La perdita di Acquaporina11 impedisce la corretta biogenesi di Policistina-1 attraverso l'accumulo di H2O2 nel Reticolo Endoplasmatico

Aquaporin11 (AQP11) is an ER-resident channel that allows passive transport of H2O2 from the ER-lumen to the cytosol, along a concentration gradient. Removing the channel, the flux of H2O2 that goes from one compartment to the other is blocked. As a consequence, cells with reduced expression of AQP11 display increased concentration of H2O2 in the ER (H2O2)ER and decreased H2O2 in the cytosol (H2O2)cytosol, leading to ER hyper-oxidation and possibly impacting cytosolic redox signalling. Aqp11-/- mice develop an early onset Polycystic Kidney Disease (PKD) and die of renal failure within three weeks of life. The pathological mechanism proposed was the impaired maturation of Polycystin-1 (PC-1), the product of PKD1 gene which is mutated in over four fifths of Autosomal Dominant Polycystic Kidney Disease (ADPKD) cases. During its biogenesis, PC-1 undergoes many posttranslational modifications in the early secretory pathway, including N-glycosylation, formation of disulfide bonds, autoproteolytic cleavage and assembly with its partner Polycystin-2 (PC-2) to generate functional polycystin complexes. This complex biogenesis is a hard task for cells: mutations in many enzymes involved in N-glycan assembly, protein folding, and quality control were found to affect PC-1 biogenesis and cause atypical PKD in humans. Thus, PC-1 processing seems particularly sensitive to perturbations in the folding machinery, and many conditions that are tolerated by other proteins prevent polycystin to reach its functional form. During my PhD studies, I sought to clarify how the absence of functional AQP11 causes PKD. Are the altered H2O2 fluxes ensuing from lack of AQP11 the cause of impaired polycystin maturation? To address these issues, I investigated whether and how perturbing H2O2 level and redox homeostasis impacted polycystin. My results provide new insights on how downregulation of AQP11 selectively inhibits PC-1 processing. Using tricks to manipulate (H2O2)ER independently by AQP11, I was able to demonstrate that the defect on PC-1 is manifested via accumulation of (H2O2)ER. In addition, I found that loss of AQP11 results in defective N-glycan processing that affects PC-1 as well as other glycoproteins. My results confirm the fundamental role of AQP11 as a H2O2 transporter across the ER membrane, and demonstrate that subtle changes in the (H2O2)ER have dramatic consequences in PC- 1 biogenesis with broad effects on glycoprotein processing.

Questa tesi investiga il ruolo di Acquaporina11 (AQP11), un canale per l'acqua ossigenata (H2O2) localizzato sulla membrana del reticolo endoplasmatico, nella corretta maturazione di Policistina-1 (PC-1), proteina la cui funzionalità è necessaria per prevenire la formazioni di cisti renali.