Quantitation of protein kinase C by immunoblot-expression in different cell lines and response to phorbol esters
Antisera have been raised against human protein kinase C and also against a synthetic peptide based on the sequence of the bovine brain enzyme (LLNQEE‐GEYYNVPIPE). These antibodies react with protein kinase C from a number of species (human, murine, rat, rabbit, bovine), indicating substantial conse...
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| Veröffentlicht in: | J. Cell. Physiol.; (United States) 1987-01, Vol.130 (1), p.111-117 |
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| creator | Stabel, S. Rodriguez-Pena, A. Young, S. Rozengurt, E. Parker, P. J. |
| description | Antisera have been raised against human protein kinase C and also against a synthetic peptide based on the sequence of the bovine brain enzyme (LLNQEE‐GEYYNVPIPE). These antibodies react with protein kinase C from a number of species (human, murine, rat, rabbit, bovine), indicating substantial conservation of epitopes. These antisera have been used to quantitate directly protein kinase C by immunoblot analysis. We show here that there is a strict correlation between the leveis of immunoreactive polypeptide and extractable calcium‐ and phospholipid‐dependent kinase activity for various cell lines. Treatment of murine, rat, and human cells with phorbol dibutyrate was found to deplete levels of immunoreactive protein kinase C severely. A detailed study of the time course of this depletion in Swiss 3T3 cells shows that it follows precisely the loss of extractable activity. On exposure to 400 nM phorbol 12,13‐dibutyrate protein kinase C was essentially undetectable by 40 hours; the half‐life of this down‐regulation was 6.7 hours. This data thus demonstrate that the loss of immunoreactive protein kinase C and of extractable calcium‐ and phospholipid‐dependent kinase activity precisely parallels the phorbol ester induced down‐regulation of binding and responsiveness in Swiss 3T3 cells. |
| doi_str_mv | 10.1002/jcp.1041300116 |
| format | Article |
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J.</creator><creatorcontrib>Stabel, S. ; Rodriguez-Pena, A. ; Young, S. ; Rozengurt, E. ; Parker, P. J. ; Imperial Cancer Research Fund, London, England</creatorcontrib><description>Antisera have been raised against human protein kinase C and also against a synthetic peptide based on the sequence of the bovine brain enzyme (LLNQEE‐GEYYNVPIPE). These antibodies react with protein kinase C from a number of species (human, murine, rat, rabbit, bovine), indicating substantial conservation of epitopes. These antisera have been used to quantitate directly protein kinase C by immunoblot analysis. We show here that there is a strict correlation between the leveis of immunoreactive polypeptide and extractable calcium‐ and phospholipid‐dependent kinase activity for various cell lines. Treatment of murine, rat, and human cells with phorbol dibutyrate was found to deplete levels of immunoreactive protein kinase C severely. A detailed study of the time course of this depletion in Swiss 3T3 cells shows that it follows precisely the loss of extractable activity. On exposure to 400 nM phorbol 12,13‐dibutyrate protein kinase C was essentially undetectable by 40 hours; the half‐life of this down‐regulation was 6.7 hours. This data thus demonstrate that the loss of immunoreactive protein kinase C and of extractable calcium‐ and phospholipid‐dependent kinase activity precisely parallels the phorbol ester induced down‐regulation of binding and responsiveness in Swiss 3T3 cells.</description><identifier>ISSN: 0021-9541</identifier><identifier>EISSN: 1097-4652</identifier><identifier>DOI: 10.1002/jcp.1041300116</identifier><identifier>PMID: 3543027</identifier><identifier>CODEN: JCLLAX</identifier><language>eng</language><publisher>Hoboken: Wiley Subscription Services, Inc., A Wiley Company</publisher><subject>550601 - Medicine- Unsealed Radionuclides in Diagnostics ; 560300 - Chemicals Metabolism & Toxicology ; AMINO ACID SEQUENCE ; AMINO ACIDS ; ANIMAL CELLS ; ANIMALS ; Antigen-Antibody Complex ; BETA DECAY RADIOISOTOPES ; BETA-MINUS DECAY RADIOISOTOPES ; Biological and medical sciences ; BIOLOGICAL EFFECTS ; CARBOXYLIC ACIDS ; CARCINOGENS ; Cattle ; Cell Line ; Cell physiology ; CONNECTIVE TISSUE CELLS ; DAYS LIVING RADIOISOTOPES ; DRUGS ; ELECTRON CAPTURE RADIOISOTOPES ; ENZYME ACTIVITY ; ENZYMES ; ESTERS ; EVEN-ODD NUCLEI ; FIBROBLASTS ; Fundamental and applied biological sciences. Psychology ; Glioma ; Humans ; IMMUNE SERUMS ; Immunologic Techniques ; INTERMEDIATE MASS NUCLEI ; IODINE 125 ; IODINE ISOTOPES ; ISOTOPES ; LIGHT NUCLEI ; LIPOTROPIC FACTORS ; MAMMALS ; MAN ; METHIONINE ; MICE ; Molecular and cellular biology ; MOLECULAR STRUCTURE ; Neuroblastoma ; NUCLEI ; ODD-EVEN NUCLEI ; ORGANIC ACIDS ; ORGANIC COMPOUNDS ; ORGANIC SULFUR COMPOUNDS ; Phorbol 12,13-Dibutyrate ; PHORBOL ESTERS ; Phorbol Esters - pharmacology ; PHOSPHORUS-GROUP TRANSFERASES ; PHOSPHOTRANSFERASES ; PRIMATES ; Protein Kinase C - analysis ; Protein Kinase C - biosynthesis ; Protein Kinase C - metabolism ; RABBITS ; RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT ; RADIOASSAY ; RADIOISOTOPES ; RADIOLOGY AND NUCLEAR MEDICINE ; RATS ; Responses to growth factors, tumor promotors, other factors ; RODENTS ; SOMATIC CELLS ; SULFUR 35 ; SULFUR ISOTOPES ; Tetradecanoylphorbol Acetate - pharmacology ; TRANSFERASES ; TUMOR CELLS ; VERTEBRATES</subject><ispartof>J. Cell. Physiol.; (United States), 1987-01, Vol.130 (1), p.111-117</ispartof><rights>Copyright © 1987 Wiley‐Liss, Inc.</rights><rights>1987 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c4346-51ef848f56cec96dba07d91353a9e13416482e98eede3cf1b0a0c5b633224ea23</citedby><cites>FETCH-LOGICAL-c4346-51ef848f56cec96dba07d91353a9e13416482e98eede3cf1b0a0c5b633224ea23</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://onlinelibrary.wiley.com/doi/pdf/10.1002%2Fjcp.1041300116$$EPDF$$P50$$Gwiley$$H</linktopdf><linktohtml>$$Uhttps://onlinelibrary.wiley.com/doi/full/10.1002%2Fjcp.1041300116$$EHTML$$P50$$Gwiley$$H</linktohtml><link.rule.ids>314,780,784,885,1417,4023,27922,27923,27924,45573,45574</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=8312419$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/3543027$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://www.osti.gov/biblio/6515544$$D View this record in Osti.gov$$Hfree_for_read</backlink></links><search><creatorcontrib>Stabel, S.</creatorcontrib><creatorcontrib>Rodriguez-Pena, A.</creatorcontrib><creatorcontrib>Young, S.</creatorcontrib><creatorcontrib>Rozengurt, E.</creatorcontrib><creatorcontrib>Parker, P. J.</creatorcontrib><creatorcontrib>Imperial Cancer Research Fund, London, England</creatorcontrib><title>Quantitation of protein kinase C by immunoblot-expression in different cell lines and response to phorbol esters</title><title>J. Cell. Physiol.; (United States)</title><addtitle>J. Cell. Physiol</addtitle><description>Antisera have been raised against human protein kinase C and also against a synthetic peptide based on the sequence of the bovine brain enzyme (LLNQEE‐GEYYNVPIPE). These antibodies react with protein kinase C from a number of species (human, murine, rat, rabbit, bovine), indicating substantial conservation of epitopes. These antisera have been used to quantitate directly protein kinase C by immunoblot analysis. We show here that there is a strict correlation between the leveis of immunoreactive polypeptide and extractable calcium‐ and phospholipid‐dependent kinase activity for various cell lines. Treatment of murine, rat, and human cells with phorbol dibutyrate was found to deplete levels of immunoreactive protein kinase C severely. A detailed study of the time course of this depletion in Swiss 3T3 cells shows that it follows precisely the loss of extractable activity. On exposure to 400 nM phorbol 12,13‐dibutyrate protein kinase C was essentially undetectable by 40 hours; the half‐life of this down‐regulation was 6.7 hours. This data thus demonstrate that the loss of immunoreactive protein kinase C and of extractable calcium‐ and phospholipid‐dependent kinase activity precisely parallels the phorbol ester induced down‐regulation of binding and responsiveness in Swiss 3T3 cells.</description><subject>550601 - Medicine- Unsealed Radionuclides in Diagnostics</subject><subject>560300 - Chemicals Metabolism & Toxicology</subject><subject>AMINO ACID SEQUENCE</subject><subject>AMINO ACIDS</subject><subject>ANIMAL CELLS</subject><subject>ANIMALS</subject><subject>Antigen-Antibody Complex</subject><subject>BETA DECAY RADIOISOTOPES</subject><subject>BETA-MINUS DECAY RADIOISOTOPES</subject><subject>Biological and medical sciences</subject><subject>BIOLOGICAL EFFECTS</subject><subject>CARBOXYLIC ACIDS</subject><subject>CARCINOGENS</subject><subject>Cattle</subject><subject>Cell Line</subject><subject>Cell physiology</subject><subject>CONNECTIVE TISSUE CELLS</subject><subject>DAYS LIVING RADIOISOTOPES</subject><subject>DRUGS</subject><subject>ELECTRON CAPTURE RADIOISOTOPES</subject><subject>ENZYME ACTIVITY</subject><subject>ENZYMES</subject><subject>ESTERS</subject><subject>EVEN-ODD NUCLEI</subject><subject>FIBROBLASTS</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glioma</subject><subject>Humans</subject><subject>IMMUNE SERUMS</subject><subject>Immunologic Techniques</subject><subject>INTERMEDIATE MASS NUCLEI</subject><subject>IODINE 125</subject><subject>IODINE ISOTOPES</subject><subject>ISOTOPES</subject><subject>LIGHT NUCLEI</subject><subject>LIPOTROPIC FACTORS</subject><subject>MAMMALS</subject><subject>MAN</subject><subject>METHIONINE</subject><subject>MICE</subject><subject>Molecular and cellular biology</subject><subject>MOLECULAR STRUCTURE</subject><subject>Neuroblastoma</subject><subject>NUCLEI</subject><subject>ODD-EVEN NUCLEI</subject><subject>ORGANIC ACIDS</subject><subject>ORGANIC COMPOUNDS</subject><subject>ORGANIC SULFUR COMPOUNDS</subject><subject>Phorbol 12,13-Dibutyrate</subject><subject>PHORBOL ESTERS</subject><subject>Phorbol Esters - pharmacology</subject><subject>PHOSPHORUS-GROUP TRANSFERASES</subject><subject>PHOSPHOTRANSFERASES</subject><subject>PRIMATES</subject><subject>Protein Kinase C - analysis</subject><subject>Protein Kinase C - biosynthesis</subject><subject>Protein Kinase C - metabolism</subject><subject>RABBITS</subject><subject>RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT</subject><subject>RADIOASSAY</subject><subject>RADIOISOTOPES</subject><subject>RADIOLOGY AND NUCLEAR MEDICINE</subject><subject>RATS</subject><subject>Responses to growth factors, tumor promotors, other factors</subject><subject>RODENTS</subject><subject>SOMATIC CELLS</subject><subject>SULFUR 35</subject><subject>SULFUR ISOTOPES</subject><subject>Tetradecanoylphorbol Acetate - pharmacology</subject><subject>TRANSFERASES</subject><subject>TUMOR CELLS</subject><subject>VERTEBRATES</subject><issn>0021-9541</issn><issn>1097-4652</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1987</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkE1rFDEYgINY6lq9ehOCeJ2a75kcZdW2UtoKfoCXkMm8Q9POJkOSxe6_b8osK548JfA-T_LyIPSGklNKCPtw5-Z6EZQTQql6hlaU6LYRSrLnaFUB2mgp6Av0Muc7QojWnB-jYy4FJ6xdofnb1obiiy0-BhxHPKdYwAd874PNgNe432G_2WxD7KdYGniYE-T8BFdo8OMICULBDqYJTz5AxjYMuDJzDNUvEc-3MfVxwpALpPwKHY12yvB6f56gH18-f1-fN5fXZxfrj5eNE1yoRlIYO9GNUjlwWg29Je2gKZfcaqBcUCU6BroDGIC7kfbEEid7xTljAizjJ-jd8m7MxZvsfAF362II4IpRkkopRIVOF8ilmHOC0czJb2zaGUrMU15T85q_eavwdhHmbb-B4YDve9b5-_3cZmenMdngfD5gHadMUF0xvWB__AS7_3xqvq5v_lmhWVxfez4cXJvujWp5K82vqzOj2c1v9fNTZxR_BLXQo1I</recordid><startdate>198701</startdate><enddate>198701</enddate><creator>Stabel, S.</creator><creator>Rodriguez-Pena, A.</creator><creator>Young, S.</creator><creator>Rozengurt, E.</creator><creator>Parker, P. J.</creator><general>Wiley Subscription Services, Inc., A Wiley Company</general><general>Wiley-Liss</general><scope>BSCLL</scope><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>OTOTI</scope></search><sort><creationdate>198701</creationdate><title>Quantitation of protein kinase C by immunoblot-expression in different cell lines and response to phorbol esters</title><author>Stabel, S. ; Rodriguez-Pena, A. ; Young, S. ; Rozengurt, E. ; Parker, P. J.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c4346-51ef848f56cec96dba07d91353a9e13416482e98eede3cf1b0a0c5b633224ea23</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1987</creationdate><topic>550601 - Medicine- Unsealed Radionuclides in Diagnostics</topic><topic>560300 - Chemicals Metabolism & Toxicology</topic><topic>AMINO ACID SEQUENCE</topic><topic>AMINO ACIDS</topic><topic>ANIMAL CELLS</topic><topic>ANIMALS</topic><topic>Antigen-Antibody Complex</topic><topic>BETA DECAY RADIOISOTOPES</topic><topic>BETA-MINUS DECAY RADIOISOTOPES</topic><topic>Biological and medical sciences</topic><topic>BIOLOGICAL EFFECTS</topic><topic>CARBOXYLIC ACIDS</topic><topic>CARCINOGENS</topic><topic>Cattle</topic><topic>Cell Line</topic><topic>Cell physiology</topic><topic>CONNECTIVE TISSUE CELLS</topic><topic>DAYS LIVING RADIOISOTOPES</topic><topic>DRUGS</topic><topic>ELECTRON CAPTURE RADIOISOTOPES</topic><topic>ENZYME ACTIVITY</topic><topic>ENZYMES</topic><topic>ESTERS</topic><topic>EVEN-ODD NUCLEI</topic><topic>FIBROBLASTS</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Glioma</topic><topic>Humans</topic><topic>IMMUNE SERUMS</topic><topic>Immunologic Techniques</topic><topic>INTERMEDIATE MASS NUCLEI</topic><topic>IODINE 125</topic><topic>IODINE ISOTOPES</topic><topic>ISOTOPES</topic><topic>LIGHT NUCLEI</topic><topic>LIPOTROPIC FACTORS</topic><topic>MAMMALS</topic><topic>MAN</topic><topic>METHIONINE</topic><topic>MICE</topic><topic>Molecular and cellular biology</topic><topic>MOLECULAR STRUCTURE</topic><topic>Neuroblastoma</topic><topic>NUCLEI</topic><topic>ODD-EVEN NUCLEI</topic><topic>ORGANIC ACIDS</topic><topic>ORGANIC COMPOUNDS</topic><topic>ORGANIC SULFUR COMPOUNDS</topic><topic>Phorbol 12,13-Dibutyrate</topic><topic>PHORBOL ESTERS</topic><topic>Phorbol Esters - pharmacology</topic><topic>PHOSPHORUS-GROUP TRANSFERASES</topic><topic>PHOSPHOTRANSFERASES</topic><topic>PRIMATES</topic><topic>Protein Kinase C - analysis</topic><topic>Protein Kinase C - biosynthesis</topic><topic>Protein Kinase C - metabolism</topic><topic>RABBITS</topic><topic>RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT</topic><topic>RADIOASSAY</topic><topic>RADIOISOTOPES</topic><topic>RADIOLOGY AND NUCLEAR MEDICINE</topic><topic>RATS</topic><topic>Responses to growth factors, tumor promotors, other factors</topic><topic>RODENTS</topic><topic>SOMATIC CELLS</topic><topic>SULFUR 35</topic><topic>SULFUR ISOTOPES</topic><topic>Tetradecanoylphorbol Acetate - pharmacology</topic><topic>TRANSFERASES</topic><topic>TUMOR CELLS</topic><topic>VERTEBRATES</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Stabel, S.</creatorcontrib><creatorcontrib>Rodriguez-Pena, A.</creatorcontrib><creatorcontrib>Young, S.</creatorcontrib><creatorcontrib>Rozengurt, E.</creatorcontrib><creatorcontrib>Parker, P. J.</creatorcontrib><creatorcontrib>Imperial Cancer Research Fund, London, England</creatorcontrib><collection>Istex</collection><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>OSTI.GOV</collection><jtitle>J. Cell. Physiol.; (United States)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Stabel, S.</au><au>Rodriguez-Pena, A.</au><au>Young, S.</au><au>Rozengurt, E.</au><au>Parker, P. J.</au><aucorp>Imperial Cancer Research Fund, London, England</aucorp><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Quantitation of protein kinase C by immunoblot-expression in different cell lines and response to phorbol esters</atitle><jtitle>J. Cell. Physiol.; (United States)</jtitle><addtitle>J. Cell. Physiol</addtitle><date>1987-01</date><risdate>1987</risdate><volume>130</volume><issue>1</issue><spage>111</spage><epage>117</epage><pages>111-117</pages><issn>0021-9541</issn><eissn>1097-4652</eissn><coden>JCLLAX</coden><abstract>Antisera have been raised against human protein kinase C and also against a synthetic peptide based on the sequence of the bovine brain enzyme (LLNQEE‐GEYYNVPIPE). These antibodies react with protein kinase C from a number of species (human, murine, rat, rabbit, bovine), indicating substantial conservation of epitopes. These antisera have been used to quantitate directly protein kinase C by immunoblot analysis. We show here that there is a strict correlation between the leveis of immunoreactive polypeptide and extractable calcium‐ and phospholipid‐dependent kinase activity for various cell lines. Treatment of murine, rat, and human cells with phorbol dibutyrate was found to deplete levels of immunoreactive protein kinase C severely. A detailed study of the time course of this depletion in Swiss 3T3 cells shows that it follows precisely the loss of extractable activity. On exposure to 400 nM phorbol 12,13‐dibutyrate protein kinase C was essentially undetectable by 40 hours; the half‐life of this down‐regulation was 6.7 hours. This data thus demonstrate that the loss of immunoreactive protein kinase C and of extractable calcium‐ and phospholipid‐dependent kinase activity precisely parallels the phorbol ester induced down‐regulation of binding and responsiveness in Swiss 3T3 cells.</abstract><cop>Hoboken</cop><pub>Wiley Subscription Services, Inc., A Wiley Company</pub><pmid>3543027</pmid><doi>10.1002/jcp.1041300116</doi><tpages>7</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0021-9541 |
| ispartof | J. Cell. Physiol.; (United States), 1987-01, Vol.130 (1), p.111-117 |
| issn | 0021-9541 1097-4652 |
| language | eng |
| recordid | cdi_crossref_primary_10_1002_jcp_1041300116 |
| source | MEDLINE; Wiley Online Library All Journals |
| subjects | 550601 - Medicine- Unsealed Radionuclides in Diagnostics 560300 - Chemicals Metabolism & Toxicology AMINO ACID SEQUENCE AMINO ACIDS ANIMAL CELLS ANIMALS Antigen-Antibody Complex BETA DECAY RADIOISOTOPES BETA-MINUS DECAY RADIOISOTOPES Biological and medical sciences BIOLOGICAL EFFECTS CARBOXYLIC ACIDS CARCINOGENS Cattle Cell Line Cell physiology CONNECTIVE TISSUE CELLS DAYS LIVING RADIOISOTOPES DRUGS ELECTRON CAPTURE RADIOISOTOPES ENZYME ACTIVITY ENZYMES ESTERS EVEN-ODD NUCLEI FIBROBLASTS Fundamental and applied biological sciences. Psychology Glioma Humans IMMUNE SERUMS Immunologic Techniques INTERMEDIATE MASS NUCLEI IODINE 125 IODINE ISOTOPES ISOTOPES LIGHT NUCLEI LIPOTROPIC FACTORS MAMMALS MAN METHIONINE MICE Molecular and cellular biology MOLECULAR STRUCTURE Neuroblastoma NUCLEI ODD-EVEN NUCLEI ORGANIC ACIDS ORGANIC COMPOUNDS ORGANIC SULFUR COMPOUNDS Phorbol 12,13-Dibutyrate PHORBOL ESTERS Phorbol Esters - pharmacology PHOSPHORUS-GROUP TRANSFERASES PHOSPHOTRANSFERASES PRIMATES Protein Kinase C - analysis Protein Kinase C - biosynthesis Protein Kinase C - metabolism RABBITS RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT RADIOASSAY RADIOISOTOPES RADIOLOGY AND NUCLEAR MEDICINE RATS Responses to growth factors, tumor promotors, other factors RODENTS SOMATIC CELLS SULFUR 35 SULFUR ISOTOPES Tetradecanoylphorbol Acetate - pharmacology TRANSFERASES TUMOR CELLS VERTEBRATES |
| title | Quantitation of protein kinase C by immunoblot-expression in different cell lines and response to phorbol esters |
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