Norcoclaurine Synthase Is a Member of the Pathogenesis-Related 10/Bet v1 Protein Family
Norcoclaurine synthase (NCS) catalyzes the first committed step in the biosynthesis of benzylisoquinoline alkaloids (BIAs). NCS from Thalictrum flavum (Tf NCS), Papaver somniferum (Ps NCS1 and Ps NCS2), and Coptis japonica (Cj PR10A) share substantial identity with pathogen-related 10 (PR10) and Bet...
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| Veröffentlicht in: | The Plant cell 2010-10, Vol.22 (10), p.3489-3503 |
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| creator | Lee, Eun-Jeong Facchini, Peter |
| description | Norcoclaurine synthase (NCS) catalyzes the first committed step in the biosynthesis of benzylisoquinoline alkaloids (BIAs). NCS from Thalictrum flavum (Tf NCS), Papaver somniferum (Ps NCS1 and Ps NCS2), and Coptis japonica (Cj PR10A) share substantial identity with pathogen-related 10 (PR10) and Bet v1 proteins, whose functions are not well understood. A distinct enzyme (Cj NCS1) with similarity to 2-oxoglutarate-dependent dioxygenases was suggested as the bona fide NCS in C. japonica. Here, we validate the exclusive role of PR10/Bet v1-type NCS enzymes in BIA metabolism. Immunolocalization of Ps NCS2 revealed its cell type-specific occurrence in phloem sieve elements, which contain all other known BIA biosynthetic enzymes. In opium poppy, NCS transcripts and proteins were abundant in root and stem, but at low levels in leaf and carpel. Silencing of NCS in opium poppy profoundly reduced alkaloid levels compared with controls. Immunoprecipitation of NCS from total protein extracts of T. flavum cells resulted in a nearly complete attenuation of NCS activity. A Ps NCS2-green fluorescent protein fusion introduced by microprojectile bombardment into opium poppy cells initially localized to the endoplasmic reticulum but subsequently sorted to the vacuole. In our hands, Cj NCS1 did not catalyze the formation of (S)-norcoclaurine from dopamine and 4-hydroxyphenylacetaldehyde. |
| doi_str_mv | 10.1105/tpc.110.077958 |
| format | Article |
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NCS from Thalictrum flavum (Tf NCS), Papaver somniferum (Ps NCS1 and Ps NCS2), and Coptis japonica (Cj PR10A) share substantial identity with pathogen-related 10 (PR10) and Bet v1 proteins, whose functions are not well understood. A distinct enzyme (Cj NCS1) with similarity to 2-oxoglutarate-dependent dioxygenases was suggested as the bona fide NCS in C. japonica. Here, we validate the exclusive role of PR10/Bet v1-type NCS enzymes in BIA metabolism. Immunolocalization of Ps NCS2 revealed its cell type-specific occurrence in phloem sieve elements, which contain all other known BIA biosynthetic enzymes. In opium poppy, NCS transcripts and proteins were abundant in root and stem, but at low levels in leaf and carpel. Silencing of NCS in opium poppy profoundly reduced alkaloid levels compared with controls. Immunoprecipitation of NCS from total protein extracts of T. flavum cells resulted in a nearly complete attenuation of NCS activity. A Ps NCS2-green fluorescent protein fusion introduced by microprojectile bombardment into opium poppy cells initially localized to the endoplasmic reticulum but subsequently sorted to the vacuole. In our hands, Cj NCS1 did not catalyze the formation of (S)-norcoclaurine from dopamine and 4-hydroxyphenylacetaldehyde.</description><identifier>ISSN: 1040-4651</identifier><identifier>EISSN: 1532-298X</identifier><identifier>DOI: 10.1105/tpc.110.077958</identifier><identifier>PMID: 21037103</identifier><language>eng</language><publisher>United States: American Society of Plant Biologists</publisher><subject>Alkaloids ; Amino Acid Sequence ; Amino acids ; Antiserum ; Benzylisoquinolines - metabolism ; Biosynthesis ; Carbon-Nitrogen Ligases - genetics ; Carbon-Nitrogen Ligases - metabolism ; Cell culture techniques ; Coptis - enzymology ; Coptis - genetics ; Enzymes ; Gene Silencing ; Molecular Sequence Data ; Opium ; Papaver - enzymology ; Papaver - genetics ; Phloem - metabolism ; Phylogeny ; Plant cells ; Proteins ; Recombinant Proteins - genetics ; Recombinant Proteins - metabolism ; Sequence Alignment ; Sieve elements ; Thalictrum - enzymology ; Thalictrum - genetics</subject><ispartof>The Plant cell, 2010-10, Vol.22 (10), p.3489-3503</ispartof><rights>2010 American Society of Plant Biologists</rights><rights>Copyright American Society of Plant Biologists Oct 2010</rights><rights>2010 American Society of Plant Biologists 2010</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c528t-5da9e38889e380b75110e9d9f6f9896f771cf6a79b555f723e626b81479c61bb3</citedby><cites>FETCH-LOGICAL-c528t-5da9e38889e380b75110e9d9f6f9896f771cf6a79b555f723e626b81479c61bb3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/25758787$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/25758787$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,315,781,785,804,886,27928,27929,58021,58254</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21037103$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Lee, Eun-Jeong</creatorcontrib><creatorcontrib>Facchini, Peter</creatorcontrib><title>Norcoclaurine Synthase Is a Member of the Pathogenesis-Related 10/Bet v1 Protein Family</title><title>The Plant cell</title><addtitle>Plant Cell</addtitle><description>Norcoclaurine synthase (NCS) catalyzes the first committed step in the biosynthesis of benzylisoquinoline alkaloids (BIAs). NCS from Thalictrum flavum (Tf NCS), Papaver somniferum (Ps NCS1 and Ps NCS2), and Coptis japonica (Cj PR10A) share substantial identity with pathogen-related 10 (PR10) and Bet v1 proteins, whose functions are not well understood. A distinct enzyme (Cj NCS1) with similarity to 2-oxoglutarate-dependent dioxygenases was suggested as the bona fide NCS in C. japonica. Here, we validate the exclusive role of PR10/Bet v1-type NCS enzymes in BIA metabolism. Immunolocalization of Ps NCS2 revealed its cell type-specific occurrence in phloem sieve elements, which contain all other known BIA biosynthetic enzymes. In opium poppy, NCS transcripts and proteins were abundant in root and stem, but at low levels in leaf and carpel. Silencing of NCS in opium poppy profoundly reduced alkaloid levels compared with controls. Immunoprecipitation of NCS from total protein extracts of T. flavum cells resulted in a nearly complete attenuation of NCS activity. A Ps NCS2-green fluorescent protein fusion introduced by microprojectile bombardment into opium poppy cells initially localized to the endoplasmic reticulum but subsequently sorted to the vacuole. In our hands, Cj NCS1 did not catalyze the formation of (S)-norcoclaurine from dopamine and 4-hydroxyphenylacetaldehyde.</description><subject>Alkaloids</subject><subject>Amino Acid Sequence</subject><subject>Amino acids</subject><subject>Antiserum</subject><subject>Benzylisoquinolines - metabolism</subject><subject>Biosynthesis</subject><subject>Carbon-Nitrogen Ligases - genetics</subject><subject>Carbon-Nitrogen Ligases - metabolism</subject><subject>Cell culture techniques</subject><subject>Coptis - enzymology</subject><subject>Coptis - genetics</subject><subject>Enzymes</subject><subject>Gene Silencing</subject><subject>Molecular Sequence Data</subject><subject>Opium</subject><subject>Papaver - enzymology</subject><subject>Papaver - genetics</subject><subject>Phloem - metabolism</subject><subject>Phylogeny</subject><subject>Plant cells</subject><subject>Proteins</subject><subject>Recombinant Proteins - genetics</subject><subject>Recombinant Proteins - metabolism</subject><subject>Sequence Alignment</subject><subject>Sieve elements</subject><subject>Thalictrum - enzymology</subject><subject>Thalictrum - genetics</subject><issn>1040-4651</issn><issn>1532-298X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2010</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>eNpdkUtv1DAUhSMEog_YsgMsNqwy9XXi2N5UgqovqUBFqWBnOZ6bmYySeLCdSvPvcZTSFha2j3Q-H_vqZNkboAsAyo_i1k5iQYVQXD7L9oEXLGdK_nqeNC1pXlYc9rKDEDaUUhCgXmZ7DGgh0trPfn513jrbmdG3A5Kb3RDXJiC5DMSQL9jX6IlrSFwjuTZx7VY4YGhD_h07E3FJgB59xkjugFx7F7EdyJnp2273KnvRmC7g6_vzMLs9O_1xcpFffTu_PPl0lVvOZMz50igspJTTTmvB0yyolqqpGiVV1QgBtqmMUDXnvBGswIpVtYRSKFtBXReH2fGcux3rHpcWh-hNp7e-7Y3faWda_a8ztGu9cneaKUWhZCng432Ad79HDFH3bbDYdWZANwYtIb1cMQmJ_PAfuXGjH9J0CRJClkJOcYsZst6F4LF5-ApQPTWmU2OT0HNj6cK7pwM84H8rSsDbGdiE6PyjzwWXQorkv5_9xjhtVr4N-vaGUSgoKChAqeIPxs6jhA</recordid><startdate>20101001</startdate><enddate>20101001</enddate><creator>Lee, Eun-Jeong</creator><creator>Facchini, Peter</creator><general>American Society of Plant Biologists</general><scope>FBQ</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>3V.</scope><scope>4T-</scope><scope>7QO</scope><scope>7TM</scope><scope>7X2</scope><scope>7X7</scope><scope>7XB</scope><scope>88A</scope><scope>88E</scope><scope>88I</scope><scope>8AF</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AFKRA</scope><scope>ATCPS</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0K</scope><scope>M0S</scope><scope>M1P</scope><scope>M2P</scope><scope>M7P</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>RC3</scope><scope>S0X</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20101001</creationdate><title>Norcoclaurine Synthase Is a Member of the Pathogenesis-Related 10/Bet v1 Protein Family</title><author>Lee, Eun-Jeong ; Facchini, Peter</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c528t-5da9e38889e380b75110e9d9f6f9896f771cf6a79b555f723e626b81479c61bb3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2010</creationdate><topic>Alkaloids</topic><topic>Amino Acid Sequence</topic><topic>Amino acids</topic><topic>Antiserum</topic><topic>Benzylisoquinolines - metabolism</topic><topic>Biosynthesis</topic><topic>Carbon-Nitrogen Ligases - genetics</topic><topic>Carbon-Nitrogen Ligases - metabolism</topic><topic>Cell culture techniques</topic><topic>Coptis - enzymology</topic><topic>Coptis - genetics</topic><topic>Enzymes</topic><topic>Gene Silencing</topic><topic>Molecular Sequence Data</topic><topic>Opium</topic><topic>Papaver - enzymology</topic><topic>Papaver - genetics</topic><topic>Phloem - metabolism</topic><topic>Phylogeny</topic><topic>Plant cells</topic><topic>Proteins</topic><topic>Recombinant Proteins - genetics</topic><topic>Recombinant Proteins - metabolism</topic><topic>Sequence Alignment</topic><topic>Sieve elements</topic><topic>Thalictrum - enzymology</topic><topic>Thalictrum - genetics</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Lee, Eun-Jeong</creatorcontrib><creatorcontrib>Facchini, Peter</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Docstoc</collection><collection>Biotechnology Research Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Agricultural Science Collection</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Biology Database (Alumni Edition)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Science Database (Alumni Edition)</collection><collection>STEM Database</collection><collection>ProQuest Pharma Collection</collection><collection>Technology Research Database</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 Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>Agricultural & Environmental Science Collection</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>Engineering Research Database</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>Agricultural Science Database</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Science Database</collection><collection>Biological Science Database</collection><collection>Biotechnology and BioEngineering Abstracts</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 Basic</collection><collection>Genetics Abstracts</collection><collection>SIRS Editorial</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>The Plant cell</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lee, Eun-Jeong</au><au>Facchini, Peter</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Norcoclaurine Synthase Is a Member of the Pathogenesis-Related 10/Bet v1 Protein Family</atitle><jtitle>The Plant cell</jtitle><addtitle>Plant Cell</addtitle><date>2010-10-01</date><risdate>2010</risdate><volume>22</volume><issue>10</issue><spage>3489</spage><epage>3503</epage><pages>3489-3503</pages><issn>1040-4651</issn><eissn>1532-298X</eissn><abstract>Norcoclaurine synthase (NCS) catalyzes the first committed step in the biosynthesis of benzylisoquinoline alkaloids (BIAs). NCS from Thalictrum flavum (Tf NCS), Papaver somniferum (Ps NCS1 and Ps NCS2), and Coptis japonica (Cj PR10A) share substantial identity with pathogen-related 10 (PR10) and Bet v1 proteins, whose functions are not well understood. A distinct enzyme (Cj NCS1) with similarity to 2-oxoglutarate-dependent dioxygenases was suggested as the bona fide NCS in C. japonica. Here, we validate the exclusive role of PR10/Bet v1-type NCS enzymes in BIA metabolism. Immunolocalization of Ps NCS2 revealed its cell type-specific occurrence in phloem sieve elements, which contain all other known BIA biosynthetic enzymes. In opium poppy, NCS transcripts and proteins were abundant in root and stem, but at low levels in leaf and carpel. Silencing of NCS in opium poppy profoundly reduced alkaloid levels compared with controls. Immunoprecipitation of NCS from total protein extracts of T. flavum cells resulted in a nearly complete attenuation of NCS activity. A Ps NCS2-green fluorescent protein fusion introduced by microprojectile bombardment into opium poppy cells initially localized to the endoplasmic reticulum but subsequently sorted to the vacuole. In our hands, Cj NCS1 did not catalyze the formation of (S)-norcoclaurine from dopamine and 4-hydroxyphenylacetaldehyde.</abstract><cop>United States</cop><pub>American Society of Plant Biologists</pub><pmid>21037103</pmid><doi>10.1105/tpc.110.077958</doi><tpages>15</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; JSTOR Archive Collection A-Z Listing; Oxford University Press Journals All Titles (1996-Current) |
| subjects | Alkaloids Amino Acid Sequence Amino acids Antiserum Benzylisoquinolines - metabolism Biosynthesis Carbon-Nitrogen Ligases - genetics Carbon-Nitrogen Ligases - metabolism Cell culture techniques Coptis - enzymology Coptis - genetics Enzymes Gene Silencing Molecular Sequence Data Opium Papaver - enzymology Papaver - genetics Phloem - metabolism Phylogeny Plant cells Proteins Recombinant Proteins - genetics Recombinant Proteins - metabolism Sequence Alignment Sieve elements Thalictrum - enzymology Thalictrum - genetics |
| title | Norcoclaurine Synthase Is a Member of the Pathogenesis-Related 10/Bet v1 Protein Family |
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