Divergent pathways of Surfactant Protein C maturation for disease-associated isoforms

Authors

• Sarah Bui, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104; PENN-CHOP Lung Biology Institute; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Anamarie Reineberg, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Dakota Jones, Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA; The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA.
• Cheng-Lun Na, Perinatal Institute and Division of Neonatology, Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center. Cincinnati OH 45221 USA.
• Joseph Kitzmiller, Perinatal Institute and Division of Neonatology, Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center. Cincinnati OH 45221 USA.
• Luis R. Rodriguez, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104; PENN-CHOP Lung Biology Institute; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104; Department of Biochemistry and Molecular Medicine, The George Washington University, Washington, D.C., 20037, USA.
• Aditi Murthy, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104; PENN-CHOP Lung Biology Institute; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Swati Iyer, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Charlotte Cooper, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Rea Chroneos, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Yaniv Tomer, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Surafel Mulugeta, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104.
• Timothy E. Weaver, Perinatal Institute and Division of Neonatology, Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center. Cincinnati OH 45221 USA.
• Darrell N. Kotton, Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA; The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA.
• Konstantinos-Dionysios Alysandratos, Center for Regenerative Medicine, Boston University and Boston Medical Center, Boston, MA 02118, USA; The Pulmonary Center and Department of Medicine, Boston University Chobanian & Avedisian School of Medicine, Boston, MA 02118, USA.
• Jeffrey A. Whitsett, Perinatal Institute and Division of Neonatology, Perinatal and Pulmonary Biology, Cincinnati Children's Hospital Medical Center. Cincinnati OH 45221 USA.
• Michael F. Beers, Pulmonary and Critical Care Division, Department of Medicine; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104; PENN-CHOP Lung Biology Institute; Perelman School of Medicine at the University of Pennsylvania; Philadelphia, PA 19104. Electronic address: mfbeers@pennmedicine.upenn.edu.

Document Type

Journal Article

Publication Date

2-5-2026

Journal

The Journal of biological chemistry

DOI

10.1016/j.jbc.2026.111252

Keywords

Alveolar Type 2 Cell; Golgi; Proprotein Convertase; Protein Maturation; Surfactant Protein C

Abstract

Surfactant Protein C (SP-C), a hydrophobic protein exclusively synthesized and secreted by alveolar type II (AT2) cells, is important for reducing alveolar surface tension in the distal lung. Chronic interstitial pulmonary diseases have been associated with SFTPC mutations. However, a detailed understanding of SP-C maturation in the secretory pathway and disruptions caused by mutations has remained incomplete. The goal of this study was to comprehensively ascertain differences in trafficking and post-translational processing between wild-type and disease-associated SP-C mutants using doxycycline-inducible mouse lung epithelial (MLE-12) cell lines expressing either wildtype SP-C or the common clinical variant SP-C, validated using primary AT2 cells isolated from a murine SP-C pulmonary fibrosis model and induced pluripotent stem cell (iPSC)-derived human AT2 cells expressing the same mutant. In all 3 models SP-C was highly concentrated in acidic Lysosomal Related Organelles (LROs) while SP-C accumulated on the plasma membrane, which was corroborated by inhibition of clathrin-mediated endocytosis, surface biotinylation, immunogold EM, immunofluorescent staining, and proteinase K protection assays supporting divergence of SP-C trafficking from SP-C. The exclusion of SP-C from normal routing occurred early in the biosynthetic pathway as Brefeldin A blocked processing of both SP-C proproteins, while a 20˚C temperature shift caused selective accumulation of a processed proSP-C intermediate, suggesting initial C-terminal cleavage of proSP-C occurs in late-Golgi/trans-Golgi network (TGN). This cleavage event was sensitive to DC1, an inhibitor of furin-related subtilisin-like proprotein convertase (PPC) family members. Site-directed mutagenesis of canonical residues K160/R167 within a predicted PPC recognition site in the proSP-C COOH domain blocked its processing. Expression constructs encoding inhibitory pre-proprotein (pp) peptide fragments of Furin and ppPC7 each inhibited cleavage of proSP-C by MLE-12 cells. Collectively, our data demonstrate that trafficking pathways for maturation of WT and mutant I73T SP-C diverge prior to the TGN where initial cleavage of the COOH-terminal SP-C propeptide occurs via a Furin-like proprotein convertase.

APA Citation

Bui, Sarah; Reineberg, Anamarie; Jones, Dakota; Na, Cheng-Lun; Kitzmiller, Joseph; Rodriguez, Luis R.; Murthy, Aditi; Iyer, Swati; Cooper, Charlotte; Chroneos, Rea; Tomer, Yaniv; Mulugeta, Surafel; Weaver, Timothy E.; Kotton, Darrell N.; Alysandratos, Konstantinos-Dionysios; Whitsett, Jeffrey A.; and Beers, Michael F., "Divergent pathways of Surfactant Protein C maturation for disease-associated isoforms" (2026). GW Authored Works. Paper 8746.
https://hsrc.himmelfarb.gwu.edu/gwhpubs/8746

Department

Biochemistry and Molecular Medicine