Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos
Imprinted genes have been implicated in early embryonic, placental, and neonatal development and alterations in expression levels of these genes can lead to growth abnormalities and embryonic lethality. However, little is known about the functions of bovine imprinted genes during the pre-implantatio...
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| description | Imprinted genes have been implicated in early embryonic, placental, and neonatal development and alterations in expression levels of these genes can lead to growth abnormalities and embryonic lethality. However, little is known about the functions of bovine imprinted genes during the pre-implantation period. Therefore, the objective of this study was to assess the influence of altered expression of imprinted genes on developmental progress of embryos using small interfering RNA (siRNA). Expression levels of 18 imprinted genes (MAGEL2, UBE3A, IGF2R, NAP1L5, TSSC4, PEG3, NDN, CDKN1C, PHLDA2, MKRN3, USP29, NNAT, PEG10, RTL1, IGF2, H19, MIM1, and XIST) were compared between embryos reaching the blastocyst stage and growth-arrested embryos (degenerates) using quantitative real-time PCR (qRT-PCR). Ten genes were found to be differentially expressed between blastocysts and degenerates. The CDKN1C gene showed the highest upregulation in blastocysts whereas PHLDA2 was highly expressed in degenerates. To assess whether the observed differential gene expression was causative or resultant of embryo degeneration, these genes were selected for functional analysis using siRNA. Injection of siRNA specific to PHLDA2 into one-cell zygotes resulted in a substantial increase in blastocyst development, whereas injection of CDKN1C-specific siRNA resulted in a 45% reduction (P = 0.0006) in blastocyst development. RNA-Seq analysis of CDKN1C-siRNA-injected vs. non-injected embryos revealed 51 differentially expressed genes with functions in apoptosis, lipid metabolism, differentiation, and cell cycle regulation. Gene ontology analysis revealed nine pathways related to cell signaling, metabolism, and nucleic acid processing. Overall, our results show that proper expression levels of the imprinted genes CDKN1C and PHLDA2 are critical for embryo development, which suggests that these genes can be used as markers for normal blastocyst formation. |
| doi_str_mv | 10.1371/journal.pone.0069490 |
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However, little is known about the functions of bovine imprinted genes during the pre-implantation period. Therefore, the objective of this study was to assess the influence of altered expression of imprinted genes on developmental progress of embryos using small interfering RNA (siRNA). Expression levels of 18 imprinted genes (MAGEL2, UBE3A, IGF2R, NAP1L5, TSSC4, PEG3, NDN, CDKN1C, PHLDA2, MKRN3, USP29, NNAT, PEG10, RTL1, IGF2, H19, MIM1, and XIST) were compared between embryos reaching the blastocyst stage and growth-arrested embryos (degenerates) using quantitative real-time PCR (qRT-PCR). Ten genes were found to be differentially expressed between blastocysts and degenerates. The CDKN1C gene showed the highest upregulation in blastocysts whereas PHLDA2 was highly expressed in degenerates. To assess whether the observed differential gene expression was causative or resultant of embryo degeneration, these genes were selected for functional analysis using siRNA. Injection of siRNA specific to PHLDA2 into one-cell zygotes resulted in a substantial increase in blastocyst development, whereas injection of CDKN1C-specific siRNA resulted in a 45% reduction (P = 0.0006) in blastocyst development. RNA-Seq analysis of CDKN1C-siRNA-injected vs. non-injected embryos revealed 51 differentially expressed genes with functions in apoptosis, lipid metabolism, differentiation, and cell cycle regulation. Gene ontology analysis revealed nine pathways related to cell signaling, metabolism, and nucleic acid processing. Overall, our results show that proper expression levels of the imprinted genes CDKN1C and PHLDA2 are critical for embryo development, which suggests that these genes can be used as markers for normal blastocyst formation.</description><identifier>ISSN: 1932-6203</identifier><identifier>EISSN: 1932-6203</identifier><identifier>DOI: 10.1371/journal.pone.0069490</identifier><identifier>PMID: 23894493</identifier><language>eng</language><publisher>United States: Public Library of Science</publisher><subject>Abnormalities ; Animal sciences ; Animals ; Apoptosis ; Biology ; Blastocysts ; Cattle ; Cdkn1c protein ; Cell cycle ; Comparative analysis ; Cyclin-Dependent Kinase Inhibitor p57 - deficiency ; Cyclin-Dependent Kinase Inhibitor p57 - genetics ; Degeneration ; Developmental stages ; DNA methylation ; Embryo Implantation - genetics ; Embryogenesis ; Embryonic development ; Embryos ; Functional analysis ; Gene expression ; Gene Knockdown Techniques ; Genes ; Genomes ; Genomics ; Implantation ; Injection ; Insulin-like growth factor II ; Insulin-like growth factor II receptors ; Kinases ; Lethality ; Lipid metabolism ; Metabolism ; Neonates ; Nuclear Proteins - deficiency ; Nuclear Proteins - genetics ; Peg3 protein ; Physiological aspects ; Placenta ; Ribonucleic acid ; RNA ; RNA sequencing ; RNA, Small Interfering - genetics ; Signaling ; siRNA ; Steroids (Organic compounds) ; Success ; Transcriptome ; Ubiquitin-protein ligase ; Zygotes</subject><ispartof>PloS one, 2013-07, Vol.8 (7), p.e69490-e69490</ispartof><rights>COPYRIGHT 2013 Public Library of Science</rights><rights>2013 Driver 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 Driver et al 2013 Driver et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c593t-d160ec1c9c10cd7d15ba29122b38e2f4f1c4ba81a722e37f9e9c025f0762e9113</citedby><cites>FETCH-LOGICAL-c593t-d160ec1c9c10cd7d15ba29122b38e2f4f1c4ba81a722e37f9e9c025f0762e9113</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/PMC3718760/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3718760/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,864,885,2102,2928,23866,27924,27925,53791,53793,79472,79473</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/23894493$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Wrenzycki, Christine</contributor><creatorcontrib>Driver, Ashley M</creatorcontrib><creatorcontrib>Huang, Wen</creatorcontrib><creatorcontrib>Kropp, Jenna</creatorcontrib><creatorcontrib>Peñagaricano, Francisco</creatorcontrib><creatorcontrib>Khatib, Hasan</creatorcontrib><title>Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos</title><title>PloS one</title><addtitle>PLoS One</addtitle><description>Imprinted genes have been implicated in early embryonic, placental, and neonatal development and alterations in expression levels of these genes can lead to growth abnormalities and embryonic lethality. However, little is known about the functions of bovine imprinted genes during the pre-implantation period. Therefore, the objective of this study was to assess the influence of altered expression of imprinted genes on developmental progress of embryos using small interfering RNA (siRNA). Expression levels of 18 imprinted genes (MAGEL2, UBE3A, IGF2R, NAP1L5, TSSC4, PEG3, NDN, CDKN1C, PHLDA2, MKRN3, USP29, NNAT, PEG10, RTL1, IGF2, H19, MIM1, and XIST) were compared between embryos reaching the blastocyst stage and growth-arrested embryos (degenerates) using quantitative real-time PCR (qRT-PCR). Ten genes were found to be differentially expressed between blastocysts and degenerates. The CDKN1C gene showed the highest upregulation in blastocysts whereas PHLDA2 was highly expressed in degenerates. To assess whether the observed differential gene expression was causative or resultant of embryo degeneration, these genes were selected for functional analysis using siRNA. Injection of siRNA specific to PHLDA2 into one-cell zygotes resulted in a substantial increase in blastocyst development, whereas injection of CDKN1C-specific siRNA resulted in a 45% reduction (P = 0.0006) in blastocyst development. RNA-Seq analysis of CDKN1C-siRNA-injected vs. non-injected embryos revealed 51 differentially expressed genes with functions in apoptosis, lipid metabolism, differentiation, and cell cycle regulation. Gene ontology analysis revealed nine pathways related to cell signaling, metabolism, and nucleic acid processing. Overall, our results show that proper expression levels of the imprinted genes CDKN1C and PHLDA2 are critical for embryo development, which suggests that these genes can be used as markers for normal blastocyst formation.</description><subject>Abnormalities</subject><subject>Animal sciences</subject><subject>Animals</subject><subject>Apoptosis</subject><subject>Biology</subject><subject>Blastocysts</subject><subject>Cattle</subject><subject>Cdkn1c protein</subject><subject>Cell cycle</subject><subject>Comparative analysis</subject><subject>Cyclin-Dependent Kinase Inhibitor p57 - deficiency</subject><subject>Cyclin-Dependent Kinase Inhibitor p57 - genetics</subject><subject>Degeneration</subject><subject>Developmental stages</subject><subject>DNA methylation</subject><subject>Embryo Implantation - genetics</subject><subject>Embryogenesis</subject><subject>Embryonic development</subject><subject>Embryos</subject><subject>Functional analysis</subject><subject>Gene expression</subject><subject>Gene Knockdown Techniques</subject><subject>Genes</subject><subject>Genomes</subject><subject>Genomics</subject><subject>Implantation</subject><subject>Injection</subject><subject>Insulin-like growth factor II</subject><subject>Insulin-like growth factor II receptors</subject><subject>Kinases</subject><subject>Lethality</subject><subject>Lipid metabolism</subject><subject>Metabolism</subject><subject>Neonates</subject><subject>Nuclear Proteins - deficiency</subject><subject>Nuclear Proteins - genetics</subject><subject>Peg3 protein</subject><subject>Physiological aspects</subject><subject>Placenta</subject><subject>Ribonucleic acid</subject><subject>RNA</subject><subject>RNA sequencing</subject><subject>RNA, Small Interfering - genetics</subject><subject>Signaling</subject><subject>siRNA</subject><subject>Steroids (Organic compounds)</subject><subject>Success</subject><subject>Transcriptome</subject><subject>Ubiquitin-protein ligase</subject><subject>Zygotes</subject><issn>1932-6203</issn><issn>1932-6203</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2013</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>DOA</sourceid><recordid>eNptUk1vEzEQXSEQLYV_gMASl_SQ4K9dry9IUQq0agQc4Gx5vbOp0117a2-C-u9xkm3VoMoHWzNv3sw8vyx7T_CMMEE-r_0mON3Oeu9ghnEhucQvslMiGZ0WFLOXT94n2ZsY1xjnrCyK19kJZaXkXLLTzF07b25r_9ch36DFxfUPskCTPheTW9vT83OkXY1-XS4v5hQFiJt2iMg6VMMWWt934AbdInOj3Qr2icpvrQPUB5jarm91yg_WOwRdFe59fJu9anQb4d14n2V_vn39vbicLn9-v1rMl1OTSzZMa1JgMMRIQ7CpRU3ySlNJKK1YCbThDTG80iXRglJgopEgDaZ5g0VBQRLCzrKPB96-9VGNUkVFOBVMMMJZQlwdELXXa9UH2-lwr7y2ah_wYaV0GKxpQdUcSwFlaptLjjXWAoSssJYF4yXfc30Zu22qDmqTVAm6PSI9zjh7o1Z-q9I_lqLAiWAyEgR_t4E4qM5GA23SD_xmNzfJC5wXuUjQT_9Bn99uRK10WsC6xqe-Zkeq5lyUpBCYlgk1ewaVTg2dNclXjU3xowJ-KDDBxxigedyRYLVz5cMwaudKNboylX14qs9j0YMN2T-xTdzO</recordid><startdate>20130722</startdate><enddate>20130722</enddate><creator>Driver, Ashley M</creator><creator>Huang, Wen</creator><creator>Kropp, Jenna</creator><creator>Peñagaricano, Francisco</creator><creator>Khatib, Hasan</creator><general>Public Library of Science</general><general>Public Library of Science (PLoS)</general><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>3V.</scope><scope>7QG</scope><scope>7QL</scope><scope>7QO</scope><scope>7RV</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TG</scope><scope>7TM</scope><scope>7U9</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8AO</scope><scope>8C1</scope><scope>8FD</scope><scope>8FE</scope><scope>8FG</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABJCF</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>ARAPS</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BGLVJ</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>D1I</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>H94</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>KB.</scope><scope>KB0</scope><scope>KL.</scope><scope>L6V</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M7N</scope><scope>M7P</scope><scope>M7S</scope><scope>NAPCQ</scope><scope>P5Z</scope><scope>P62</scope><scope>P64</scope><scope>PATMY</scope><scope>PDBOC</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>PTHSS</scope><scope>PYCSY</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope></search><sort><creationdate>20130722</creationdate><title>Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos</title><author>Driver, Ashley M ; Huang, Wen ; Kropp, Jenna ; Peñagaricano, Francisco ; Khatib, Hasan</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c593t-d160ec1c9c10cd7d15ba29122b38e2f4f1c4ba81a722e37f9e9c025f0762e9113</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2013</creationdate><topic>Abnormalities</topic><topic>Animal sciences</topic><topic>Animals</topic><topic>Apoptosis</topic><topic>Biology</topic><topic>Blastocysts</topic><topic>Cattle</topic><topic>Cdkn1c protein</topic><topic>Cell cycle</topic><topic>Comparative analysis</topic><topic>Cyclin-Dependent Kinase Inhibitor p57 - deficiency</topic><topic>Cyclin-Dependent Kinase Inhibitor p57 - genetics</topic><topic>Degeneration</topic><topic>Developmental stages</topic><topic>DNA methylation</topic><topic>Embryo Implantation - genetics</topic><topic>Embryogenesis</topic><topic>Embryonic development</topic><topic>Embryos</topic><topic>Functional analysis</topic><topic>Gene expression</topic><topic>Gene Knockdown Techniques</topic><topic>Genes</topic><topic>Genomes</topic><topic>Genomics</topic><topic>Implantation</topic><topic>Injection</topic><topic>Insulin-like growth factor II</topic><topic>Insulin-like growth factor II receptors</topic><topic>Kinases</topic><topic>Lethality</topic><topic>Lipid metabolism</topic><topic>Metabolism</topic><topic>Neonates</topic><topic>Nuclear Proteins - deficiency</topic><topic>Nuclear Proteins - genetics</topic><topic>Peg3 protein</topic><topic>Physiological aspects</topic><topic>Placenta</topic><topic>Ribonucleic acid</topic><topic>RNA</topic><topic>RNA sequencing</topic><topic>RNA, Small Interfering - 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>DOAJ Directory of Open Access Journals</collection><jtitle>PloS one</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Driver, Ashley M</au><au>Huang, Wen</au><au>Kropp, Jenna</au><au>Peñagaricano, Francisco</au><au>Khatib, Hasan</au><au>Wrenzycki, Christine</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos</atitle><jtitle>PloS one</jtitle><addtitle>PLoS One</addtitle><date>2013-07-22</date><risdate>2013</risdate><volume>8</volume><issue>7</issue><spage>e69490</spage><epage>e69490</epage><pages>e69490-e69490</pages><issn>1932-6203</issn><eissn>1932-6203</eissn><abstract>Imprinted genes have been implicated in early embryonic, placental, and neonatal development and alterations in expression levels of these genes can lead to growth abnormalities and embryonic lethality. However, little is known about the functions of bovine imprinted genes during the pre-implantation period. Therefore, the objective of this study was to assess the influence of altered expression of imprinted genes on developmental progress of embryos using small interfering RNA (siRNA). Expression levels of 18 imprinted genes (MAGEL2, UBE3A, IGF2R, NAP1L5, TSSC4, PEG3, NDN, CDKN1C, PHLDA2, MKRN3, USP29, NNAT, PEG10, RTL1, IGF2, H19, MIM1, and XIST) were compared between embryos reaching the blastocyst stage and growth-arrested embryos (degenerates) using quantitative real-time PCR (qRT-PCR). Ten genes were found to be differentially expressed between blastocysts and degenerates. The CDKN1C gene showed the highest upregulation in blastocysts whereas PHLDA2 was highly expressed in degenerates. To assess whether the observed differential gene expression was causative or resultant of embryo degeneration, these genes were selected for functional analysis using siRNA. Injection of siRNA specific to PHLDA2 into one-cell zygotes resulted in a substantial increase in blastocyst development, whereas injection of CDKN1C-specific siRNA resulted in a 45% reduction (P = 0.0006) in blastocyst development. RNA-Seq analysis of CDKN1C-siRNA-injected vs. non-injected embryos revealed 51 differentially expressed genes with functions in apoptosis, lipid metabolism, differentiation, and cell cycle regulation. Gene ontology analysis revealed nine pathways related to cell signaling, metabolism, and nucleic acid processing. Overall, our results show that proper expression levels of the imprinted genes CDKN1C and PHLDA2 are critical for embryo development, which suggests that these genes can be used as markers for normal blastocyst formation.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>23894493</pmid><doi>10.1371/journal.pone.0069490</doi><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; DOAJ Directory of Open Access Journals; Public Library of Science (PLoS); EZB-FREE-00999 freely available EZB journals; PubMed Central; Free Full-Text Journals in Chemistry |
| subjects | Abnormalities Animal sciences Animals Apoptosis Biology Blastocysts Cattle Cdkn1c protein Cell cycle Comparative analysis Cyclin-Dependent Kinase Inhibitor p57 - deficiency Cyclin-Dependent Kinase Inhibitor p57 - genetics Degeneration Developmental stages DNA methylation Embryo Implantation - genetics Embryogenesis Embryonic development Embryos Functional analysis Gene expression Gene Knockdown Techniques Genes Genomes Genomics Implantation Injection Insulin-like growth factor II Insulin-like growth factor II receptors Kinases Lethality Lipid metabolism Metabolism Neonates Nuclear Proteins - deficiency Nuclear Proteins - genetics Peg3 protein Physiological aspects Placenta Ribonucleic acid RNA RNA sequencing RNA, Small Interfering - genetics Signaling siRNA Steroids (Organic compounds) Success Transcriptome Ubiquitin-protein ligase Zygotes |
| title | Knockdown of CDKN1C (p57(kip2)) and PHLDA2 results in developmental changes in bovine pre-implantation embryos |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-08T00%3A08%3A29IST&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=Knockdown%20of%20CDKN1C%20(p57(kip2))%20and%20PHLDA2%20results%20in%20developmental%20changes%20in%20bovine%20pre-implantation%20embryos&rft.jtitle=PloS%20one&rft.au=Driver,%20Ashley%20M&rft.date=2013-07-22&rft.volume=8&rft.issue=7&rft.spage=e69490&rft.epage=e69490&rft.pages=e69490-e69490&rft.issn=1932-6203&rft.eissn=1932-6203&rft_id=info:doi/10.1371/journal.pone.0069490&rft_dat=%3Cgale_plos_%3EA478167028%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=1427373143&rft_id=info:pmid/23894493&rft_galeid=A478167028&rft_doaj_id=oai_doaj_org_article_d4097e8f4f5940a0a7e79b0a96348443&rfr_iscdi=true |