Repurposing the Damage Repair Protein Methyl Guanine Methyl Transferase as a Ligand Inducible Fusion Degron
We successfully repurpose the DNA repair protein methylguanine methyltransferase (MGMT) as an inducible degron for protein fusions. MGMT is a suicide protein that removes alkyl groups from the O6 position of guanine (O6G) and is thereafter quickly degraded by the ubiquitin proteasome pathway (UPP)....
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| Veröffentlicht in: | ACS chemical biology 2022-01, Vol.17 (1), p.24-31 |
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| creator | Murawska, Gosia M Vogel, Caspar Jan, Max Lu, Xinyan Schild, Matthias Slabicki, Mikolaj Zou, Charles Zhanybekova, Saule Manojkumar, Manisha Petzold, Georg Kaiser, Peter Thomä, Nicolas Ebert, Benjamin Gillingham, Dennis |
| description | We successfully repurpose the DNA repair protein methylguanine methyltransferase (MGMT) as an inducible degron for protein fusions. MGMT is a suicide protein that removes alkyl groups from the O6 position of guanine (O6G) and is thereafter quickly degraded by the ubiquitin proteasome pathway (UPP). Starting with MGMT pseudosubstrates (benzylguanine and lomeguatrib), we first demonstrate that these lead to potent MGMT depletion while affecting little else in the proteome. We then show that fusion proteins of MGMT undergo rapid UPP-dependent degradation in response to pseudosubstrates. Mechanistic studies confirm the involvement of the UPP, while revealing that at least two E3 ligase classes can degrade MGMT depending on cell-line and expression type (native or ectopic). We also demonstrate the technique’s versatility with two clinically relevant examples: degradation of KRASG12C and a chimeric antigen receptor. |
| doi_str_mv | 10.1021/acschembio.1c00771 |
| format | Article |
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MGMT is a suicide protein that removes alkyl groups from the O6 position of guanine (O6G) and is thereafter quickly degraded by the ubiquitin proteasome pathway (UPP). Starting with MGMT pseudosubstrates (benzylguanine and lomeguatrib), we first demonstrate that these lead to potent MGMT depletion while affecting little else in the proteome. We then show that fusion proteins of MGMT undergo rapid UPP-dependent degradation in response to pseudosubstrates. Mechanistic studies confirm the involvement of the UPP, while revealing that at least two E3 ligase classes can degrade MGMT depending on cell-line and expression type (native or ectopic). We also demonstrate the technique’s versatility with two clinically relevant examples: degradation of KRASG12C and a chimeric antigen receptor.</description><identifier>ISSN: 1554-8929</identifier><identifier>EISSN: 1554-8937</identifier><identifier>DOI: 10.1021/acschembio.1c00771</identifier><identifier>PMID: 34982531</identifier><language>eng</language><publisher>United States: American Chemical Society</publisher><subject>Cell Line ; CRISPR-Cas Systems ; DNA Damage ; DNA Modification Methylases - antagonists & inhibitors ; DNA Modification Methylases - genetics ; DNA Modification Methylases - metabolism ; DNA Repair ; DNA Repair Enzymes - antagonists & inhibitors ; DNA Repair Enzymes - genetics ; DNA Repair Enzymes - metabolism ; Humans ; Ligands ; Tumor Suppressor Proteins - antagonists & inhibitors ; Tumor Suppressor Proteins - genetics ; Tumor Suppressor Proteins - metabolism</subject><ispartof>ACS chemical biology, 2022-01, Vol.17 (1), p.24-31</ispartof><rights>2022 The Authors. Published by American Chemical Society</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a386t-f41ecc100b97bab04fa341c5e9d15e4d2fbb717af13af58a9e0820f73b04edc93</citedby><cites>FETCH-LOGICAL-a386t-f41ecc100b97bab04fa341c5e9d15e4d2fbb717af13af58a9e0820f73b04edc93</cites><orcidid>0000-0002-3672-8699</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/acschembio.1c00771$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/acschembio.1c00771$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/34982531$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Murawska, Gosia M</creatorcontrib><creatorcontrib>Vogel, Caspar</creatorcontrib><creatorcontrib>Jan, Max</creatorcontrib><creatorcontrib>Lu, Xinyan</creatorcontrib><creatorcontrib>Schild, Matthias</creatorcontrib><creatorcontrib>Slabicki, Mikolaj</creatorcontrib><creatorcontrib>Zou, Charles</creatorcontrib><creatorcontrib>Zhanybekova, Saule</creatorcontrib><creatorcontrib>Manojkumar, Manisha</creatorcontrib><creatorcontrib>Petzold, Georg</creatorcontrib><creatorcontrib>Kaiser, Peter</creatorcontrib><creatorcontrib>Thomä, Nicolas</creatorcontrib><creatorcontrib>Ebert, Benjamin</creatorcontrib><creatorcontrib>Gillingham, Dennis</creatorcontrib><title>Repurposing the Damage Repair Protein Methyl Guanine Methyl Transferase as a Ligand Inducible Fusion Degron</title><title>ACS chemical biology</title><addtitle>ACS Chem. Biol</addtitle><description>We successfully repurpose the DNA repair protein methylguanine methyltransferase (MGMT) as an inducible degron for protein fusions. MGMT is a suicide protein that removes alkyl groups from the O6 position of guanine (O6G) and is thereafter quickly degraded by the ubiquitin proteasome pathway (UPP). Starting with MGMT pseudosubstrates (benzylguanine and lomeguatrib), we first demonstrate that these lead to potent MGMT depletion while affecting little else in the proteome. We then show that fusion proteins of MGMT undergo rapid UPP-dependent degradation in response to pseudosubstrates. Mechanistic studies confirm the involvement of the UPP, while revealing that at least two E3 ligase classes can degrade MGMT depending on cell-line and expression type (native or ectopic). We also demonstrate the technique’s versatility with two clinically relevant examples: degradation of KRASG12C and a chimeric antigen receptor.</description><subject>Cell Line</subject><subject>CRISPR-Cas Systems</subject><subject>DNA Damage</subject><subject>DNA Modification Methylases - antagonists & inhibitors</subject><subject>DNA Modification Methylases - genetics</subject><subject>DNA Modification Methylases - metabolism</subject><subject>DNA Repair</subject><subject>DNA Repair Enzymes - antagonists & inhibitors</subject><subject>DNA Repair Enzymes - genetics</subject><subject>DNA Repair Enzymes - metabolism</subject><subject>Humans</subject><subject>Ligands</subject><subject>Tumor Suppressor Proteins - antagonists & inhibitors</subject><subject>Tumor Suppressor Proteins - genetics</subject><subject>Tumor Suppressor Proteins - metabolism</subject><issn>1554-8929</issn><issn>1554-8937</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2022</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kD1PwzAQhi0EglL4AwzII0uLHefLIypQKhWBUJmji3NODYld7GTg35OqpWxM57Oe95XuIeSKsylnEb8FFdQa29K4KVeMZRk_IiOeJPEklyI7PrwjeUbOQ_hgLBZpLk_JmYhlHiWCj8jnG256v3HB2Jp2a6T30EKNdPgG4-mrdx0aS5-xW383dN6DNRZ_15UHGzR6CEghUKBLU4Ot6MJWvTJlg_SxD8ZZeo-1d_aCnGhoAl7u55i8Pz6sZk-T5ct8MbtbTkDkaTfRMUelOGOlzEooWaxBxFwlKCueYFxFuiwznoHmAnSSg0SWR0xnYkCxUlKMyc2ud-PdV4-hK1oTFDYNWHR9KKKUpzJJRLZFox2qvAvBoy423rTgvwvOiq3k4k9ysZc8hK73_X3ZYnWI_FodgOkOGMLFh-u9Hc79r_EHEBaL3g</recordid><startdate>20220121</startdate><enddate>20220121</enddate><creator>Murawska, Gosia M</creator><creator>Vogel, Caspar</creator><creator>Jan, Max</creator><creator>Lu, Xinyan</creator><creator>Schild, Matthias</creator><creator>Slabicki, Mikolaj</creator><creator>Zou, Charles</creator><creator>Zhanybekova, Saule</creator><creator>Manojkumar, Manisha</creator><creator>Petzold, Georg</creator><creator>Kaiser, Peter</creator><creator>Thomä, Nicolas</creator><creator>Ebert, Benjamin</creator><creator>Gillingham, Dennis</creator><general>American Chemical Society</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>7X8</scope><orcidid>https://orcid.org/0000-0002-3672-8699</orcidid></search><sort><creationdate>20220121</creationdate><title>Repurposing the Damage Repair Protein Methyl Guanine Methyl Transferase as a Ligand Inducible Fusion Degron</title><author>Murawska, Gosia M ; Vogel, Caspar ; Jan, Max ; Lu, Xinyan ; Schild, Matthias ; Slabicki, Mikolaj ; Zou, Charles ; Zhanybekova, Saule ; Manojkumar, Manisha ; Petzold, Georg ; Kaiser, Peter ; Thomä, Nicolas ; Ebert, Benjamin ; Gillingham, Dennis</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a386t-f41ecc100b97bab04fa341c5e9d15e4d2fbb717af13af58a9e0820f73b04edc93</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2022</creationdate><topic>Cell Line</topic><topic>CRISPR-Cas Systems</topic><topic>DNA Damage</topic><topic>DNA Modification Methylases - antagonists & inhibitors</topic><topic>DNA Modification Methylases - genetics</topic><topic>DNA Modification Methylases - metabolism</topic><topic>DNA Repair</topic><topic>DNA Repair Enzymes - antagonists & inhibitors</topic><topic>DNA Repair Enzymes - genetics</topic><topic>DNA Repair Enzymes - metabolism</topic><topic>Humans</topic><topic>Ligands</topic><topic>Tumor Suppressor Proteins - antagonists & inhibitors</topic><topic>Tumor Suppressor Proteins - genetics</topic><topic>Tumor Suppressor Proteins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Murawska, Gosia M</creatorcontrib><creatorcontrib>Vogel, Caspar</creatorcontrib><creatorcontrib>Jan, Max</creatorcontrib><creatorcontrib>Lu, Xinyan</creatorcontrib><creatorcontrib>Schild, Matthias</creatorcontrib><creatorcontrib>Slabicki, Mikolaj</creatorcontrib><creatorcontrib>Zou, Charles</creatorcontrib><creatorcontrib>Zhanybekova, Saule</creatorcontrib><creatorcontrib>Manojkumar, Manisha</creatorcontrib><creatorcontrib>Petzold, Georg</creatorcontrib><creatorcontrib>Kaiser, Peter</creatorcontrib><creatorcontrib>Thomä, Nicolas</creatorcontrib><creatorcontrib>Ebert, Benjamin</creatorcontrib><creatorcontrib>Gillingham, Dennis</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>ACS chemical biology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Murawska, Gosia M</au><au>Vogel, Caspar</au><au>Jan, Max</au><au>Lu, Xinyan</au><au>Schild, Matthias</au><au>Slabicki, Mikolaj</au><au>Zou, Charles</au><au>Zhanybekova, Saule</au><au>Manojkumar, Manisha</au><au>Petzold, Georg</au><au>Kaiser, Peter</au><au>Thomä, Nicolas</au><au>Ebert, Benjamin</au><au>Gillingham, Dennis</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Repurposing the Damage Repair Protein Methyl Guanine Methyl Transferase as a Ligand Inducible Fusion Degron</atitle><jtitle>ACS chemical biology</jtitle><addtitle>ACS Chem. 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| subjects | Cell Line CRISPR-Cas Systems DNA Damage DNA Modification Methylases - antagonists & inhibitors DNA Modification Methylases - genetics DNA Modification Methylases - metabolism DNA Repair DNA Repair Enzymes - antagonists & inhibitors DNA Repair Enzymes - genetics DNA Repair Enzymes - metabolism Humans Ligands Tumor Suppressor Proteins - antagonists & inhibitors Tumor Suppressor Proteins - genetics Tumor Suppressor Proteins - metabolism |
| title | Repurposing the Damage Repair Protein Methyl Guanine Methyl Transferase as a Ligand Inducible Fusion Degron |
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