metabolic and transcriptional regulators such as sirtuin 1 (SIRT1) and the nuclear receptor farnesoid X receptor (FXR), leading to inhibition of SREBP-1-driven lipogenic gene expression and attenuation of excessive fatty acid synthesis (14,15). This regulatory axis integrates redox sensing with transcriptional control of lipid metabolism, thereby constraining metabolic overactivation under physiological conditions. In contrast, loss of H₂S bioavailability derepresses lipogenic programs, enhances the activity of canonical lipogenic enzymes, and accelerates hepatic lipid accumulation, thereby promoting steatotic transformation. In parallel, H₂S-mediated protein S-sulfhydration facilitates hepatic lipid clearance by sustaining autophagic flux and lipophagy, processes essential for lipid droplet turnover and intracellular lipid quality control. Mechanistically, reduced H₂S availability impairs ULK1 activation, disrupts autophagosome formation, and compromises lipid droplet degradation, ultimately exacerbating steatotic injury under metabolic stress (16). This highlights autophagy as a key redox-sensitive effector pathway downstream of H₂S signaling in hepatic lipid homeostasis. Moreover, disruption of H₂S signaling further perturbs hepatic cholesterol homeostasis in metabolic disease states by attenuating Keap1-Nrf2 signaling and downstream LRP1 expression, thereby limiting hepatic clearance of cholesterol-rich lipoproteins and aggravating hepatic lipid burden (9). Collectively, H₂S-dependent protein S-sulfhydration functions as an integrative redox-sensitive regulatory network that coordinates lipid synthesis, degradation, and cholesterol handling. By coupling sulfur metabolism to multiple layers of hepatic lipid regulation, this pathway critically shapes hepatic lipid homeostasis, disease susceptibility, and the progression of MASLD, and represents a potential therapeutic target.

Beyond hepatic contexts, protein S-sulfhydration has emerged as a conserved redox-sensitive mechanism regulating lipid metabolic processes across extrahepatic tissues. Although direct evidence in the liver remains limited, accumulating studies in metabolic tissues provide compelling proof-of-concept that protein S-sulfhydration can fine-tune distinct steps of lipid handling in a highly context-dependent manner. In the heart, S-sulfhydration limits fatty acid uptake by modulating vesicle trafficking-dependent membrane localization of the predominant fatty acid transporter CD36, thereby restraining lipid influx and attenuating cardiomyocyte lipotoxic stress (10). Given the evolutionary conservation of vesicular trafficking machinery and lipid transport systems, similar redox-sensitive regulatory mechanisms may operate in hepatocytes to dynamically control fatty acid uptake under metabolic stress. Likewise, protein S-sulfhydration has emerged as an important regulator of mitochondrial fatty acid oxidation (FAO), as impaired S-sulfhydration of key β-oxidation enzymes is associated with defective FAO, mitochondrial dysfunction, and heightened metabolic stress in diabetic kidney disease (17). These observations highlight a critical role for sulfur-dependent redox modifications in maintaining mitochondrial metabolic competence across tissues with high energetic demand. In parallel, studies in cardiomyocytes demonstrate that S-sulfhydration restrains triglyceride synthesis by promoting the turnover of key triglyceride-synthesizing enzymes, thereby limiting lipid droplet expansion and preventing excessive intracellular lipid storage (18). Together, the conservation of these regulatory principles across metabolically active tissues supports the concept that protein S-sulfhydration functions as a fundamental redox-sensitive mechanism coordinating lipid uptake, utilization, and storage. Extrapolated to the liver, these findings suggest that analogous S-sulfhydration-dependent pathways may critically influence hepatic lipid homeostasis, and that their disruption could contribute to the multifaceted metabolic disturbances underlying MASLD pathogenesis.

From a translational perspective, therapeutic strategies aimed at restoring H₂S-mediated protein S-sulfhydration have attracted increasing attention as potential interventions for lipid metabolic disorders, particularly MASLD. Exogenous H₂S donors are the most extensively explored pharmacological tools, yet they differ markedly in kinetics, bioavailability, tissue distribution, and safety. Fast-releasing sulfide salts such as sodium hydrosulfide (NaHS) enable acute S-sulfhydration modulation and are useful for proof-of-concept studies; however, their rapid H₂S burst often leads to supraphysiological exposure and dose-dependent toxicity, limiting translational applicability (19). In contrast, slow-releasing donors such as GYY4137 provide sustained and more physiologically relevant H₂S delivery, resulting in more stable modulation of redox signaling and conferring metabolic benefits in preclinical models of metabolic dysfunction (20). More recently, mitochondria-targeted H₂S donors have further expanded the therapeutic landscape by enabling spatially restricted enhancement of S-sulfhydration within key metabolic organelles, thereby improving efficacy while reducing off-target effects (21).

To date, clinical evaluation of H₂S donors has been limited to early-phase studies of compounds such as SG1002 and ATB-346, which have demonstrated modest anti-inflammatory or cytoprotective effects in cardiovascular and inflammatory settings (22,23). However, these studies have not assessed metabolic endpoints, and no clinical trials have specifically investigated their efficacy in MASLD or other lipid metabolic diseases. Consequently, there remains a substantial gap between extensive mechanistic evidence supporting the role of H₂S-mediated S-sulfhydration in metabolic regulation and its clinical translation. This gap is further compounded by incomplete understanding of tissue-specific H₂S pharmacodynamics, variability in endogenous sulfur metabolism, and the lack of biomarkers to monitor S-sulfhydration in vivo, all of which hinder rational dose optimization and patient stratification. Despite these challenges, the convergence of preclinical metabolic benefits and early clinical safety data suggests that targeting H₂S signaling may represent a promising but still largely untapped therapeutic avenue for MASLD.

Collectively, protein S-sulfhydration emerges as a dynamic redox-sensitive regulatory layer that integrates lipid metabolic processes across synthesis, utilization, and storage, thereby maintaining hepatic lipid homeostasis under physiological conditions. In the context of MASLD, disruption of H₂S signaling leads to coordinated impairment of multiple metabolic nodes, including lipogenesis, FAO, autophagy-mediated lipid clearance, and cholesterol handling, ultimately promoting hepatic lipid overload and disease progression. Importantly, evidence from extrahepatic metabolic tissues further supports the concept that S-sulfhydration functions as a conserved mechanism linking redox state to lipid metabolic control. Although direct hepatic evidence remains limited, these findings collectively suggest that restoring physiological S-sulfhydration represents a promising strategy to re-establish metabolic balance in MASLD. Future studies are warranted to define liver-specific regulatory targets, improve spatial and temporal control of H₂S delivery, and translate these mechanistic insights into clinically viable therapeutic approaches.

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