Synthesis and processing of hydrophobic surfactant protein sp-c by
isolated rat type ii cells.
Beers, Michael F., Catherine Lomax.
Institute for Environmental Medicine, University of Pennsylvania
School of Medicine, Philadelphia, Pennsylvania 19104-6068, Pulmonary
and Critical Care Division, Department of Medicine, Hospital of the
University of Pennsylvania, Philadelphia, Pennsylvania 19104
APStracts 2:0118L, 1995.
Surfactant protein C (SP-C) is a 3.7 kDa hydrophobic peptide isolated
from organic extracts of pulmonary surfactant which is secreted by
alveolar type II cells after synthesis and post-translational
processing of a 21 kDa proSP-C peptide (SP-C21). Previously
characterized epitope-specific proSP-C antisera were used to study
early proteolytic steps of proSP-C processing by adult rat type II
cells. Western blotting and immunocytochemistry using antiNPROSP-C
(epitope = Met10-Phe23) each demonstrated marked attenuation of
proSP-C protein expression by culture on plastic. Processing was
therefore studied by metabolic labeling of freshly isolated type II
cells maintained in suspension in serum-free media. Using anti
-NPROSP-C, immunoprecipitation of cell lysates continuously labeled
for 4 hrs with 35S-methionine demonstrated radiolabeled bands of Mr
21-, 16-, and 10-6,000 while anti-CTERMSP-C (epitope = Ser149-Ser166)
failed to detect 35S-bands of Mr < 16,000. Pulse-chase studies
demonstrated synthesis of 35S-proSP-C21 with a time dependent
appearance of 16 kDa and 10-6 kDa forms which was blocked by addition
of Brefeldin A. SP-C precursors were not detected in the media.
Quantitative analysis of the major bands by direct (-counting
indicated a precursor-product relationship between SP-C21 and SP-C16.
These results demonstrate the utility of freshly isolated type II
cells for characterization of SP-C synthetic pathways and show that
early proSP-C processing events include synthesis of a 21 kDa primary
translation product followed by extensive intracellular proteolysis
of the proSP-C C-terminus in subcellular compartments of type II
cells which are distal to the trans-Golgi network.
Received 12 January 1995; accepted in final form 27 June 1995.
APS Manuscript Number L9-5.
Article publication pending Am. J. Physiol. (Lung Cell. Mol.
Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 18 July 1995.