Integrating Proteome-Wide Association Studies and Single-Cell Transcriptomics Identifies GSTT2B as a Causal Mediator and Prioritizes COL4A1 in Diabetic Retinopathy

Diabetic retinopathy (DR) is a leading cause of vision loss, yet its systemic proteomic mediators remain largely elusive. This study aimed to identify causal plasma proteins, map their cell-type-specific localization in the retina, and experimentally validate their expression under disease-relevant stress. We conducted a proteome-wide association study (PWAS) integrating UK Biobank plasma pQTL data (N = 53,022) with DR GWAS summary statistics. Causal relationships were inferred utilizing summary-data-based Mendelian randomization (SMR) and Bayesian colocalization. Prioritized candidates were mapped to the Human and Mouse Retina Cell Atlases via single-nucleus RNA sequencing (snRNA-seq). Finally, to substantiate the computational findings, in vitro validation of COL4A1 was performed in ARPE-19 cells cultured under hyperglycemic conditions utilizing quantitative real-time PCR (qPCR) and transcriptomic dataset re-analysis. The PWAS identified 26 proteins significantly associated with DR. Subsequent causal inference prioritized 12 high-confidence candidates, including GSTT2B, COL4A1, PAM, and GALNT3. Notably, GSTT2B emerged as a Tier-1 protective causal protein (Z = −3.609; PSMR = 1.22 × 10−4). snRNA-seq mapping revealed that GSTT2B is robustly expressed in Müller glia and the retinal pigment epithelium (RPE), whereas COL4A1 is prominently enriched in vascular compartments. These specific expression signatures exhibited partial conservation across species with notable cell-type specific variations. Crucially, in vitro validation confirmed that COL4A1 mRNA expression is significantly upregulated under high-glucose stress. Furthermore, druggability analysis highlighted actionable targets, identifying GSTT2B as a highly probable causal mediator and COL4A1 as a prioritized candidate for structural intervention. This study provides robust genetic, single-cell, and experimental evidence implicating specific plasma proteins in DR pathogenesis. The identification of GSTT2B-mediated protective pathways and the hyperglycemia-induced upregulation of COL4A1 offer a high-resolution molecular atlas to guide drug repositioning and precision therapeutic strategies.