Mutation of Serine 32 to Threonine in Peroxiredoxin 6 Preserves Its Structure and Enzymatic Function but Abolishes Its Trafficking to Lamellar Bodies

Peroxiredoxin 6 (Prdx6), a bifunctional protein with phospholipase A2 (aiPLA2) and GSH peroxidase activities, protects lungs from oxidative stress and participates in lung surfactant phospholipid turnover. Prdx6 has been localized to both cytosol and lamellar bodies (LB) in lung epithelium, and its organellar targeting sequence has been identified. We propose that Prdx6 LB targeting facilitates its role in the metabolism of lung surfactant phosphatidylcholine (PC). Ser-32 has been identified as the active site in Prdx6 for aiPLA2 activity, and this activity was abolished by the mutation of serine 32 to alanine (S32A). However, aiPLA2 activity was unaffected by mutation of serine 32 in Prdx6 to threonine (S32T). Prdx6 protein expression and aiPLA2 activity were normal in the whole lung of a "knock-in" mouse model carrying an S32T mutation in the Prdx6 gene but were absent from isolated LB. Analyses by proximity ligation assay in lung sections demonstrated the inability of S32T Prdx6 to bind to the chaperone protein, 14-3-3ϵ, that is required for LB targeting. The content of total phospholipid, PC, and disaturated PC in lung tissue homogenate, bronchoalveolar lavage fluid, and lung LB was increased significantly in Prdx6-S32T mutant lungs, whereas degradation of internalized [(3)H]dipalmitoyl-PC was significantly decreased. Thus, Thr can substitute for Ser for the enzymatic activities of Prdx6 but not for its targeting to LB. These results confirm an important role for LB Prdx6 in the degradation and remodeling of lung surfactant phosphatidylcholine.