Endogenous human MDM2-C is highly expressed in human cancers and functions as a p53-independent growth activator
Human cancers over-expressing mdm2, through a T to G variation at a single nucleotide polymorphism at position 309 (mdm2 SNP309), have functionally inactivated p53 that is not effectively degraded. They also have high expression of the alternatively spliced transcript, mdm2-C. Alternatively spliced...
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| description | Human cancers over-expressing mdm2, through a T to G variation at a single nucleotide polymorphism at position 309 (mdm2 SNP309), have functionally inactivated p53 that is not effectively degraded. They also have high expression of the alternatively spliced transcript, mdm2-C. Alternatively spliced mdm2 transcripts are expressed in many forms of human cancer and when they are exogenously expressed they transform human cells. However no study to date has detected endogenous MDM2 protein isoforms. Studies with exogenous expression of splice variants have been carried out with mdm2-A and mdm2-B, but the mdm2-C isoform has remained virtually unexplored. We addressed the cellular influence of exogenously expressed MDM2-C, and asked if endogenous MDM2-C protein was present in human cancers. To detect endogenous MDM2-C protein, we created a human MDM2-C antibody to the splice junction epitope of exons four and ten (MDM2 C410) and validated the antibody with in vitro translated full length MDM2 compared to MDM2-C. Interestingly, we discovered that MDM2-C co-migrates with MDM2-FL at approximately 98 kDa. Using the validated C410 antibody, we detected high expression of endogenous MDM2-C in human cancer cell lines and human cancer tissues. In the estrogen receptor positive (ER+) mdm2 G/G SNP309 breast cancer cell line, T47D, we observed an increase in endogenous MDM2-C protein with estrogen treatment. MDM2-C localized to the nucleus and the cytoplasm. We examined the biological activity of MDM2-C by exogenously expressing the protein and observed that MDM2-C did not efficiently target p53 for degradation or reduce p53 transcriptional activity. Exogenous expression of MDM2-C in p53-null human cancer cells increased colony formation, indicating p53-independent tumorigenic properties. Our data indicate a role for MDM2-C that does not require the inhibition of p53 for increasing cancer cell proliferation and survival. |
| doi_str_mv | 10.1371/journal.pone.0077643 |
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They also have high expression of the alternatively spliced transcript, mdm2-C. Alternatively spliced mdm2 transcripts are expressed in many forms of human cancer and when they are exogenously expressed they transform human cells. However no study to date has detected endogenous MDM2 protein isoforms. Studies with exogenous expression of splice variants have been carried out with mdm2-A and mdm2-B, but the mdm2-C isoform has remained virtually unexplored. We addressed the cellular influence of exogenously expressed MDM2-C, and asked if endogenous MDM2-C protein was present in human cancers. To detect endogenous MDM2-C protein, we created a human MDM2-C antibody to the splice junction epitope of exons four and ten (MDM2 C410) and validated the antibody with in vitro translated full length MDM2 compared to MDM2-C. Interestingly, we discovered that MDM2-C co-migrates with MDM2-FL at approximately 98 kDa. Using the validated C410 antibody, we detected high expression of endogenous MDM2-C in human cancer cell lines and human cancer tissues. In the estrogen receptor positive (ER+) mdm2 G/G SNP309 breast cancer cell line, T47D, we observed an increase in endogenous MDM2-C protein with estrogen treatment. MDM2-C localized to the nucleus and the cytoplasm. We examined the biological activity of MDM2-C by exogenously expressing the protein and observed that MDM2-C did not efficiently target p53 for degradation or reduce p53 transcriptional activity. Exogenous expression of MDM2-C in p53-null human cancer cells increased colony formation, indicating p53-independent tumorigenic properties. Our data indicate a role for MDM2-C that does not require the inhibition of p53 for increasing cancer cell proliferation and survival.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0077643</identifier><identifier>PMID: 24147044</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Alternative splicing ; Antibodies ; Apoptosis ; Biochemistry ; Biological activity ; Biology ; Blotting, Northern ; Blotting, Western ; Breast cancer ; C protein ; Cancer ; Cell cycle ; Cell Line, Tumor ; Cell proliferation ; Cell survival ; Cells (Biology) ; Cytoplasm ; Deoxyribonucleic acid ; Departments ; DNA ; Epitopes ; Estrogen receptors ; Estrogens ; Exons ; Fluorescent Antibody Technique ; Gene amplification ; Genetic aspects ; Genetic polymorphisms ; Humans ; Immunohistochemistry ; Immunoprecipitation ; Isoforms ; MDM2 protein ; Microscopy, Confocal ; Monoclonal antibodies ; Neoplasms - genetics ; Neoplasms - metabolism ; Nuclei ; p53 Protein ; Polymorphism ; Protein Isoforms - genetics ; Protein Isoforms - metabolism ; Proteins ; Proto-Oncogene Proteins c-mdm2 - genetics ; Proto-Oncogene Proteins c-mdm2 - metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Single-nucleotide polymorphism ; Transcription ; Tumor cell lines ; Tumor proteins ; Tumor Suppressor Protein p53 - genetics ; Tumor Suppressor Protein p53 - metabolism ; Tumorigenesis</subject><ispartof>PloS one, 2013-10, Vol.8 (10), p.e77643-e77643</ispartof><rights>COPYRIGHT 2013 Public Library of Science</rights><rights>2013 Okoro et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License: https://creativecommons.org/licenses/by/4.0/ (the “License”), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited. Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2013 Okoro et al 2013 Okoro et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c692t-282caf57cb2421d11f6ccc42d6c910dd5d4109fe43caff85fe124d1aa83831c3</citedby><cites>FETCH-LOGICAL-c692t-282caf57cb2421d11f6ccc42d6c910dd5d4109fe43caff85fe124d1aa83831c3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3795673/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3795673/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,860,881,2096,2915,23847,27903,27904,53769,53771,79346,79347</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/24147044$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Deb, Sumitra</contributor><creatorcontrib>Okoro, Danielle R</creatorcontrib><creatorcontrib>Arva, Nicoleta</creatorcontrib><creatorcontrib>Gao, Chong</creatorcontrib><creatorcontrib>Polotskaia, Alla</creatorcontrib><creatorcontrib>Puente, Cindy</creatorcontrib><creatorcontrib>Rosso, Melissa</creatorcontrib><creatorcontrib>Bargonetti, Jill</creatorcontrib><title>Endogenous human MDM2-C is highly expressed in human cancers and functions as a p53-independent growth activator</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Human cancers over-expressing mdm2, through a T to G variation at a single nucleotide polymorphism at position 309 (mdm2 SNP309), have functionally inactivated p53 that is not effectively degraded. They also have high expression of the alternatively spliced transcript, mdm2-C. Alternatively spliced mdm2 transcripts are expressed in many forms of human cancer and when they are exogenously expressed they transform human cells. However no study to date has detected endogenous MDM2 protein isoforms. Studies with exogenous expression of splice variants have been carried out with mdm2-A and mdm2-B, but the mdm2-C isoform has remained virtually unexplored. We addressed the cellular influence of exogenously expressed MDM2-C, and asked if endogenous MDM2-C protein was present in human cancers. To detect endogenous MDM2-C protein, we created a human MDM2-C antibody to the splice junction epitope of exons four and ten (MDM2 C410) and validated the antibody with in vitro translated full length MDM2 compared to MDM2-C. Interestingly, we discovered that MDM2-C co-migrates with MDM2-FL at approximately 98 kDa. Using the validated C410 antibody, we detected high expression of endogenous MDM2-C in human cancer cell lines and human cancer tissues. In the estrogen receptor positive (ER+) mdm2 G/G SNP309 breast cancer cell line, T47D, we observed an increase in endogenous MDM2-C protein with estrogen treatment. MDM2-C localized to the nucleus and the cytoplasm. We examined the biological activity of MDM2-C by exogenously expressing the protein and observed that MDM2-C did not efficiently target p53 for degradation or reduce p53 transcriptional activity. Exogenous expression of MDM2-C in p53-null human cancer cells increased colony formation, indicating p53-independent tumorigenic properties. Our data indicate a role for MDM2-C that does not require the inhibition of p53 for increasing cancer cell proliferation and survival.</description><subject>Alternative splicing</subject><subject>Antibodies</subject><subject>Apoptosis</subject><subject>Biochemistry</subject><subject>Biological activity</subject><subject>Biology</subject><subject>Blotting, Northern</subject><subject>Blotting, Western</subject><subject>Breast cancer</subject><subject>C protein</subject><subject>Cancer</subject><subject>Cell cycle</subject><subject>Cell Line, Tumor</subject><subject>Cell proliferation</subject><subject>Cell survival</subject><subject>Cells (Biology)</subject><subject>Cytoplasm</subject><subject>Deoxyribonucleic acid</subject><subject>Departments</subject><subject>DNA</subject><subject>Epitopes</subject><subject>Estrogen receptors</subject><subject>Estrogens</subject><subject>Exons</subject><subject>Fluorescent Antibody Technique</subject><subject>Gene amplification</subject><subject>Genetic aspects</subject><subject>Genetic polymorphisms</subject><subject>Humans</subject><subject>Immunohistochemistry</subject><subject>Immunoprecipitation</subject><subject>Isoforms</subject><subject>MDM2 protein</subject><subject>Microscopy, Confocal</subject><subject>Monoclonal antibodies</subject><subject>Neoplasms - genetics</subject><subject>Neoplasms - metabolism</subject><subject>Nuclei</subject><subject>p53 Protein</subject><subject>Polymorphism</subject><subject>Protein Isoforms - genetics</subject><subject>Protein Isoforms - metabolism</subject><subject>Proteins</subject><subject>Proto-Oncogene Proteins c-mdm2 - genetics</subject><subject>Proto-Oncogene Proteins c-mdm2 - metabolism</subject><subject>Reverse Transcriptase Polymerase Chain Reaction</subject><subject>Single-nucleotide polymorphism</subject><subject>Transcription</subject><subject>Tumor cell lines</subject><subject>Tumor proteins</subject><subject>Tumor Suppressor Protein p53 - 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human MDM2-C is highly expressed in human cancers and functions as a p53-independent growth activator</title><author>Okoro, Danielle R ; Arva, Nicoleta ; Gao, Chong ; Polotskaia, Alla ; Puente, Cindy ; Rosso, Melissa ; Bargonetti, Jill</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c692t-282caf57cb2421d11f6ccc42d6c910dd5d4109fe43caff85fe124d1aa83831c3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2013</creationdate><topic>Alternative splicing</topic><topic>Antibodies</topic><topic>Apoptosis</topic><topic>Biochemistry</topic><topic>Biological activity</topic><topic>Biology</topic><topic>Blotting, Northern</topic><topic>Blotting, Western</topic><topic>Breast cancer</topic><topic>C protein</topic><topic>Cancer</topic><topic>Cell cycle</topic><topic>Cell Line, Tumor</topic><topic>Cell proliferation</topic><topic>Cell survival</topic><topic>Cells 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One</addtitle><date>2013-10-11</date><risdate>2013</risdate><volume>8</volume><issue>10</issue><spage>e77643</spage><epage>e77643</epage><pages>e77643-e77643</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Human cancers over-expressing mdm2, through a T to G variation at a single nucleotide polymorphism at position 309 (mdm2 SNP309), have functionally inactivated p53 that is not effectively degraded. They also have high expression of the alternatively spliced transcript, mdm2-C. Alternatively spliced mdm2 transcripts are expressed in many forms of human cancer and when they are exogenously expressed they transform human cells. However no study to date has detected endogenous MDM2 protein isoforms. Studies with exogenous expression of splice variants have been carried out with mdm2-A and mdm2-B, but the mdm2-C isoform has remained virtually unexplored. We addressed the cellular influence of exogenously expressed MDM2-C, and asked if endogenous MDM2-C protein was present in human cancers. To detect endogenous MDM2-C protein, we created a human MDM2-C antibody to the splice junction epitope of exons four and ten (MDM2 C410) and validated the antibody with in vitro translated full length MDM2 compared to MDM2-C. Interestingly, we discovered that MDM2-C co-migrates with MDM2-FL at approximately 98 kDa. Using the validated C410 antibody, we detected high expression of endogenous MDM2-C in human cancer cell lines and human cancer tissues. In the estrogen receptor positive (ER+) mdm2 G/G SNP309 breast cancer cell line, T47D, we observed an increase in endogenous MDM2-C protein with estrogen treatment. MDM2-C localized to the nucleus and the cytoplasm. We examined the biological activity of MDM2-C by exogenously expressing the protein and observed that MDM2-C did not efficiently target p53 for degradation or reduce p53 transcriptional activity. Exogenous expression of MDM2-C in p53-null human cancer cells increased colony formation, indicating p53-independent tumorigenic properties. Our data indicate a role for MDM2-C that does not require the inhibition of p53 for increasing cancer cell proliferation and survival.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>24147044</pmid><doi>10.1371/journal.pone.0077643</doi><tpages>e77643</tpages><oa>free_for_read</oa></addata></record> |
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| recordid | cdi_plos_journals_1441432716 |
| source | Public Library of Science (PLoS) Journals Open Access; MEDLINE; Full-Text Journals in Chemistry (Open access); DOAJ Directory of Open Access Journals; PubMed Central; EZB Electronic Journals Library |
| subjects | Alternative splicing Antibodies Apoptosis Biochemistry Biological activity Biology Blotting, Northern Blotting, Western Breast cancer C protein Cancer Cell cycle Cell Line, Tumor Cell proliferation Cell survival Cells (Biology) Cytoplasm Deoxyribonucleic acid Departments DNA Epitopes Estrogen receptors Estrogens Exons Fluorescent Antibody Technique Gene amplification Genetic aspects Genetic polymorphisms Humans Immunohistochemistry Immunoprecipitation Isoforms MDM2 protein Microscopy, Confocal Monoclonal antibodies Neoplasms - genetics Neoplasms - metabolism Nuclei p53 Protein Polymorphism Protein Isoforms - genetics Protein Isoforms - metabolism Proteins Proto-Oncogene Proteins c-mdm2 - genetics Proto-Oncogene Proteins c-mdm2 - metabolism Reverse Transcriptase Polymerase Chain Reaction Single-nucleotide polymorphism Transcription Tumor cell lines Tumor proteins Tumor Suppressor Protein p53 - genetics Tumor Suppressor Protein p53 - metabolism Tumorigenesis |
| title | Endogenous human MDM2-C is highly expressed in human cancers and functions as a p53-independent growth activator |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-27T20%3A31%3A40IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-gale_plos_&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Endogenous%20human%20MDM2-C%20is%20highly%20expressed%20in%20human%20cancers%20and%20functions%20as%20a%20p53-independent%20growth%20activator&rft.jtitle=PloS%20one&rft.au=Okoro,%20Danielle%20R&rft.date=2013-10-11&rft.volume=8&rft.issue=10&rft.spage=e77643&rft.epage=e77643&rft.pages=e77643-e77643&rft.issn=1932-6203&rft.eissn=1932-6203&rft_id=info:doi/10.1371/journal.pone.0077643&rft_dat=%3Cgale_plos_%3EA478321376%3C/gale_plos_%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=1441432716&rft_id=info:pmid/24147044&rft_galeid=A478321376&rft_doaj_id=oai_doaj_org_article_cd20aa0d90ef480fb82b0c31ef920fa5&rfr_iscdi=true |