Pulmonary surfactant contains, in addition to lipids, proteins of MW 26,000-40,000 (SAP-A) and of MW 4,000-50,000. The latter group of proteins are hydrophobic and copurify with the lipids. The various roles played in lung surfactant activity by surfactant associated proteins are currently a subject of debate and interest. Fetal lung maturity and the risk of respiratory distress syndrome (RDS) can be predicted by the assay of SAP-A in the amniotic fluid samples. In order to treat RDS in premature infants, recent clinical trials have successfully used preparations of exogenous surfactant which are free of SAP-A, but contain the hydrophobic proteins. This suggests an important role for hydrophobic proteins in surfactant function. Our recent results indicate that human SAP-A gene encodes both SAP-A and hydrophobic surfactant protein (HSAP). We have raised antisera to the hydrophobic proteins extracted from rat and human pulmonary surfactant. Immunoblot analysis of human surfactant revealed our antiserum has reactivity with hydrophobic surfactant proteins in the MW range 4,000-50,000. This antiserum lacks reactivity with SAP-A. This antiserum was then used to screen a lambda gtll library of human lung cDNA. A positive clone was isolated and the cDNA insert sequenced. The sequence of this cDNA bears homology to a portion of the SAP-A gene that is part of the untranslated portion of SAP-A mRNA. Upon Northern analysis, this clone identified a 2.5 kb mRNA (SAP-A mRNA is of this size). It is not clear whether the gene transcribes separate mRNAs for the two proteins, or a polypeptide is synthesized which is later cleaved to yield SAP-A and the hydrophobic protein. In the case of several other mosaic genes, production of multiple mRNAs or polypeptides is developmentally regulated and cell-specific. Further work is proposed on the identity and structural characterization of the hydrophobic protein, time of its appearance, developmental and hormonal modulation of the protein and its mRNA. Elucidation of the mechanism(s) whereby SAP-A gene expresses the two proteins is also proposed. Expression studies on the hydrophobic protein will be performed in concert with our studies on SAP-A. SAP-A shows molecular and charge heterogeneity as well as developmental changes in the amounts and heterogeneity. Further studies are proposed to determine whether SAP-A is expressed by separate genes in fetal and adult lung.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
5R01HL028193-06
Application #
3339625
Study Section
Human Embryology and Development Subcommittee 1 (HED)
Project Start
1983-04-01
Project End
1993-03-31
Budget Start
1990-04-01
Budget End
1991-03-31
Support Year
6
Fiscal Year
1990
Total Cost
Indirect Cost
Name
University of Pittsburgh
Department
Type
Schools of Medicine
DUNS #
053785812
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Singh, J; Singh, G; Katyal, S L et al. (1993) Immunohistochemistry of a human type I pneumocyte-associated protein in lung. Microsc Res Tech 26:357-65
Stripp, B R; Sawaya, P L; Luse, D S et al. (1992) cis-acting elements that confer lung epithelial cell expression of the CC10 gene. J Biol Chem 267:14703-12
Katyal, S L; Singh, G; Locker, J (1992) Characterization of a second human pulmonary surfactant-associated protein SP-A gene. Am J Respir Cell Mol Biol 6:446-52
Singh, G; Katyal, S L (1991) Pulmonary proteins as cell-specific markers. Exp Lung Res 17:245-53
Singh, G; Katyal, S L; Brown, W E et al. (1991) Identification, isolation, and partial characterization of a 7.5-kDa surfactant-associated protein. Exp Lung Res 17:559-67
Singh, G; Katyal, S L; Brown, W E et al. (1990) Clara cell 10 kDa protein (CC10): comparison of structure and function to uteroglobin. Biochim Biophys Acta 1039:348-55
Singh, G; Katyal, S L; Brown, W E et al. (1988) Amino-acid and cDNA nucleotide sequences of human Clara cell 10 kDa protein. Biochim Biophys Acta 950:329-37
Singh, G; Singh, J; Katyal, S L et al. (1988) Identification, cellular localization, isolation, and characterization of human Clara cell-specific 10 KD protein. J Histochem Cytochem 36:73-80
Katyal, S L; Singh, G; Ryan, L et al. (1988) Hydrophobic surfactant-associated proteins: electrophoretic and immunologic analyses and cellular localization in human lung. Exp Lung Res 14:655-69
Singh, G; Katyal, S L; Brown, W E et al. (1988) Pulmonary lysozyme--a secretory protein of type II pneumocytes in the rat. Am Rev Respir Dis 138:1261-7

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