Loading…

The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan

Summary Mixed‐linkage glucan (MLG) is a significant cell wall carbohydrate in grasses and an important carbon source for human consumption and biofuel production. MLG biosynthesis depends on the biochemical activity of membrane spanning glucan synthases encoded by the CSLH and CSLF cellulose synthas...

Full description

Saved in:
Bibliographic Details
Published in:The Plant journal : for cell and molecular biology 2015-02, Vol.81 (4), p.537-547
Main Authors: Kim, Sang‐Jin, Zemelis, Starla, Keegstra, Kenneth, Brandizzi, Federica
Format: Article
Language:English
Subjects:
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
cited_by
cites
container_end_page 547
container_issue 4
container_start_page 537
container_title The Plant journal : for cell and molecular biology
container_volume 81
creator Kim, Sang‐Jin
Zemelis, Starla
Keegstra, Kenneth
Brandizzi, Federica
description Summary Mixed‐linkage glucan (MLG) is a significant cell wall carbohydrate in grasses and an important carbon source for human consumption and biofuel production. MLG biosynthesis depends on the biochemical activity of membrane spanning glucan synthases encoded by the CSLH and CSLF cellulose synthase‐like gene families. CSLF proteins are the best characterized to date but relatively little information is known about their topology with respect to the biosynthetic membranes. In this study, we report on the topology of CSLF6 protein derived from the model grass species Brachypodium distachyon (BdCSLF6) when it is expressed in heterologous systems. Using live cell imaging and immuno‐electron microscopy analyses of tobacco epidermal cells expressing BdCSLF6, we demonstrate that a functional yellow fluorescent protein (YFP) fusion of BdCSLF6 is localized to the Golgi apparatus and that the Golgi localization of BdCSLF6 is sufficient for MLG biosynthesis. By implementing protease protection assays of BdCSLF6 expressed in the yeast Pichia pastoris, we also demonstrate that the catalytic domain, the N‐terminus and the C‐ terminus of the protein are exposed in the cytosol. Furthermore, we found that BdCSLF6 is capable of producing MLG not only in tobacco cells but also in Pichia, which generally does not produce MLG. Together, these results support the conclusion that BdCSLF6 can produce both of the linkages present in the (1,3;1,4)‐β‐d‐glucan chain of MLG and that the product is channelled at the Golgi into the secretory pathway for deposition into the cell wall.
doi_str_mv 10.1111/tpj.12748
format article
fullrecord <record><control><sourceid>proquest_pubme</sourceid><recordid>TN_cdi_proquest_miscellaneous_1655522449</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><sourcerecordid>1655522449</sourcerecordid><originalsourceid>FETCH-LOGICAL-j3808-55ff9b1f70c73ac5c8fc50012e06eccdb190aa45de287ecf2bd4ee129c007ee03</originalsourceid><addsrcrecordid>eNpdkc1u1DAUha0KRIfCoi-ALLHpJq1_42RZjVp-NBJInUrsIse5mXpw4hA7KmHFijXPyJPgTFsW3M298vmuZZ-D0Ckl5zTVRRz255QpURyhFeW5zDjlX56hFSlzkilB2TF6GcKeEKp4Ll6gYyalVEQUK_RrewfYzNEPTofOGuy80c7-0NH6HvsWx0XXUbs5JjXYCMvp-mZzneMwDYMfY8Aamzvd9-Bsv8Odb8Dh1o-H3dr6MPdpCjYsm539Ds2fn78T-lXvAO_cZHT_Cj1vtQvw-rGfoNvrq-36fbb59O7D-nKT7XlBikzKti1r2ipiFNdGmqI1Mn2LAcnBmKamJdFayAZYocC0rG4EAGWlIUQBEH6Czh7uHUb_bYIQq84GA87pHvwUKponZxgTokzo2__QvZ_GPr1uoYQqGCc0UW8eqanuoKmG0XZ6nKsnhxNw8QDcWwfzP52SaomuStFVh-iq7eePh4H_BdIDjk8</addsrcrecordid><sourcetype>Aggregation Database</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>1654782301</pqid></control><display><type>article</type><title>The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan</title><source>Wiley-Blackwell Read &amp; Publish Collection</source><source>EZB Electronic Journals Library</source><creator>Kim, Sang‐Jin ; Zemelis, Starla ; Keegstra, Kenneth ; Brandizzi, Federica</creator><creatorcontrib>Kim, Sang‐Jin ; Zemelis, Starla ; Keegstra, Kenneth ; Brandizzi, Federica</creatorcontrib><description>Summary Mixed‐linkage glucan (MLG) is a significant cell wall carbohydrate in grasses and an important carbon source for human consumption and biofuel production. MLG biosynthesis depends on the biochemical activity of membrane spanning glucan synthases encoded by the CSLH and CSLF cellulose synthase‐like gene families. CSLF proteins are the best characterized to date but relatively little information is known about their topology with respect to the biosynthetic membranes. In this study, we report on the topology of CSLF6 protein derived from the model grass species Brachypodium distachyon (BdCSLF6) when it is expressed in heterologous systems. Using live cell imaging and immuno‐electron microscopy analyses of tobacco epidermal cells expressing BdCSLF6, we demonstrate that a functional yellow fluorescent protein (YFP) fusion of BdCSLF6 is localized to the Golgi apparatus and that the Golgi localization of BdCSLF6 is sufficient for MLG biosynthesis. By implementing protease protection assays of BdCSLF6 expressed in the yeast Pichia pastoris, we also demonstrate that the catalytic domain, the N‐terminus and the C‐ terminus of the protein are exposed in the cytosol. Furthermore, we found that BdCSLF6 is capable of producing MLG not only in tobacco cells but also in Pichia, which generally does not produce MLG. Together, these results support the conclusion that BdCSLF6 can produce both of the linkages present in the (1,3;1,4)‐β‐d‐glucan chain of MLG and that the product is channelled at the Golgi into the secretory pathway for deposition into the cell wall.</description><identifier>ISSN: 0960-7412</identifier><identifier>EISSN: 1365-313X</identifier><identifier>DOI: 10.1111/tpj.12748</identifier><identifier>PMID: 25557048</identifier><language>eng</language><publisher>England: Blackwell Publishing Ltd</publisher><subject>Biofuels ; Biosynthesis ; Botany ; Brachypodium ; Brachypodium - genetics ; Carbon sources ; Cells, Cultured ; Cellulose ; CSLF6 ; Cytoplasm ; Cytoplasm - metabolism ; enzyme topology ; Glucans - biosynthesis ; Golgi ; Golgi Apparatus - metabolism ; Grasses ; mixed‐linkage glucan ; Plant Proteins - genetics ; Plant Proteins - metabolism ; Protein Transport ; Proteins ; Topology ; Yeasts</subject><ispartof>The Plant journal : for cell and molecular biology, 2015-02, Vol.81 (4), p.537-547</ispartof><rights>2014 The Authors The Plant Journal © 2014 John Wiley &amp; Sons Ltd</rights><rights>2014 The Authors The Plant Journal © 2014 John Wiley &amp; Sons Ltd.</rights><rights>Copyright © 2015 John Wiley &amp; Sons Ltd and the Society for Experimental Biology</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/25557048$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kim, Sang‐Jin</creatorcontrib><creatorcontrib>Zemelis, Starla</creatorcontrib><creatorcontrib>Keegstra, Kenneth</creatorcontrib><creatorcontrib>Brandizzi, Federica</creatorcontrib><title>The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan</title><title>The Plant journal : for cell and molecular biology</title><addtitle>Plant J</addtitle><description>Summary Mixed‐linkage glucan (MLG) is a significant cell wall carbohydrate in grasses and an important carbon source for human consumption and biofuel production. MLG biosynthesis depends on the biochemical activity of membrane spanning glucan synthases encoded by the CSLH and CSLF cellulose synthase‐like gene families. CSLF proteins are the best characterized to date but relatively little information is known about their topology with respect to the biosynthetic membranes. In this study, we report on the topology of CSLF6 protein derived from the model grass species Brachypodium distachyon (BdCSLF6) when it is expressed in heterologous systems. Using live cell imaging and immuno‐electron microscopy analyses of tobacco epidermal cells expressing BdCSLF6, we demonstrate that a functional yellow fluorescent protein (YFP) fusion of BdCSLF6 is localized to the Golgi apparatus and that the Golgi localization of BdCSLF6 is sufficient for MLG biosynthesis. By implementing protease protection assays of BdCSLF6 expressed in the yeast Pichia pastoris, we also demonstrate that the catalytic domain, the N‐terminus and the C‐ terminus of the protein are exposed in the cytosol. Furthermore, we found that BdCSLF6 is capable of producing MLG not only in tobacco cells but also in Pichia, which generally does not produce MLG. Together, these results support the conclusion that BdCSLF6 can produce both of the linkages present in the (1,3;1,4)‐β‐d‐glucan chain of MLG and that the product is channelled at the Golgi into the secretory pathway for deposition into the cell wall.</description><subject>Biofuels</subject><subject>Biosynthesis</subject><subject>Botany</subject><subject>Brachypodium</subject><subject>Brachypodium - genetics</subject><subject>Carbon sources</subject><subject>Cells, Cultured</subject><subject>Cellulose</subject><subject>CSLF6</subject><subject>Cytoplasm</subject><subject>Cytoplasm - metabolism</subject><subject>enzyme topology</subject><subject>Glucans - biosynthesis</subject><subject>Golgi</subject><subject>Golgi Apparatus - metabolism</subject><subject>Grasses</subject><subject>mixed‐linkage glucan</subject><subject>Plant Proteins - genetics</subject><subject>Plant Proteins - metabolism</subject><subject>Protein Transport</subject><subject>Proteins</subject><subject>Topology</subject><subject>Yeasts</subject><issn>0960-7412</issn><issn>1365-313X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2015</creationdate><recordtype>article</recordtype><recordid>eNpdkc1u1DAUha0KRIfCoi-ALLHpJq1_42RZjVp-NBJInUrsIse5mXpw4hA7KmHFijXPyJPgTFsW3M298vmuZZ-D0Ckl5zTVRRz255QpURyhFeW5zDjlX56hFSlzkilB2TF6GcKeEKp4Ll6gYyalVEQUK_RrewfYzNEPTofOGuy80c7-0NH6HvsWx0XXUbs5JjXYCMvp-mZzneMwDYMfY8Aamzvd9-Bsv8Odb8Dh1o-H3dr6MPdpCjYsm539Ds2fn78T-lXvAO_cZHT_Cj1vtQvw-rGfoNvrq-36fbb59O7D-nKT7XlBikzKti1r2ipiFNdGmqI1Mn2LAcnBmKamJdFayAZYocC0rG4EAGWlIUQBEH6Czh7uHUb_bYIQq84GA87pHvwUKponZxgTokzo2__QvZ_GPr1uoYQqGCc0UW8eqanuoKmG0XZ6nKsnhxNw8QDcWwfzP52SaomuStFVh-iq7eePh4H_BdIDjk8</recordid><startdate>201502</startdate><enddate>201502</enddate><creator>Kim, Sang‐Jin</creator><creator>Zemelis, Starla</creator><creator>Keegstra, Kenneth</creator><creator>Brandizzi, Federica</creator><general>Blackwell Publishing Ltd</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>7QO</scope><scope>7QP</scope><scope>7QR</scope><scope>7TM</scope><scope>8FD</scope><scope>FR3</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>201502</creationdate><title>The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan</title><author>Kim, Sang‐Jin ; Zemelis, Starla ; Keegstra, Kenneth ; Brandizzi, Federica</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-j3808-55ff9b1f70c73ac5c8fc50012e06eccdb190aa45de287ecf2bd4ee129c007ee03</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2015</creationdate><topic>Biofuels</topic><topic>Biosynthesis</topic><topic>Botany</topic><topic>Brachypodium</topic><topic>Brachypodium - genetics</topic><topic>Carbon sources</topic><topic>Cells, Cultured</topic><topic>Cellulose</topic><topic>CSLF6</topic><topic>Cytoplasm</topic><topic>Cytoplasm - metabolism</topic><topic>enzyme topology</topic><topic>Glucans - biosynthesis</topic><topic>Golgi</topic><topic>Golgi Apparatus - metabolism</topic><topic>Grasses</topic><topic>mixed‐linkage glucan</topic><topic>Plant Proteins - genetics</topic><topic>Plant Proteins - metabolism</topic><topic>Protein Transport</topic><topic>Proteins</topic><topic>Topology</topic><topic>Yeasts</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kim, Sang‐Jin</creatorcontrib><creatorcontrib>Zemelis, Starla</creatorcontrib><creatorcontrib>Keegstra, Kenneth</creatorcontrib><creatorcontrib>Brandizzi, Federica</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>Biotechnology Research Abstracts</collection><collection>Calcium &amp; Calcified Tissue Abstracts</collection><collection>Chemoreception Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>The Plant journal : for cell and molecular biology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kim, Sang‐Jin</au><au>Zemelis, Starla</au><au>Keegstra, Kenneth</au><au>Brandizzi, Federica</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan</atitle><jtitle>The Plant journal : for cell and molecular biology</jtitle><addtitle>Plant J</addtitle><date>2015-02</date><risdate>2015</risdate><volume>81</volume><issue>4</issue><spage>537</spage><epage>547</epage><pages>537-547</pages><issn>0960-7412</issn><eissn>1365-313X</eissn><abstract>Summary Mixed‐linkage glucan (MLG) is a significant cell wall carbohydrate in grasses and an important carbon source for human consumption and biofuel production. MLG biosynthesis depends on the biochemical activity of membrane spanning glucan synthases encoded by the CSLH and CSLF cellulose synthase‐like gene families. CSLF proteins are the best characterized to date but relatively little information is known about their topology with respect to the biosynthetic membranes. In this study, we report on the topology of CSLF6 protein derived from the model grass species Brachypodium distachyon (BdCSLF6) when it is expressed in heterologous systems. Using live cell imaging and immuno‐electron microscopy analyses of tobacco epidermal cells expressing BdCSLF6, we demonstrate that a functional yellow fluorescent protein (YFP) fusion of BdCSLF6 is localized to the Golgi apparatus and that the Golgi localization of BdCSLF6 is sufficient for MLG biosynthesis. By implementing protease protection assays of BdCSLF6 expressed in the yeast Pichia pastoris, we also demonstrate that the catalytic domain, the N‐terminus and the C‐ terminus of the protein are exposed in the cytosol. Furthermore, we found that BdCSLF6 is capable of producing MLG not only in tobacco cells but also in Pichia, which generally does not produce MLG. Together, these results support the conclusion that BdCSLF6 can produce both of the linkages present in the (1,3;1,4)‐β‐d‐glucan chain of MLG and that the product is channelled at the Golgi into the secretory pathway for deposition into the cell wall.</abstract><cop>England</cop><pub>Blackwell Publishing Ltd</pub><pmid>25557048</pmid><doi>10.1111/tpj.12748</doi><tpages>11</tpages><oa>free_for_read</oa></addata></record>
fulltext fulltext
identifier ISSN: 0960-7412
ispartof The Plant journal : for cell and molecular biology, 2015-02, Vol.81 (4), p.537-547
issn 0960-7412
1365-313X
language eng
recordid cdi_proquest_miscellaneous_1655522449
source Wiley-Blackwell Read & Publish Collection; EZB Electronic Journals Library
subjects Biofuels
Biosynthesis
Botany
Brachypodium
Brachypodium - genetics
Carbon sources
Cells, Cultured
Cellulose
CSLF6
Cytoplasm
Cytoplasm - metabolism
enzyme topology
Glucans - biosynthesis
Golgi
Golgi Apparatus - metabolism
Grasses
mixed‐linkage glucan
Plant Proteins - genetics
Plant Proteins - metabolism
Protein Transport
Proteins
Topology
Yeasts
title The cytoplasmic localization of the catalytic site of CSLF6 supports a channeling model for the biosynthesis of mixed‐linkage glucan
url http://sfxeu10.hosted.exlibrisgroup.com/loughborough?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2024-12-24T12%3A26%3A15IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-proquest_pubme&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=The%20cytoplasmic%20localization%20of%20the%20catalytic%20site%20of%20CSLF6%20supports%20a%20channeling%20model%20for%20the%20biosynthesis%20of%20mixed%E2%80%90linkage%20glucan&rft.jtitle=The%20Plant%20journal%20:%20for%20cell%20and%20molecular%20biology&rft.au=Kim,%20Sang%E2%80%90Jin&rft.date=2015-02&rft.volume=81&rft.issue=4&rft.spage=537&rft.epage=547&rft.pages=537-547&rft.issn=0960-7412&rft.eissn=1365-313X&rft_id=info:doi/10.1111/tpj.12748&rft_dat=%3Cproquest_pubme%3E1655522449%3C/proquest_pubme%3E%3Cgrp_id%3Ecdi_FETCH-LOGICAL-j3808-55ff9b1f70c73ac5c8fc50012e06eccdb190aa45de287ecf2bd4ee129c007ee03%3C/grp_id%3E%3Coa%3E%3C/oa%3E%3Curl%3E%3C/url%3E&rft_id=info:oai/&rft_pqid=1654782301&rft_id=info:pmid/25557048&rfr_iscdi=true