The Golgi Apparatus-Hosted ATP7A-Fibulin-4 Complex Mediates Activation of LOX to Promote Kidney Fibrosis

Background: Lysyl oxidase (LOX) is a copper-dependent monoamine oxidase whose primary function is the covalent cross-linking of collagen and elastin in extracellular matrix (ECM), however, the regulation of LOX activity in renal fibrosis is not well understood. Methods: To explore the relationship between cellular copper overload and LOX activity, we employed copper chelator tetrathiomolybdate and implemented specific knockdown of copper transporter1 (CTR1) both in vivo and in vitro. Subsequently, we evaluated LOX activity and activated LOX expression by LOX assay kit and western blot. The specific regulatory mechanism of LOX activity was elucidated by knocking-down expression of ATPase copper transporting alpha (ATP7A) and Fibulin-4 (FBLN4) in vitro or in vivo. Results: 1) Expression of LOX and extracellular matrix (ECM) cross-linking were markedly increased in fibrotic kidneys of ischemia/reperfusion (IR) induced renal fibrosis model. Treatment with tetrathiomolybdate or the knockdown of CTR1 resulted in a reduction in copper levels, leading to decrease LOX activity and ameliorated renal fibrosis. 2) The upregulation of ATP7A expression increased copper levels in the Golgi apparatus of renal tubular epithelial cells, resulting in enhanced LOX activity and ECM crosslinking, thereby promoting the progression of renal fibrosis. 3) FBLN4 was essential for ATP7A-transfering copper to LOX and formed a ternary complex of ATP7A-FBLN4-LOX in renal tubular epithelial cells. Conclusion: Our results suggest that the ATP7A-FBLN4 complex facilitates the aberrant accmulation of copper in the Golgi apparatuses, thereby promoting the activation of LOX to catalyze ECM cross-linking and contribute to the progression of renal fibrosis. Funding: Government Support – Non-U.S.