Discussion

In this study, we integrated compartment-specific transcriptomic analysis, interaction network analysis, and SMR-based genetic inference to investigate the potential involvement of FUNDC1-associated mitochondrial regulatory networks in diabetic nephropathy (DN). Rather than establishing FUNDC1 as a direct causal driver, our analysis identified EHHADH as the key candidate gene within the FUNDC1-related network. EHHADH exhibited consistent downregulation across distinct renal compartments and was further supported by genetic association analysis, suggesting that altered EHHADH-associated metabolic regulation may represent a shared molecular feature associated with DN progression.

Mitochondrial dysfunction is increasingly recognized as a central pathological mechanism underlying DN development. Hyperglycemia-induced metabolic stress, excessive reactive oxygen species generation, and impaired mitochondrial quality control collectively contribute to progressive renal injury [13]. However, DN is not a homogeneous disease restricted to a single renal compartment. Increasing evidence indicates that disruption of the physiological interaction between glomerular and tubular compartments plays a critical role in disease progression. Podocyte injury can initiate downstream tubular metabolic stress, whereas tubular dysfunction further accelerates glomerular deterioration through inflammatory and metabolic feedback loops [14]. Therefore, identifying molecular regulators shared across different renal compartments may provide new insights into the coordinated progression of DN.

FUNDC1, an outer mitochondrial membrane mitophagy receptor, has been extensively implicated in mitochondrial quality control under metabolic and hypoxic stress conditions. Previous studies suggested that impaired FUNDC1-mediated mitophagy contributes to renal injury in diabetic models, partly through regulation of oxidative stress and mitochondrial apoptosis [15]. Consistent with these findings, FUNDC1-associated genes identified in our study were significantly enriched in mitochondrial organization, mitophagy, and energy metabolism pathways. However, our SMR analysis did not provide direct genetic evidence supporting FUNDC1 itself as a causal gene for DN. Therefore, FUNDC1 should be interpreted as an upstream molecular framework that guided the identification of mitochondrial regulatory candidates rather than as a genetically validated therapeutic target.

Among the FUNDC1-associated genes, EHHADH emerged as the primary candidate supported by both transcriptomic and genetic evidence. EHHADH encodes an enzyme involved in peroxisomal fatty acid β-oxidation and plays an essential role in maintaining lipid metabolic homeostasis in renal tubular epithelial cells. Previous experimental studies have suggested that EHHADH deficiency aggravates tubulointerstitial injury in diabetic kidney disease by disrupting peroxisomal function and promoting abnormal pexophagy, ultimately enhancing oxidative stress and inflammatory responses [16]. Our findings extend these observations by showing that reduced EHHADH expression is observed at the transcriptomic level and is further supported by SMR-based genetic association analysis.

The consistent reduction of EHHADH across glomerular and tubular compartments provides additional biological implications. Although renal compartments experience distinct pathological stresses during DN progression, including podocyte dysfunction in glomeruli and metabolic overload in tubular epithelial cells, both compartments are highly dependent on mitochondrial energy metabolism. The cross-compartment reduction of EHHADH suggests that impaired lipid metabolic regulation may represent a common molecular response during DN progression [17]. Importantly, while the biological functions of EHHADH have been primarily characterized in tubular cells, its downregulation in glomerular tissues indicates that EHHADH-associated metabolic disturbance may extend beyond tubular injury and participate in broader renal homeostasis disruption.

From a translational perspective, the SMR evidence obtained from whole-blood eQTL data provides additional significance. Although blood-derived genetic instruments cannot directly represent kidney-specific regulation, the identification of EHHADH as genetically associated with DN susceptibility suggests that its regulatory variation may reflect systemic biological processes linked to disease risk. Compared with purely tissue-level observational changes, genetic evidence is less susceptible to reverse causality and disease-induced secondary alterations [18]. Therefore, EHHADH may represent a potential molecular candidate associated with DN susceptibility., although kidney-specific eQTL and clinical cohort evaluation will be required to confirm its applicability.

Mechanistically, our pathway analyses indicated that FUNDC1 and EHHADH converge on mitochondrial metabolic regulation, suggesting a potential biological association between mitochondrial quality control and lipid metabolic regulation. EHHADH may primarily maintain metabolic substrate utilization through fatty acid oxidation, whereas FUNDC1 contributes to mitochondrial quality control through selective removal of damaged mitochondria. Similar coordinated regulation between different mitochondrial quality control processes has been reported previously, supporting the possibility that metabolic maintenance and mitochondrial clearance pathways may act cooperatively under stress conditions [19]. However, whether FUNDC1 directly regulates EHHADH expression or whether both genes represent parallel responses to diabetic stress remains unknown and requires experimental investigation.

Furthermore, transcription factor analysis identified 17 shared transcription factors potentially involved in the regulation of FUNDC1 and EHHADH. Among these candidates, E2F4 was prioritized based on integrated enrichment evidence and biological relevance. As a member of the E2F transcription factor family, E2F4 has been implicated in cellular stress responses, metabolic regulation, and renal pathological processes, including clear cell renal carcinoma [20,21]. Although diabetic nephropathy and renal cancer represent distinct diseases, they share certain pathological features, such as oxidative stress, inflammatory activation, and metabolic reprogramming. However, the current analysis does not establish a dominant role for E2F4 over other shared transcription factors; therefore, E2F4 should be considered a computationally prioritized candidate requiring further experimental validation.

Several limitations should be acknowledged. First, although SMR provides genetic evidence supporting an association between EHHADH expression and diabetic nephropathy risk, the eQTL dataset used in this study was derived from whole blood rather than kidney tissue. Therefore, kidney-specific regulatory effects require further confirmation using renal eQTL resources. Second, the present study was primarily based on transcriptomic integration and computational analyses, and experimental validation of the FUNDC1–EHHADH relationship, including functional perturbation, protein interaction, and histological confirmation, was not performed. Third, although cross-compartment transcriptomic analysis showed a consistent downregulation pattern of EHHADH, independent external cohorts with larger sample sizes are needed to further evaluate the robustness of these findings. Finally, the predicted transcriptional regulation by E2F4 and the proposed coordination between mitophagy and metabolic pathways remain hypothesis-generating and require further mechanistic studies.

Overall, this study identifies EHHADH as a genetically supported candidate gene within a FUNDC1-associated mitochondrial regulatory network in DN. These findings highlight the importance of metabolic-mitochondrial coupling in coordinating renal compartment injury and provide a potential molecular basis for future biomarker development and therapeutic exploration.