RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis
Mammalian spermatogenesis involves formation of haploid cells from the male germline and then a complex morphological transformation to generate motile sperm. Focusing on meiotic prophase, some tissue-specific transcription factors are known (A-MYB) or suspected (RFX2) to play important roles in mod...
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description | Mammalian spermatogenesis involves formation of haploid cells from the male germline and then a complex morphological transformation to generate motile sperm. Focusing on meiotic prophase, some tissue-specific transcription factors are known (A-MYB) or suspected (RFX2) to play important roles in modulating gene expression in pachytene spermatocytes. The current work was initiated to identify both downstream and upstream regulatory connections for Rfx2.
Searches of pachytene up-regulated genes identified high affinity RFX binding sites (X boxes) in promoter regions of several new genes: Adam5, Pdcl2, and Spag6. We confirmed a strong promoter-region X-box for Alf, a germ cell-specific variant of general transcription factor TFIIA. Using Alf as an example of a target gene, we showed that its promoter is stimulated by RFX2 in transfected cells and used ChIP analysis to show that the promoter is occupied by RFX2 in vivo. Turning to upstream regulation of the Rfx2 promoter, we identified a cluster of three binding sites (MBS) for the MYB family of transcription factors. Because testis is one of the few sites of A-myb expression, and because spermatogenesis arrests in pachytene in A-myb knockout mice, the MBS cluster implicates Rfx2 as an A-myb target. Electrophoretic gel-shift, ChIP, and co-transfection assays all support a role for these MYB sites in Rfx2 expression. Further, Rfx2 expression was virtually eliminated in A-myb knockout testes. Immunohistology on testis sections showed that A-MYB expression is up-regulated only after pachytene spermatocytes have clearly moved away from the tubule wall, which correlates with onset of RFX2 expression, whereas B-MYB expression, by contrast, is prevalent only in earlier spermatocytes and spermatogonia.
With an expanding list of likely target genes, RFX2 is potentially an important transcriptional regulator in pachytene spermatocytes. Rfx2 itself is a good candidate to be regulated by A-MYB, which is essential for meiotic progression. If Alf is a genuine RFX2 target, then A-myb, Rfx2, and Alf may form part of a transcriptional network that is vital for completion of meiosis and preparation for post-meiotic differentiation. |
doi_str_mv | 10.1186/1471-213X-9-63 |
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Searches of pachytene up-regulated genes identified high affinity RFX binding sites (X boxes) in promoter regions of several new genes: Adam5, Pdcl2, and Spag6. We confirmed a strong promoter-region X-box for Alf, a germ cell-specific variant of general transcription factor TFIIA. Using Alf as an example of a target gene, we showed that its promoter is stimulated by RFX2 in transfected cells and used ChIP analysis to show that the promoter is occupied by RFX2 in vivo. Turning to upstream regulation of the Rfx2 promoter, we identified a cluster of three binding sites (MBS) for the MYB family of transcription factors. Because testis is one of the few sites of A-myb expression, and because spermatogenesis arrests in pachytene in A-myb knockout mice, the MBS cluster implicates Rfx2 as an A-myb target. Electrophoretic gel-shift, ChIP, and co-transfection assays all support a role for these MYB sites in Rfx2 expression. Further, Rfx2 expression was virtually eliminated in A-myb knockout testes. Immunohistology on testis sections showed that A-MYB expression is up-regulated only after pachytene spermatocytes have clearly moved away from the tubule wall, which correlates with onset of RFX2 expression, whereas B-MYB expression, by contrast, is prevalent only in earlier spermatocytes and spermatogonia.
With an expanding list of likely target genes, RFX2 is potentially an important transcriptional regulator in pachytene spermatocytes. Rfx2 itself is a good candidate to be regulated by A-MYB, which is essential for meiotic progression. If Alf is a genuine RFX2 target, then A-myb, Rfx2, and Alf may form part of a transcriptional network that is vital for completion of meiosis and preparation for post-meiotic differentiation.</description><identifier>ISSN: 1471-213X</identifier><identifier>EISSN: 1471-213X</identifier><identifier>DOI: 10.1186/1471-213X-9-63</identifier><identifier>PMID: 20003220</identifier><language>eng</language><publisher>England: BioMed Central Ltd</publisher><subject>Animals ; Base Sequence ; Boxes ; DNA binding proteins ; DNA-Binding Proteins - genetics ; DNA-Binding Proteins - metabolism ; Experiments ; Gene expression ; Genes ; Genetic aspects ; Genetics ; Kinases ; Life sciences ; Male ; Mice ; Mice, Knockout ; Molecular Sequence Data ; Physiological aspects ; Plasmids ; Promoter Regions, Genetic ; Proto-Oncogene Proteins c-myb - metabolism ; Regulation ; Regulatory Factor X Transcription Factors ; Research article ; Sperm ; Spermatogenesis ; Stem cells ; Testis - metabolism ; Trans-Activators - metabolism ; Transcription Factors - genetics ; Transcription Factors - metabolism</subject><ispartof>BMC developmental biology, 2009-12, Vol.9 (1), p.63-63, Article 63</ispartof><rights>COPYRIGHT 2009 BioMed Central Ltd.</rights><rights>2009 Horvath et al; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.</rights><rights>Copyright ©2009 Horvath et al; licensee BioMed Central Ltd. 2009 Horvath et al; licensee BioMed Central Ltd.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-b613t-a46f10bb682c41f87b3420d29b43fa13818fea8fd20d2bc4d7343e2808265e553</citedby><cites>FETCH-LOGICAL-b613t-a46f10bb682c41f87b3420d29b43fa13818fea8fd20d2bc4d7343e2808265e553</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/PMC2797782/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC2797782/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,24781,27903,27904,53770,53772,75485,75486</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/20003220$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Horvath, Gary C</creatorcontrib><creatorcontrib>Kistler, Malathi K</creatorcontrib><creatorcontrib>Kistler, W Stephen</creatorcontrib><title>RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis</title><title>BMC developmental biology</title><addtitle>BMC Dev Biol</addtitle><description>Mammalian spermatogenesis involves formation of haploid cells from the male germline and then a complex morphological transformation to generate motile sperm. Focusing on meiotic prophase, some tissue-specific transcription factors are known (A-MYB) or suspected (RFX2) to play important roles in modulating gene expression in pachytene spermatocytes. The current work was initiated to identify both downstream and upstream regulatory connections for Rfx2.
Searches of pachytene up-regulated genes identified high affinity RFX binding sites (X boxes) in promoter regions of several new genes: Adam5, Pdcl2, and Spag6. We confirmed a strong promoter-region X-box for Alf, a germ cell-specific variant of general transcription factor TFIIA. Using Alf as an example of a target gene, we showed that its promoter is stimulated by RFX2 in transfected cells and used ChIP analysis to show that the promoter is occupied by RFX2 in vivo. Turning to upstream regulation of the Rfx2 promoter, we identified a cluster of three binding sites (MBS) for the MYB family of transcription factors. Because testis is one of the few sites of A-myb expression, and because spermatogenesis arrests in pachytene in A-myb knockout mice, the MBS cluster implicates Rfx2 as an A-myb target. Electrophoretic gel-shift, ChIP, and co-transfection assays all support a role for these MYB sites in Rfx2 expression. Further, Rfx2 expression was virtually eliminated in A-myb knockout testes. Immunohistology on testis sections showed that A-MYB expression is up-regulated only after pachytene spermatocytes have clearly moved away from the tubule wall, which correlates with onset of RFX2 expression, whereas B-MYB expression, by contrast, is prevalent only in earlier spermatocytes and spermatogonia.
With an expanding list of likely target genes, RFX2 is potentially an important transcriptional regulator in pachytene spermatocytes. Rfx2 itself is a good candidate to be regulated by A-MYB, which is essential for meiotic progression. If Alf is a genuine RFX2 target, then A-myb, Rfx2, and Alf may form part of a transcriptional network that is vital for completion of meiosis and preparation for post-meiotic differentiation.</description><subject>Animals</subject><subject>Base Sequence</subject><subject>Boxes</subject><subject>DNA binding proteins</subject><subject>DNA-Binding Proteins - genetics</subject><subject>DNA-Binding Proteins - metabolism</subject><subject>Experiments</subject><subject>Gene expression</subject><subject>Genes</subject><subject>Genetic aspects</subject><subject>Genetics</subject><subject>Kinases</subject><subject>Life sciences</subject><subject>Male</subject><subject>Mice</subject><subject>Mice, Knockout</subject><subject>Molecular Sequence Data</subject><subject>Physiological aspects</subject><subject>Plasmids</subject><subject>Promoter Regions, Genetic</subject><subject>Proto-Oncogene Proteins c-myb - metabolism</subject><subject>Regulation</subject><subject>Regulatory Factor X Transcription Factors</subject><subject>Research article</subject><subject>Sperm</subject><subject>Spermatogenesis</subject><subject>Stem cells</subject><subject>Testis - metabolism</subject><subject>Trans-Activators - metabolism</subject><subject>Transcription Factors - genetics</subject><subject>Transcription Factors - metabolism</subject><issn>1471-213X</issn><issn>1471-213X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2009</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><recordid>eNp1Uk1v1DAQtRCIlsKVI7K4oB5S_JE4zgVpqVqoVIRUqFROluOMg6vE3toJH_8eh11WXRDywdabN29mngeh55ScUCrFa1rWtGCU3xRNIfgDdLgDHt57H6AnKd0SQmtJxWN0wAghnDFyiK6vzm8YdglrbLTvXKcnwF347tMUQY9Yj-vBWQcRB4tXxYcvb3GEfh705ILHzuMxzAlwWkMc9RR68JBceooeWT0keLa9j9D1-dnn0_fF5cd3F6ery6IVlE-FLoWlpG2FZKakVtYtLxnpWNOW3GrKJZUWtLTdAram7GpecmCSSCYqqCp-hN5sdNdzO0JnwE9RD2od3ajjTxW0U_sR776qPnxTrG7qWrIssNoItC78R2A_YsKoFlvVYqtqlOBZ49W2iRjuZkiTGl0yMAzaQzZH5aYZLcnvdl_-xbwNc_TZIdWQ_CeiYgvpZEPq9QDKeRtyYZNPB6MzwYN1GV8xWtGG1ULmhOO9hMyZ4MfU6zkldfHpap-7FTcxpBTB7kalRC0L9e9wL-47vKP_2SD-Cw3fxU0</recordid><startdate>20091209</startdate><enddate>20091209</enddate><creator>Horvath, Gary C</creator><creator>Kistler, Malathi K</creator><creator>Kistler, W Stephen</creator><general>BioMed Central Ltd</general><general>BioMed Central</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>ISR</scope><scope>3V.</scope><scope>7SS</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M7P</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20091209</creationdate><title>RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis</title><author>Horvath, Gary C ; Kistler, Malathi K ; Kistler, W Stephen</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-b613t-a46f10bb682c41f87b3420d29b43fa13818fea8fd20d2bc4d7343e2808265e553</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2009</creationdate><topic>Animals</topic><topic>Base Sequence</topic><topic>Boxes</topic><topic>DNA binding proteins</topic><topic>DNA-Binding Proteins - genetics</topic><topic>DNA-Binding Proteins - metabolism</topic><topic>Experiments</topic><topic>Gene expression</topic><topic>Genes</topic><topic>Genetic aspects</topic><topic>Genetics</topic><topic>Kinases</topic><topic>Life sciences</topic><topic>Male</topic><topic>Mice</topic><topic>Mice, Knockout</topic><topic>Molecular Sequence Data</topic><topic>Physiological aspects</topic><topic>Plasmids</topic><topic>Promoter Regions, Genetic</topic><topic>Proto-Oncogene Proteins c-myb - metabolism</topic><topic>Regulation</topic><topic>Regulatory Factor X Transcription Factors</topic><topic>Research article</topic><topic>Sperm</topic><topic>Spermatogenesis</topic><topic>Stem cells</topic><topic>Testis - metabolism</topic><topic>Trans-Activators - metabolism</topic><topic>Transcription Factors - genetics</topic><topic>Transcription Factors - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Horvath, Gary C</creatorcontrib><creatorcontrib>Kistler, Malathi K</creatorcontrib><creatorcontrib>Kistler, W Stephen</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Gale In Context: Science</collection><collection>ProQuest Central (Corporate)</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Health & Medical Collection (Proquest)</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>ProQuest Central (Alumni)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>ProQuest Natural Science Collection</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>PML(ProQuest Medical Library)</collection><collection>ProQuest Biological Science Journals</collection><collection>Publicly Available Content Database</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central China</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>BMC developmental biology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Horvath, Gary C</au><au>Kistler, Malathi K</au><au>Kistler, W Stephen</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis</atitle><jtitle>BMC developmental biology</jtitle><addtitle>BMC Dev Biol</addtitle><date>2009-12-09</date><risdate>2009</risdate><volume>9</volume><issue>1</issue><spage>63</spage><epage>63</epage><pages>63-63</pages><artnum>63</artnum><issn>1471-213X</issn><eissn>1471-213X</eissn><abstract>Mammalian spermatogenesis involves formation of haploid cells from the male germline and then a complex morphological transformation to generate motile sperm. Focusing on meiotic prophase, some tissue-specific transcription factors are known (A-MYB) or suspected (RFX2) to play important roles in modulating gene expression in pachytene spermatocytes. The current work was initiated to identify both downstream and upstream regulatory connections for Rfx2.
Searches of pachytene up-regulated genes identified high affinity RFX binding sites (X boxes) in promoter regions of several new genes: Adam5, Pdcl2, and Spag6. We confirmed a strong promoter-region X-box for Alf, a germ cell-specific variant of general transcription factor TFIIA. Using Alf as an example of a target gene, we showed that its promoter is stimulated by RFX2 in transfected cells and used ChIP analysis to show that the promoter is occupied by RFX2 in vivo. Turning to upstream regulation of the Rfx2 promoter, we identified a cluster of three binding sites (MBS) for the MYB family of transcription factors. Because testis is one of the few sites of A-myb expression, and because spermatogenesis arrests in pachytene in A-myb knockout mice, the MBS cluster implicates Rfx2 as an A-myb target. Electrophoretic gel-shift, ChIP, and co-transfection assays all support a role for these MYB sites in Rfx2 expression. Further, Rfx2 expression was virtually eliminated in A-myb knockout testes. Immunohistology on testis sections showed that A-MYB expression is up-regulated only after pachytene spermatocytes have clearly moved away from the tubule wall, which correlates with onset of RFX2 expression, whereas B-MYB expression, by contrast, is prevalent only in earlier spermatocytes and spermatogonia.
With an expanding list of likely target genes, RFX2 is potentially an important transcriptional regulator in pachytene spermatocytes. Rfx2 itself is a good candidate to be regulated by A-MYB, which is essential for meiotic progression. If Alf is a genuine RFX2 target, then A-myb, Rfx2, and Alf may form part of a transcriptional network that is vital for completion of meiosis and preparation for post-meiotic differentiation.</abstract><cop>England</cop><pub>BioMed Central Ltd</pub><pmid>20003220</pmid><doi>10.1186/1471-213X-9-63</doi><oa>free_for_read</oa></addata></record> |
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subjects | Animals Base Sequence Boxes DNA binding proteins DNA-Binding Proteins - genetics DNA-Binding Proteins - metabolism Experiments Gene expression Genes Genetic aspects Genetics Kinases Life sciences Male Mice Mice, Knockout Molecular Sequence Data Physiological aspects Plasmids Promoter Regions, Genetic Proto-Oncogene Proteins c-myb - metabolism Regulation Regulatory Factor X Transcription Factors Research article Sperm Spermatogenesis Stem cells Testis - metabolism Trans-Activators - metabolism Transcription Factors - genetics Transcription Factors - metabolism |
title | RFX2 is a candidate downstream amplifier of A-MYB regulation in mouse spermatogenesis |
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