Characterization and Function of 3-Hydroxy-3-Methylglutaryl-CoA Reductase in Populus trichocarpa: Overexpression of PtHMGR Enhances Terpenoids in Transgenic Poplar.
Identifieur interne : 000A89 ( Main/Exploration ); précédent : 000A88; suivant : 000A90Characterization and Function of 3-Hydroxy-3-Methylglutaryl-CoA Reductase in Populus trichocarpa: Overexpression of PtHMGR Enhances Terpenoids in Transgenic Poplar.
Auteurs : Hui Wei [République populaire de Chine] ; Chen Xu [République populaire de Chine] ; Ali Movahedi [République populaire de Chine] ; Weibo Sun [République populaire de Chine] ; Dawei Li [République populaire de Chine] ; Qiang Zhuge [République populaire de Chine]Source :
- Frontiers in plant science [ 1664-462X ] ; 2019.
Abstract
In the mevalonic acid (MVA) pathway, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is considered the first rate-limiting enzyme in isoprenoid biosynthesis. In this study, we cloned a full-length cDNA from Populus trichocarpa with an open reading frame of 1,734 bp. The deduced PtHMGR sequence contained two HMG-CoA motifs and two NADPH motifs, which exhibited homology with HMGR proteins from other species. Subsequently, truncated PtHMGR was expressed in Escherichia coli BL21 (DE3) cells, and enzyme activity analysis revealed that the truncated PtHMGR protein could catalyze the reaction of HMG-CoA and NADPH to form MVA. Relative expression analysis suggests that PtHMGR expression varies among tissues and that PtHMGR responds significantly to abscisic acid (ABA), NaCl, PEG6000, hydrogen peroxide (H2O2), and cold stresses. We used polymerase chain reaction (PCR) analysis to select transgenic Nanlin 895 poplars (Populus× euramericana cv.) and quantitative reverse-transcription PCR (qRT-PCR) to show that PtHMGR expression levels were 3- to 10-fold higher in transgenic lines than in wild-type (WT) poplars. qRT-PCR was also used to determine transcript levels of methylerythritol phosphate (MEP)-, MVA-, and downstream-related genes, indicating that overexpression of PtHMGR not only affects expression levels of MVA-related genes, but also those of MEP-related genes. We also measured the content of terpenoids including ABA, gibberellic acid (GA), carotenes, and lycopene. PtHMGR overexpression significantly increased ABA, GA, carotene, and lycopene content, indicating that PtHMGR participates in the regulation of terpenoid compound synthesis.
DOI: 10.3389/fpls.2019.01476
PubMed: 31803212
PubMed Central: PMC6872958
Affiliations:
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Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Xu, Chen" sort="Xu, Chen" uniqKey="Xu C" first="Chen" last="Xu">Chen Xu</name><affiliation wicri:level="1"><nlm:affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation><affiliation wicri:level="1"><nlm:affiliation>Jiangsu Provincial Key Construction Laboratory of Special Biomass Resource Utilization, Nanjing Xiaozhuang University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Jiangsu Provincial Key Construction Laboratory of Special Biomass Resource Utilization, Nanjing Xiaozhuang University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Movahedi, Ali" sort="Movahedi, Ali" uniqKey="Movahedi A" first="Ali" last="Movahedi">Ali Movahedi</name><affiliation wicri:level="1"><nlm:affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Sun, Weibo" sort="Sun, Weibo" uniqKey="Sun W" first="Weibo" last="Sun">Weibo Sun</name><affiliation wicri:level="1"><nlm:affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Li, Dawei" sort="Li, Dawei" uniqKey="Li D" first="Dawei" last="Li">Dawei Li</name><affiliation wicri:level="1"><nlm:affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Zhuge, Qiang" sort="Zhuge, Qiang" uniqKey="Zhuge Q" first="Qiang" last="Zhuge">Qiang Zhuge</name><affiliation wicri:level="1"><nlm:affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author></analytic><series><title level="j">Frontiers in plant science</title><idno type="ISSN">1664-462X</idno><imprint><date when="2019" type="published">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">In the mevalonic acid (MVA) pathway, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is considered the first rate-limiting enzyme in isoprenoid biosynthesis. In this study, we cloned a full-length cDNA from <i>Populus trichocarpa</i> with an open reading frame of 1,734 bp. The deduced PtHMGR sequence contained two HMG-CoA motifs and two NADPH motifs, which exhibited homology with HMGR proteins from other species. Subsequently, truncated PtHMGR was expressed in <i>Escherichia coli</i> BL21 (DE3) cells, and enzyme activity analysis revealed that the truncated PtHMGR protein could catalyze the reaction of HMG-CoA and NADPH to form MVA. Relative expression analysis suggests that <i>PtHMGR</i> expression varies among tissues and that <i>PtHMGR</i> responds significantly to abscisic acid (ABA), NaCl, PEG<sub>6000</sub>, hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>), and cold stresses. We used polymerase chain reaction (PCR) analysis to select transgenic Nanlin 895 poplars (<i>Populus× euramericana</i> cv.) and quantitative reverse-transcription PCR (qRT-PCR) to show that <i>PtHMGR</i> expression levels were 3- to 10-fold higher in transgenic lines than in wild-type (WT) poplars. qRT-PCR was also used to determine transcript levels of methylerythritol phosphate (MEP)-, MVA-, and downstream-related genes, indicating that overexpression of <i>PtHMGR</i> not only affects expression levels of MVA-related genes, but also those of MEP-related genes. We also measured the content of terpenoids including ABA, gibberellic acid (GA), carotenes, and lycopene. <i>PtHMGR</i> overexpression significantly increased ABA, GA, carotene, and lycopene content, indicating that PtHMGR participates in the regulation of terpenoid compound synthesis.</div></front></TEI><pubmed><MedlineCitation Status="PubMed-not-MEDLINE" Owner="NLM"><PMID Version="1">31803212</PMID><DateRevised><Year>2020</Year><Month>10</Month><Day>01</Day></DateRevised><Article PubModel="Electronic-eCollection"><Journal><ISSN IssnType="Print">1664-462X</ISSN><JournalIssue CitedMedium="Print"><Volume>10</Volume><PubDate><Year>2019</Year></PubDate></JournalIssue><Title>Frontiers in plant science</Title><ISOAbbreviation>Front Plant Sci</ISOAbbreviation></Journal><ArticleTitle>Characterization and Function of 3-Hydroxy-3-Methylglutaryl-CoA Reductase in <i>Populus trichocarpa</i>: Overexpression of <i>PtHMGR</i> Enhances Terpenoids in Transgenic Poplar.</ArticleTitle><Pagination><MedlinePgn>1476</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.3389/fpls.2019.01476</ELocationID><Abstract><AbstractText>In the mevalonic acid (MVA) pathway, 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is considered the first rate-limiting enzyme in isoprenoid biosynthesis. In this study, we cloned a full-length cDNA from <i>Populus trichocarpa</i> with an open reading frame of 1,734 bp. The deduced PtHMGR sequence contained two HMG-CoA motifs and two NADPH motifs, which exhibited homology with HMGR proteins from other species. Subsequently, truncated PtHMGR was expressed in <i>Escherichia coli</i> BL21 (DE3) cells, and enzyme activity analysis revealed that the truncated PtHMGR protein could catalyze the reaction of HMG-CoA and NADPH to form MVA. Relative expression analysis suggests that <i>PtHMGR</i> expression varies among tissues and that <i>PtHMGR</i> responds significantly to abscisic acid (ABA), NaCl, PEG<sub>6000</sub>, hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>), and cold stresses. We used polymerase chain reaction (PCR) analysis to select transgenic Nanlin 895 poplars (<i>Populus× euramericana</i> cv.) and quantitative reverse-transcription PCR (qRT-PCR) to show that <i>PtHMGR</i> expression levels were 3- to 10-fold higher in transgenic lines than in wild-type (WT) poplars. qRT-PCR was also used to determine transcript levels of methylerythritol phosphate (MEP)-, MVA-, and downstream-related genes, indicating that overexpression of <i>PtHMGR</i> not only affects expression levels of MVA-related genes, but also those of MEP-related genes. We also measured the content of terpenoids including ABA, gibberellic acid (GA), carotenes, and lycopene. <i>PtHMGR</i> overexpression significantly increased ABA, GA, carotene, and lycopene content, indicating that PtHMGR participates in the regulation of terpenoid compound synthesis.</AbstractText><CopyrightInformation>Copyright © 2019 Wei, Xu, Movahedi, Sun, Li and Zhuge.</CopyrightInformation></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Wei</LastName><ForeName>Hui</ForeName><Initials>H</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Xu</LastName><ForeName>Chen</ForeName><Initials>C</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo><AffiliationInfo><Affiliation>Jiangsu Provincial Key Construction Laboratory of Special Biomass Resource Utilization, Nanjing Xiaozhuang University, Nanjing, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Movahedi</LastName><ForeName>Ali</ForeName><Initials>A</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Sun</LastName><ForeName>Weibo</ForeName><Initials>W</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Li</LastName><ForeName>Dawei</ForeName><Initials>D</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Zhuge</LastName><ForeName>Qiang</ForeName><Initials>Q</Initials><AffiliationInfo><Affiliation>Co-Innovation Center for Sustainable Forestry in Southern China, Key Laboratory of Forest Genetics & Biotechnology, Ministry of Education, College of Biology and the Environment, Nanjing Forestry University, Nanjing, China.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2019</Year><Month>11</Month><Day>15</Day></ArticleDate></Article><MedlineJournalInfo><Country>Switzerland</Country><MedlineTA>Front Plant Sci</MedlineTA><NlmUniqueID>101568200</NlmUniqueID><ISSNLinking>1664-462X</ISSNLinking></MedlineJournalInfo><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Populus trichocarpa</Keyword><Keyword MajorTopicYN="N">PtHMGR</Keyword><Keyword MajorTopicYN="N">carotene</Keyword><Keyword MajorTopicYN="N">lycopene</Keyword><Keyword MajorTopicYN="N">terpenoid</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2019</Year><Month>04</Month><Day>28</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2019</Year><Month>10</Month><Day>24</Day></PubMedPubDate><PubMedPubDate 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Dawei" sort="Li, Dawei" uniqKey="Li D" first="Dawei" last="Li">Dawei Li</name><name sortKey="Movahedi, Ali" sort="Movahedi, Ali" uniqKey="Movahedi A" first="Ali" last="Movahedi">Ali Movahedi</name><name sortKey="Sun, Weibo" sort="Sun, Weibo" uniqKey="Sun W" first="Weibo" last="Sun">Weibo Sun</name><name sortKey="Xu, Chen" sort="Xu, Chen" uniqKey="Xu C" first="Chen" last="Xu">Chen Xu</name><name sortKey="Xu, Chen" sort="Xu, Chen" uniqKey="Xu C" first="Chen" last="Xu">Chen Xu</name><name sortKey="Zhuge, Qiang" sort="Zhuge, Qiang" uniqKey="Zhuge Q" first="Qiang" last="Zhuge">Qiang Zhuge</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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