Localization, synthesis, and processing of surfactant protein SP-C in rat lung analyzed by epitope-specific antipeptide antibodies
Surfactant protein C (SP-C), a 3.7-kDa hydrophobic lung-specific protein, is synthesized and secreted by pulmonary type II cells through proteolytic processing of a 21-kDa propeptide (SP-C21) by currently undefined pathways. Previously, we reported the production of a polyclonal antibody against rat...
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| Veröffentlicht in: | The Journal of biological chemistry 1994-08, Vol.269 (32), p.20318-20328 |
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| creator | BEERS, M. F KIM, C. Y CHANDRA DODIA FISHER, A. B |
| description | Surfactant protein C (SP-C), a 3.7-kDa hydrophobic lung-specific protein, is synthesized and secreted by pulmonary type II
cells through proteolytic processing of a 21-kDa propeptide (SP-C21) by currently undefined pathways. Previously, we reported
the production of a polyclonal antibody against rat SP-C21 (anti-CPROSP-C) using a synthetic peptide as the immunizing antigen
(Beers, M. F., Wali, A., Eckenhoff, M. E. F., Feinstein, S., Fisher, J. H., and Fisher, A. B. (1992) Am. J. Respir. Cell.
Mol. Biol. 7, 368-378). In this study, two additional epitope-specific proSP-C antibodies produced using synthetic peptide
sequences were utilized to examine synthetic processing of SP-C. Anti-NPROSP-C (Met10-Gln23) and anti-CTERMSP-C (Ser149-Ser166)
recognized native proSP-C21 produced from in vitro translation of SP-C cDNA. Immunocytochemistry using anti-NPROSP-C confirmed
the localization of proSP-C peptides exclusively in type II cells. Western analysis of subcellular fractions identified a
single 21-kDa band in microsomes and a 16-kDa form in lamellar bodies each recognized by all three antisera, while anti-NPROSP-C
also uniquely identified a prominent 5-6-kDa form in lamellar bodies. In a perfused rat lung model labeled with [35S]cysteine/methionine,
immunoprecipitation of lung homogenate and lamellar body fractions identified early appearances of 35S-labeled 21-, 18-, and
16-kDa SP-C forms in homogenate and a 16-kDa intermediate form in lamellar bodies. Anti-NPROSP-C also exclusively detected
time-dependent appearances of 5-10-kDa proSP-C forms in lamellar bodies and homogenates. Processing of proSP-C21 was completely
blocked by inclusion of brefeldin A (15 micrograms/ml) in the perfusate. These results demonstrate that synthetic peptides
can be used to produce epitope-specific antisera which recognize more hydrophilic domains of proSP-C and show that proSP-C
processing occurs intracellularly in subcellular compartments of type II cells which are distal to the trans-Golgi network. |
| doi_str_mv | 10.1016/s0021-9258(17)31994-4 |
| format | Article |
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cells through proteolytic processing of a 21-kDa propeptide (SP-C21) by currently undefined pathways. Previously, we reported
the production of a polyclonal antibody against rat SP-C21 (anti-CPROSP-C) using a synthetic peptide as the immunizing antigen
(Beers, M. F., Wali, A., Eckenhoff, M. E. F., Feinstein, S., Fisher, J. H., and Fisher, A. B. (1992) Am. J. Respir. Cell.
Mol. Biol. 7, 368-378). In this study, two additional epitope-specific proSP-C antibodies produced using synthetic peptide
sequences were utilized to examine synthetic processing of SP-C. Anti-NPROSP-C (Met10-Gln23) and anti-CTERMSP-C (Ser149-Ser166)
recognized native proSP-C21 produced from in vitro translation of SP-C cDNA. Immunocytochemistry using anti-NPROSP-C confirmed
the localization of proSP-C peptides exclusively in type II cells. Western analysis of subcellular fractions identified a
single 21-kDa band in microsomes and a 16-kDa form in lamellar bodies each recognized by all three antisera, while anti-NPROSP-C
also uniquely identified a prominent 5-6-kDa form in lamellar bodies. In a perfused rat lung model labeled with [35S]cysteine/methionine,
immunoprecipitation of lung homogenate and lamellar body fractions identified early appearances of 35S-labeled 21-, 18-, and
16-kDa SP-C forms in homogenate and a 16-kDa intermediate form in lamellar bodies. Anti-NPROSP-C also exclusively detected
time-dependent appearances of 5-10-kDa proSP-C forms in lamellar bodies and homogenates. Processing of proSP-C21 was completely
blocked by inclusion of brefeldin A (15 micrograms/ml) in the perfusate. These results demonstrate that synthetic peptides
can be used to produce epitope-specific antisera which recognize more hydrophilic domains of proSP-C and show that proSP-C
processing occurs intracellularly in subcellular compartments of type II cells which are distal to the trans-Golgi network.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1016/s0021-9258(17)31994-4</identifier><identifier>PMID: 7519606</identifier><identifier>CODEN: JBCHA3</identifier><language>eng</language><publisher>Bethesda, MD: American Society for Biochemistry and Molecular Biology</publisher><subject>Amino Acid Sequence ; Analytical, structural and metabolic biochemistry ; Animals ; Antibodies - immunology ; Antibody Specificity ; Biological and medical sciences ; Epitopes - immunology ; Fundamental and applied biological sciences. Psychology ; Immunohistochemistry ; In Vitro Techniques ; Lung - metabolism ; Miscellaneous ; Molecular and cellular biology ; Molecular genetics ; Molecular Sequence Data ; Peptides - immunology ; Protein Processing, Post-Translational ; Proteins ; Proteolipids - biosynthesis ; Proteolipids - immunology ; Proteolipids - metabolism ; Pulmonary Surfactants - biosynthesis ; Pulmonary Surfactants - immunology ; Pulmonary Surfactants - metabolism ; Rats ; Rats, Sprague-Dawley ; Subcellular Fractions - metabolism ; Translation. Translation factors. Protein processing</subject><ispartof>The Journal of biological chemistry, 1994-08, Vol.269 (32), p.20318-20328</ispartof><rights>1994 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c475t-c041ec784edf7f7f41737755a57684a0d0a56207c07c3cfe69a9f42ad5a3f2533</citedby><cites>FETCH-LOGICAL-c475t-c041ec784edf7f7f41737755a57684a0d0a56207c07c3cfe69a9f42ad5a3f2533</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=4259422$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/7519606$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>BEERS, M. F</creatorcontrib><creatorcontrib>KIM, C. Y</creatorcontrib><creatorcontrib>CHANDRA DODIA</creatorcontrib><creatorcontrib>FISHER, A. B</creatorcontrib><title>Localization, synthesis, and processing of surfactant protein SP-C in rat lung analyzed by epitope-specific antipeptide antibodies</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>Surfactant protein C (SP-C), a 3.7-kDa hydrophobic lung-specific protein, is synthesized and secreted by pulmonary type II
cells through proteolytic processing of a 21-kDa propeptide (SP-C21) by currently undefined pathways. Previously, we reported
the production of a polyclonal antibody against rat SP-C21 (anti-CPROSP-C) using a synthetic peptide as the immunizing antigen
(Beers, M. F., Wali, A., Eckenhoff, M. E. F., Feinstein, S., Fisher, J. H., and Fisher, A. B. (1992) Am. J. Respir. Cell.
Mol. Biol. 7, 368-378). In this study, two additional epitope-specific proSP-C antibodies produced using synthetic peptide
sequences were utilized to examine synthetic processing of SP-C. Anti-NPROSP-C (Met10-Gln23) and anti-CTERMSP-C (Ser149-Ser166)
recognized native proSP-C21 produced from in vitro translation of SP-C cDNA. Immunocytochemistry using anti-NPROSP-C confirmed
the localization of proSP-C peptides exclusively in type II cells. Western analysis of subcellular fractions identified a
single 21-kDa band in microsomes and a 16-kDa form in lamellar bodies each recognized by all three antisera, while anti-NPROSP-C
also uniquely identified a prominent 5-6-kDa form in lamellar bodies. In a perfused rat lung model labeled with [35S]cysteine/methionine,
immunoprecipitation of lung homogenate and lamellar body fractions identified early appearances of 35S-labeled 21-, 18-, and
16-kDa SP-C forms in homogenate and a 16-kDa intermediate form in lamellar bodies. Anti-NPROSP-C also exclusively detected
time-dependent appearances of 5-10-kDa proSP-C forms in lamellar bodies and homogenates. Processing of proSP-C21 was completely
blocked by inclusion of brefeldin A (15 micrograms/ml) in the perfusate. These results demonstrate that synthetic peptides
can be used to produce epitope-specific antisera which recognize more hydrophilic domains of proSP-C and show that proSP-C
processing occurs intracellularly in subcellular compartments of type II cells which are distal to the trans-Golgi network.</description><subject>Amino Acid Sequence</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>Animals</subject><subject>Antibodies - immunology</subject><subject>Antibody Specificity</subject><subject>Biological and medical sciences</subject><subject>Epitopes - immunology</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Immunohistochemistry</subject><subject>In Vitro Techniques</subject><subject>Lung - metabolism</subject><subject>Miscellaneous</subject><subject>Molecular and cellular biology</subject><subject>Molecular genetics</subject><subject>Molecular Sequence Data</subject><subject>Peptides - immunology</subject><subject>Protein Processing, Post-Translational</subject><subject>Proteins</subject><subject>Proteolipids - biosynthesis</subject><subject>Proteolipids - immunology</subject><subject>Proteolipids - metabolism</subject><subject>Pulmonary Surfactants - biosynthesis</subject><subject>Pulmonary Surfactants - immunology</subject><subject>Pulmonary Surfactants - metabolism</subject><subject>Rats</subject><subject>Rats, Sprague-Dawley</subject><subject>Subcellular Fractions - metabolism</subject><subject>Translation. Translation factors. Protein processing</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1994</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpFkF2LEzEUhoMoa139CQu5EFHY0XxncinFLygorIJ3Ic2cbCPTmXFOBule-stNd0vNByfwPicJDyFXnL3ljJt3yJjgjRO6fc3tG8mdU416RFactbKRmv98TFZn5Cl5hviL1aEcvyAXVnNnmFmRv5sxhj7fhZLH4ZriYSg7wIzXNAwdneYxAmIebumYKC5zCrGEoRyDAnmgN9-aNa11DoX2S8XCEPrDHXR0e6Aw5TJO0OAEMacca1jyBFPJHdyft2OXAZ-TJyn0CC9O9ZL8-Pjh-_pzs_n66cv6_aaJyurSRKY4RNsq6JKtU3ErrdU6aGtaFVjHgjaC2ViXjAmMCy4pETodZBJaykvy6uHe-vnfC2Dx-4wR-j4MMC7orTHC1F1B_QDGeUScIflpzvswHzxn_uje3xzF-qNYz62_d-9V7bs6PbBs99Cdu06ya_7ylAes0tMchpjxjCmhnRLiP7bLt7s_eQa_zWPcwd4L47wUXjDJW_kP7BeaOg</recordid><startdate>19940812</startdate><enddate>19940812</enddate><creator>BEERS, M. F</creator><creator>KIM, C. Y</creator><creator>CHANDRA DODIA</creator><creator>FISHER, A. B</creator><general>American Society for Biochemistry and Molecular Biology</general><scope>IQODW</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>19940812</creationdate><title>Localization, synthesis, and processing of surfactant protein SP-C in rat lung analyzed by epitope-specific antipeptide antibodies</title><author>BEERS, M. F ; KIM, C. Y ; CHANDRA DODIA ; FISHER, A. B</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c475t-c041ec784edf7f7f41737755a57684a0d0a56207c07c3cfe69a9f42ad5a3f2533</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1994</creationdate><topic>Amino Acid Sequence</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>Animals</topic><topic>Antibodies - immunology</topic><topic>Antibody Specificity</topic><topic>Biological and medical sciences</topic><topic>Epitopes - immunology</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Immunohistochemistry</topic><topic>In Vitro Techniques</topic><topic>Lung - metabolism</topic><topic>Miscellaneous</topic><topic>Molecular and cellular biology</topic><topic>Molecular genetics</topic><topic>Molecular Sequence Data</topic><topic>Peptides - immunology</topic><topic>Protein Processing, Post-Translational</topic><topic>Proteins</topic><topic>Proteolipids - biosynthesis</topic><topic>Proteolipids - immunology</topic><topic>Proteolipids - metabolism</topic><topic>Pulmonary Surfactants - biosynthesis</topic><topic>Pulmonary Surfactants - immunology</topic><topic>Pulmonary Surfactants - metabolism</topic><topic>Rats</topic><topic>Rats, Sprague-Dawley</topic><topic>Subcellular Fractions - metabolism</topic><topic>Translation. Translation factors. Protein processing</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>BEERS, M. F</creatorcontrib><creatorcontrib>KIM, C. Y</creatorcontrib><creatorcontrib>CHANDRA DODIA</creatorcontrib><creatorcontrib>FISHER, A. B</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>BEERS, M. F</au><au>KIM, C. Y</au><au>CHANDRA DODIA</au><au>FISHER, A. B</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Localization, synthesis, and processing of surfactant protein SP-C in rat lung analyzed by epitope-specific antipeptide antibodies</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>1994-08-12</date><risdate>1994</risdate><volume>269</volume><issue>32</issue><spage>20318</spage><epage>20328</epage><pages>20318-20328</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><coden>JBCHA3</coden><abstract>Surfactant protein C (SP-C), a 3.7-kDa hydrophobic lung-specific protein, is synthesized and secreted by pulmonary type II
cells through proteolytic processing of a 21-kDa propeptide (SP-C21) by currently undefined pathways. Previously, we reported
the production of a polyclonal antibody against rat SP-C21 (anti-CPROSP-C) using a synthetic peptide as the immunizing antigen
(Beers, M. F., Wali, A., Eckenhoff, M. E. F., Feinstein, S., Fisher, J. H., and Fisher, A. B. (1992) Am. J. Respir. Cell.
Mol. Biol. 7, 368-378). In this study, two additional epitope-specific proSP-C antibodies produced using synthetic peptide
sequences were utilized to examine synthetic processing of SP-C. Anti-NPROSP-C (Met10-Gln23) and anti-CTERMSP-C (Ser149-Ser166)
recognized native proSP-C21 produced from in vitro translation of SP-C cDNA. Immunocytochemistry using anti-NPROSP-C confirmed
the localization of proSP-C peptides exclusively in type II cells. Western analysis of subcellular fractions identified a
single 21-kDa band in microsomes and a 16-kDa form in lamellar bodies each recognized by all three antisera, while anti-NPROSP-C
also uniquely identified a prominent 5-6-kDa form in lamellar bodies. In a perfused rat lung model labeled with [35S]cysteine/methionine,
immunoprecipitation of lung homogenate and lamellar body fractions identified early appearances of 35S-labeled 21-, 18-, and
16-kDa SP-C forms in homogenate and a 16-kDa intermediate form in lamellar bodies. Anti-NPROSP-C also exclusively detected
time-dependent appearances of 5-10-kDa proSP-C forms in lamellar bodies and homogenates. Processing of proSP-C21 was completely
blocked by inclusion of brefeldin A (15 micrograms/ml) in the perfusate. These results demonstrate that synthetic peptides
can be used to produce epitope-specific antisera which recognize more hydrophilic domains of proSP-C and show that proSP-C
processing occurs intracellularly in subcellular compartments of type II cells which are distal to the trans-Golgi network.</abstract><cop>Bethesda, MD</cop><pub>American Society for Biochemistry and Molecular Biology</pub><pmid>7519606</pmid><doi>10.1016/s0021-9258(17)31994-4</doi><tpages>11</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | The Journal of biological chemistry, 1994-08, Vol.269 (32), p.20318-20328 |
| issn | 0021-9258 1083-351X |
| language | eng |
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| source | MEDLINE; EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | Amino Acid Sequence Analytical, structural and metabolic biochemistry Animals Antibodies - immunology Antibody Specificity Biological and medical sciences Epitopes - immunology Fundamental and applied biological sciences. Psychology Immunohistochemistry In Vitro Techniques Lung - metabolism Miscellaneous Molecular and cellular biology Molecular genetics Molecular Sequence Data Peptides - immunology Protein Processing, Post-Translational Proteins Proteolipids - biosynthesis Proteolipids - immunology Proteolipids - metabolism Pulmonary Surfactants - biosynthesis Pulmonary Surfactants - immunology Pulmonary Surfactants - metabolism Rats Rats, Sprague-Dawley Subcellular Fractions - metabolism Translation. Translation factors. Protein processing |
| title | Localization, synthesis, and processing of surfactant protein SP-C in rat lung analyzed by epitope-specific antipeptide antibodies |
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