Processing of pulmonary surfactant protein B by napsin and cathepsin H.
Surfactant protein B (SP-B) is an essential constituent of pulmonary surfactant. SP-B is synthesized in alveolar type II cells as a preproprotein and processed to the mature peptide by the cleavage of NH2- and COOH-terminal peptides. An aspartyl protease has been suggested to cleave the NH2-terminal propeptide resulting in a 25-kDa intermediate. Napsin, an aspartyl protease expressed in alveolar type II cells, was detected in fetal lung homogenates as early as day 16 of gestation, 1 day before the onset of SP-B expression and processing. Napsin was localized to multivesicular bodies, the site of SP-B proprotein processing in type II cells. Incubation of SP-B proprotein from type II cells with a crude membrane extract from napsin-transfected cells resulted in enhanced levels of a 25-kDa intermediate. Purified napsin cleaved a recombinant SP-B/EGFP fusion protein within the NH2-terminal propeptide between Leu178 and Pro179, 22 amino acids upstream of the NH2 terminus of mature SP-B. Cathepsin H, a cysteine protease also implicated in pro-SP-B processing, cleaved SP-B/EGFP fusion protein 13 amino acids upstream of the NH2 terminus of mature SP-B. Napsin did not cleave the COOH-terminal peptide, whereas cathepsin H cleaved the boundary between mature SP-B and the COOH-terminal peptide and at several other sites within the COOH-terminal peptide. Knockdown of napsin by small interfering RNA resulted in decreased levels of mature SP-B and mature SP-C in type II cells. These results suggest that napsin, cathepsin H, and at least one other enzyme are involved in maturation of the biologically active SP-B peptide.
Ueno
Takayuki
T
Cancer Center Karolinska, Department of Oncology-Pathology, Karolinska Institute and Hospital, SE-171 76 Stockholm, Sweden.
Linder
Stig
S
Na
Cheng-Lun
CL
Rice
Ward R
WR
Johansson
Jan
J
Weaver
Timothy E
TE
D000818:Animals; D016282:Aspartic Acid Endopeptidases / Q000378:metabolism*; D056655:Cathepsin H; D002403:Cathepsins / Q000378:metabolism*; D002478:Cells, Cultured; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000378:metabolism / Q000648:ultrastructure; D008168:Lung / Q000378:metabolism / Q000648:ultrastructure; D051379:Mice; D011499:Protein Processing, Post-Translational; D037701:Pulmonary Surfactant-Associated Protein B / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism
D037701:Pulmonary Surfactant-Associated Protein B; D011993:Recombinant Fusion Proteins; D002403:Cathepsins; D003546:Cysteine Endopeptidases; C534126:CTSH protein, human; D056655:Cathepsin H; C534127:Ctsh protein, mouse; D016282:Aspartic Acid Endopeptidases; C104073:Kdap protein, mouse