Abstract Body: Fabry disease is an X-linked lysosomal storage disorder caused by pathogenic mutations in the GLA gene, resulting in deficient α-galactosidase A (α-gal A) activity and progressive accumulation of globotriaosylceramide (Gb3) and its deacylated metabolite lyso-Gb3 in multiple cell types and organs. At the cellular level, lyso-Gb3 acts not only as a storage metabolite but also as a bioactive signaling molecule that provokes chronic inflammation, proteotoxic stress, mitochondrial dysfunction, impaired autophagic flux, and premature cellular senescence, while at the tissue level it drives progressive vascular and renal pathology. The C-terminus of Hsc70-interacting protein (CHIP), a stress-responsive ubiquitin E3 ligase that integrates molecular chaperone activity with proteostasis and organelle quality control, has been implicated in diverse protein misfolding diseases; however, its contribution to lyso-Gb3–driven Fabry pathology has remained largely unexplored. In this study, we demonstrate that CHIP serves as a critical protective regulator against lyso-Gb3–induced cellular and tissue dysfunction using complementary in vitro and in vivo models. Exposure of wild-type fibroblasts to lyso-Gb3 led to a dose-dependent reduction in CHIP expression, while fibroblasts harboring GLA mutations (p.R301G/o and p.R301Q/o) exhibited marked suppression of CHIP levels accompanied by diminished α-gal A activity and excessive lyso-Gb3 accumulation. These pathological conditions were associated with increased A11-positive toxic protein oligomers and ubiquitin-conjugated aggregates. CHIP restoration enhanced α-gal A activity, reduced lyso-Gb3 accumulation, and broadly mitigated inflammation, necroptosis, autophagy–lysosome dysfunction, mitochondrial respiratory defects, and cellular senescence. In a murine model subjected to sustained lyso-Gb3 administration, CHIP haploinsufficiency exacerbated vascular relaxation defects and significantly elevated blood urea nitrogen, creatinine, and uric acid levels, indicating aggravated renal functional impairment. Histopathological analysis revealed characteristic Fabry-associated renal lesions. Collectively, these findings identify CHIP as a central node linking α-gal A activity, proteostasis, and organelle quality control to the suppression of lyso-Gb3–driven Fabry disease pathology, thereby highlighting CHIP as a promising therapeutic target for mitigating both cellular dysfunction and progressive organ damage in Fabry disease.