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Site-specific monoubiquitination downregulates Rab5 by disrupting effector binding and guanine nucleotide conversion
Rab GTPases, which are involved in intracellular trafficking pathways, have recently been reported to be ubiquitinated. However, the functions of ubiquitinated Rab proteins remain unexplored. Here we show that Rab5 is monoubiquitinated on K116, K140, and K165. Upon co-transfection with ubiquitin, Ra...
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| Published in: | eLife 2017-10, Vol.6 |
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| creator | Shin, Donghyuk Na, Wooju Lee, Ji-Hyung Kim, Gyuhee Baek, Jiseok Park, Seok Hee Choi, Cheol Yong Lee, Sangho |
| description | Rab GTPases, which are involved in intracellular trafficking pathways, have recently been reported to be ubiquitinated. However, the functions of ubiquitinated Rab proteins remain unexplored. Here we show that Rab5 is monoubiquitinated on K116, K140, and K165. Upon co-transfection with ubiquitin, Rab5 exhibited abnormalities in endosomal localization and EGF-induced EGF receptor degradation. Rab5 K140R and K165R mutants restored these abnormalities, whereas K116R did not. We derived structural models of individual monoubiquitinated Rab5 proteins (mUbRab5s) by solution scattering and observed different conformational flexibilities in a site-specific manner. Structural analysis combined with biochemical data revealed that interactions with downstream effectors were impeded in mUbRab5
, whereas GDP release and GTP loading activities were altered in mUbRab5
. By contrast, mUbRab5
apparently had no effect. We propose a regulatory mechanism of Rab5 where monoubiquitination downregulates effector recruitment and GDP/GTP conversion in a site-specific manner. |
| doi_str_mv | 10.7554/eLife.29154 |
| format | article |
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, whereas GDP release and GTP loading activities were altered in mUbRab5
. By contrast, mUbRab5
apparently had no effect. We propose a regulatory mechanism of Rab5 where monoubiquitination downregulates effector recruitment and GDP/GTP conversion in a site-specific manner.</description><identifier>ISSN: 2050-084X</identifier><identifier>EISSN: 2050-084X</identifier><identifier>DOI: 10.7554/eLife.29154</identifier><identifier>PMID: 28968219</identifier><language>eng</language><publisher>England: eLife Science Publications, Ltd</publisher><subject>Analysis ; Biochemistry ; Biochemistry and Chemical Biology ; Cell Line ; DNA Mutational Analysis ; Down-Regulation ; Epidermal growth factors ; G proteins ; Gross domestic product ; Guanine ; Guanine Nucleotides - metabolism ; Guanosine triphosphate ; Humans ; Hydrolysis ; Immunoglobulins ; Infection ; Localization ; Protein Binding ; Protein Conformation ; Proteins ; Purines ; Rab5 ; rab5 GTP-Binding Proteins - chemistry ; rab5 GTP-Binding Proteins - genetics ; rab5 GTP-Binding Proteins - metabolism ; SAXS ; Scattering, Small Angle ; Structural Biology and Molecular Biophysics ; Transfection ; Ubiquitin ; Ubiquitination</subject><ispartof>eLife, 2017-10, Vol.6</ispartof><rights>COPYRIGHT 2017 eLife Science Publications, Ltd.</rights><rights>2017, Shin et al. This work is licensed under the Creative Commons Attribution License ( https://creativecommons.org/licenses/by/3.0/ ) (the “License”). Notwithstanding the ProQuest Terms and Conditions, you may use this content in accordance with the terms of the License.</rights><rights>2017, Shin et al 2017 Shin et al</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c533t-7cc7bca08b4eb4df97431dbc114d186c8b25946c0c40b5dfb573bd4cd651fe2f3</citedby><cites>FETCH-LOGICAL-c533t-7cc7bca08b4eb4df97431dbc114d186c8b25946c0c40b5dfb573bd4cd651fe2f3</cites><orcidid>0000-0002-8272-6133 ; 0000-0003-3886-4579</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.proquest.com/docview/1958831591/fulltextPDF?pq-origsite=primo$$EPDF$$P50$$Gproquest$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.proquest.com/docview/1958831591?pq-origsite=primo$$EHTML$$P50$$Gproquest$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,25732,27903,27904,36991,36992,44569,53770,53772,74873</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/28968219$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Shin, Donghyuk</creatorcontrib><creatorcontrib>Na, Wooju</creatorcontrib><creatorcontrib>Lee, Ji-Hyung</creatorcontrib><creatorcontrib>Kim, Gyuhee</creatorcontrib><creatorcontrib>Baek, Jiseok</creatorcontrib><creatorcontrib>Park, Seok Hee</creatorcontrib><creatorcontrib>Choi, Cheol Yong</creatorcontrib><creatorcontrib>Lee, Sangho</creatorcontrib><title>Site-specific monoubiquitination downregulates Rab5 by disrupting effector binding and guanine nucleotide conversion</title><title>eLife</title><addtitle>Elife</addtitle><description>Rab GTPases, which are involved in intracellular trafficking pathways, have recently been reported to be ubiquitinated. However, the functions of ubiquitinated Rab proteins remain unexplored. Here we show that Rab5 is monoubiquitinated on K116, K140, and K165. Upon co-transfection with ubiquitin, Rab5 exhibited abnormalities in endosomal localization and EGF-induced EGF receptor degradation. Rab5 K140R and K165R mutants restored these abnormalities, whereas K116R did not. We derived structural models of individual monoubiquitinated Rab5 proteins (mUbRab5s) by solution scattering and observed different conformational flexibilities in a site-specific manner. Structural analysis combined with biochemical data revealed that interactions with downstream effectors were impeded in mUbRab5
, whereas GDP release and GTP loading activities were altered in mUbRab5
. By contrast, mUbRab5
apparently had no effect. We propose a regulatory mechanism of Rab5 where monoubiquitination downregulates effector recruitment and GDP/GTP conversion in a site-specific manner.</description><subject>Analysis</subject><subject>Biochemistry</subject><subject>Biochemistry and Chemical Biology</subject><subject>Cell Line</subject><subject>DNA Mutational Analysis</subject><subject>Down-Regulation</subject><subject>Epidermal growth factors</subject><subject>G proteins</subject><subject>Gross domestic product</subject><subject>Guanine</subject><subject>Guanine Nucleotides - metabolism</subject><subject>Guanosine triphosphate</subject><subject>Humans</subject><subject>Hydrolysis</subject><subject>Immunoglobulins</subject><subject>Infection</subject><subject>Localization</subject><subject>Protein Binding</subject><subject>Protein Conformation</subject><subject>Proteins</subject><subject>Purines</subject><subject>Rab5</subject><subject>rab5 GTP-Binding Proteins - chemistry</subject><subject>rab5 GTP-Binding Proteins - genetics</subject><subject>rab5 GTP-Binding Proteins - metabolism</subject><subject>SAXS</subject><subject>Scattering, Small Angle</subject><subject>Structural Biology and Molecular Biophysics</subject><subject>Transfection</subject><subject>Ubiquitin</subject><subject>Ubiquitination</subject><issn>2050-084X</issn><issn>2050-084X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><sourceid>PIMPY</sourceid><sourceid>DOA</sourceid><recordid>eNpdkttrFDEUhwdRbKl98l0GfFFka64zmRehFC8LC0Kr4FvI5WRMmU22Saba_97sbq1d85Jw8vElJ_k1zUuMznrO2XtYeQdnZMCcPWmOCeJogQT78fTR-qg5zfka1dEzIfDwvDkiYugEwcNxU658gUXegPHOm3YdQ5y1v5l98UEVH0Nr46-QYJwnVSC3l0rzVt-11uc0byo0tuAcmBJTq32w24IKth1nFXyANsxmgli8hdbEcAspV-eL5plTU4bT-_mk-f7p47eLL4vV18_Li_PVwnBKy6I3ptdGIaEZaGbd0DOKrTYYM4tFZ4QmfGCdQYYhza3TvKfaMmM7jh0QR0-a5d5ro7qWm-TXKt3JqLzcFWIapUrF1xtKjvRAO0CGaMoUp7onPWe92Oq41VBdH_auzazXYA2EktR0ID3cCf6nHOOt5B2pHlwFb-4FKd7MkItc-2xgmlSAOGeJayuM8qGjFX39H3od5xTqU1WKC0ExH_A_alS1AR9crOearVSed4R2lPRkS709oOovFPhdRjXnLJdXl4fsuz1rUsw5gXtoDyO5jZvcxU3u4lbpV49f5IH9Gy76B_2j0n4</recordid><startdate>20171002</startdate><enddate>20171002</enddate><creator>Shin, Donghyuk</creator><creator>Na, Wooju</creator><creator>Lee, Ji-Hyung</creator><creator>Kim, Gyuhee</creator><creator>Baek, Jiseok</creator><creator>Park, Seok Hee</creator><creator>Choi, Cheol Yong</creator><creator>Lee, Sangho</creator><general>eLife Science Publications, Ltd</general><general>eLife Sciences Publications Ltd</general><general>eLife Sciences Publications, Ltd</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>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>88I</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>M2P</scope><scope>M7P</scope><scope>PIMPY</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>PRINS</scope><scope>Q9U</scope><scope>7X8</scope><scope>5PM</scope><scope>DOA</scope><orcidid>https://orcid.org/0000-0002-8272-6133</orcidid><orcidid>https://orcid.org/0000-0003-3886-4579</orcidid></search><sort><creationdate>20171002</creationdate><title>Site-specific monoubiquitination downregulates Rab5 by disrupting effector binding and guanine nucleotide conversion</title><author>Shin, Donghyuk ; Na, Wooju ; Lee, Ji-Hyung ; Kim, Gyuhee ; Baek, Jiseok ; Park, Seok Hee ; Choi, Cheol Yong ; Lee, Sangho</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c533t-7cc7bca08b4eb4df97431dbc114d186c8b25946c0c40b5dfb573bd4cd651fe2f3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Analysis</topic><topic>Biochemistry</topic><topic>Biochemistry and Chemical Biology</topic><topic>Cell Line</topic><topic>DNA Mutational Analysis</topic><topic>Down-Regulation</topic><topic>Epidermal growth factors</topic><topic>G proteins</topic><topic>Gross domestic product</topic><topic>Guanine</topic><topic>Guanine Nucleotides - 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Academic</collection><collection>PubMed Central (Full Participant titles)</collection><collection>Directory of Open Access Journals</collection><jtitle>eLife</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Shin, Donghyuk</au><au>Na, Wooju</au><au>Lee, Ji-Hyung</au><au>Kim, Gyuhee</au><au>Baek, Jiseok</au><au>Park, Seok Hee</au><au>Choi, Cheol Yong</au><au>Lee, Sangho</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Site-specific monoubiquitination downregulates Rab5 by disrupting effector binding and guanine nucleotide conversion</atitle><jtitle>eLife</jtitle><addtitle>Elife</addtitle><date>2017-10-02</date><risdate>2017</risdate><volume>6</volume><issn>2050-084X</issn><eissn>2050-084X</eissn><abstract>Rab GTPases, which are involved in intracellular trafficking pathways, have recently been reported to be ubiquitinated. However, the functions of ubiquitinated Rab proteins remain unexplored. Here we show that Rab5 is monoubiquitinated on K116, K140, and K165. Upon co-transfection with ubiquitin, Rab5 exhibited abnormalities in endosomal localization and EGF-induced EGF receptor degradation. Rab5 K140R and K165R mutants restored these abnormalities, whereas K116R did not. We derived structural models of individual monoubiquitinated Rab5 proteins (mUbRab5s) by solution scattering and observed different conformational flexibilities in a site-specific manner. Structural analysis combined with biochemical data revealed that interactions with downstream effectors were impeded in mUbRab5
, whereas GDP release and GTP loading activities were altered in mUbRab5
. By contrast, mUbRab5
apparently had no effect. We propose a regulatory mechanism of Rab5 where monoubiquitination downregulates effector recruitment and GDP/GTP conversion in a site-specific manner.</abstract><cop>England</cop><pub>eLife Science Publications, Ltd</pub><pmid>28968219</pmid><doi>10.7554/eLife.29154</doi><orcidid>https://orcid.org/0000-0002-8272-6133</orcidid><orcidid>https://orcid.org/0000-0003-3886-4579</orcidid><oa>free_for_read</oa></addata></record> |
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| subjects | Analysis Biochemistry Biochemistry and Chemical Biology Cell Line DNA Mutational Analysis Down-Regulation Epidermal growth factors G proteins Gross domestic product Guanine Guanine Nucleotides - metabolism Guanosine triphosphate Humans Hydrolysis Immunoglobulins Infection Localization Protein Binding Protein Conformation Proteins Purines Rab5 rab5 GTP-Binding Proteins - chemistry rab5 GTP-Binding Proteins - genetics rab5 GTP-Binding Proteins - metabolism SAXS Scattering, Small Angle Structural Biology and Molecular Biophysics Transfection Ubiquitin Ubiquitination |
| title | Site-specific monoubiquitination downregulates Rab5 by disrupting effector binding and guanine nucleotide conversion |
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