Surfactant protein C precursor is palmitoylated and associates with subcellular membranes.
Surfactant protein C (SP-C) is a 3.7 kDa, hydrophobic protein that enhances the adsorption of phospholipids in pulmonary surfactant. SP-C is generated by proteolytic processing of a 21 kDa precursor. Murine fetal lung explant cultures and a Chinese hamster ovary cell line expressing recombinant human SP-C gene (CHO/SPC) were used to determine the subcellular location and post-translational modification(s) of proSP-C. After in vitro translation, proSP-C of Mr = 21,000 was generated. ProSP-C was associated with canine pancreatic microsomes during co-translation and was partially protected from digestion with proteinase K, supporting the concept that proSP-C enters but does not completely traverse the membrane of the endoplasmic reticulum (ER). Association of proSP-C isoforms of 21 and 26 kDa with intracellular membranes was demonstrated by subcellular fractionation of CHO/SPC cells. Pulse/chase experiments demonstrated that the 21 kDa SP-C proprotein was synthesized first and after 15 min was modified to produce a 26 kDa isoform in CHO/SPC cells or a 24 kDa isoform in murine fetal lung. Both the 21 and 26 kDa proSP-C isoforms were detected after labelling CHO/SPC cells with [3H]palmitic acid. The formation of the 26 kDa proSP-C isoform in CHO/SPC cells and the 24 kDa proSP-C isoform in murine fetal lung was blocked by cerulenin, an inhibitor of fatty acid synthesis. In conclusion, proSP-C is associated with subcellular membranes. ProSP-C is palmitoylated and undergoes additional post-translational modification that is blocked by an inhibitor of fatty acid synthesis.
Vorbroker
D K
DK
Division of Pulmonary Biology, Children's Hospital Research Foundation, Cincinnati, OH 45229.
Dey
C
C
Weaver
T E
TE
Whitsett
J A
JA
D000818:Animals; D016466:CHO Cells; D002458:Cell Fractionation; D002569:Cerulenin / Q000494:pharmacology; D006224:Cricetinae; D004285:Dogs; D016521:Electrophoresis, Gel, Pulsed-Field; D005227:Fatty Acids / Q000096:biosynthesis; D007425:Intracellular Membranes / Q000378:metabolism*; D008168:Lung / Q000196:embryology / Q000378:metabolism; D051379:Mice; D008861:Microsomes / Q000378:metabolism; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism*; D010179:Pancreas / Q000378:metabolism; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D011510:Proteolipids / Q000235:genetics / Q000378:metabolism*; D011663:Pulmonary Surfactants / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D005227:Fatty Acids; D010169:Palmitic Acids; D011498:Protein Precursors; D011510:Proteolipids; D011663:Pulmonary Surfactants; D011994:Recombinant Proteins; D002569:Cerulenin; D019308:Palmitic Acid