Genome-wide CRISPRi screen identifies enhanced autolithotrophic phenotypes in acetogenic bacterium Eubacterium limosum
Acetogenic bacteria are a unique biocatalyst that highly promises to develop the sustainable bioconversion of carbon oxides (e.g., CO and CO ) into multicarbon biochemicals. Genotype-phenotype relationships are important for engineering their metabolic capability to enhance their biocatalytic perfor...
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| Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 2023-02, Vol.120 (6), p.e2216244120-e2216244120 |
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| creator | Shin, Jongoh Bae, Jiyun Lee, Hyeonsik Kang, Seulgi Jin, Sangrak Song, Yoseb Cho, Suhyung Cho, Byung-Kwan |
| description | Acetogenic bacteria are a unique biocatalyst that highly promises to develop the sustainable bioconversion of carbon oxides (e.g., CO and CO
) into multicarbon biochemicals. Genotype-phenotype relationships are important for engineering their metabolic capability to enhance their biocatalytic performance; however, systemic investigation on the fitness contribution of individual gene has been limited. Here, we report genome-scale CRISPR interference screening using 41,939 guide RNAs designed from the
genome, one of the model acetogenic species, where all genes were targeted for transcriptional suppression. We investigated the fitness contributions of 96% of the total genes identified, revealing the gene fitness and essentiality for heterotrophic and autotrophic metabolisms. Our data show that the Wood-Ljungdahl pathway, membrane regeneration, membrane protein biosynthesis, and butyrate synthesis are essential for autotrophic acetogenesis in
. Furthermore, we discovered genes that are repression targets that unbiasedly increased autotrophic growth rates fourfold and acetoin production 1.5-fold compared to the wild-type strain under CO
-H
conditions. These results provide insight for understanding acetogenic metabolism and genome engineering in acetogenic bacteria. |
| doi_str_mv | 10.1073/pnas.2216244120 |
| format | Article |
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) into multicarbon biochemicals. Genotype-phenotype relationships are important for engineering their metabolic capability to enhance their biocatalytic performance; however, systemic investigation on the fitness contribution of individual gene has been limited. Here, we report genome-scale CRISPR interference screening using 41,939 guide RNAs designed from the
genome, one of the model acetogenic species, where all genes were targeted for transcriptional suppression. We investigated the fitness contributions of 96% of the total genes identified, revealing the gene fitness and essentiality for heterotrophic and autotrophic metabolisms. Our data show that the Wood-Ljungdahl pathway, membrane regeneration, membrane protein biosynthesis, and butyrate synthesis are essential for autotrophic acetogenesis in
. Furthermore, we discovered genes that are repression targets that unbiasedly increased autotrophic growth rates fourfold and acetoin production 1.5-fold compared to the wild-type strain under CO
-H
conditions. These results provide insight for understanding acetogenic metabolism and genome engineering in acetogenic bacteria.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.2216244120</identifier><identifier>PMID: 36716373</identifier><language>eng</language><publisher>United States: National Academy of Sciences</publisher><subject>Acetogenesis ; Acetoin ; Autotrophic Processes ; Bacteria ; Bioconversion ; Biological Sciences ; Biosynthesis ; Carbon dioxide ; Carbon Dioxide - metabolism ; CRISPR ; Eubacterium - genetics ; Eubacterium - metabolism ; Eurytium limosum ; Fitness ; Gene silencing ; Genes ; Genome, Bacterial ; Genomes ; Genotypes ; Growth rate ; Membrane proteins ; Membranes ; Metabolic engineering ; Phenotypes ; Protein biosynthesis</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 2023-02, Vol.120 (6), p.e2216244120-e2216244120</ispartof><rights>Copyright National Academy of Sciences Feb 7, 2023</rights><rights>Copyright © 2023 the Author(s). Published by PNAS. 2023</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c421t-3e4f84d59e1ee606746f90cd8cfe707014e337f8d7e580df3d15cfd0809e86693</citedby><cites>FETCH-LOGICAL-c421t-3e4f84d59e1ee606746f90cd8cfe707014e337f8d7e580df3d15cfd0809e86693</cites><orcidid>0000-0003-4788-4184 ; 0000-0003-4363-5148</orcidid></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC9963998/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC9963998/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,723,776,780,881,27903,27904,53770,53772</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/36716373$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Shin, Jongoh</creatorcontrib><creatorcontrib>Bae, Jiyun</creatorcontrib><creatorcontrib>Lee, Hyeonsik</creatorcontrib><creatorcontrib>Kang, Seulgi</creatorcontrib><creatorcontrib>Jin, Sangrak</creatorcontrib><creatorcontrib>Song, Yoseb</creatorcontrib><creatorcontrib>Cho, Suhyung</creatorcontrib><creatorcontrib>Cho, Byung-Kwan</creatorcontrib><title>Genome-wide CRISPRi screen identifies enhanced autolithotrophic phenotypes in acetogenic bacterium Eubacterium limosum</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>Acetogenic bacteria are a unique biocatalyst that highly promises to develop the sustainable bioconversion of carbon oxides (e.g., CO and CO
) into multicarbon biochemicals. Genotype-phenotype relationships are important for engineering their metabolic capability to enhance their biocatalytic performance; however, systemic investigation on the fitness contribution of individual gene has been limited. Here, we report genome-scale CRISPR interference screening using 41,939 guide RNAs designed from the
genome, one of the model acetogenic species, where all genes were targeted for transcriptional suppression. We investigated the fitness contributions of 96% of the total genes identified, revealing the gene fitness and essentiality for heterotrophic and autotrophic metabolisms. Our data show that the Wood-Ljungdahl pathway, membrane regeneration, membrane protein biosynthesis, and butyrate synthesis are essential for autotrophic acetogenesis in
. Furthermore, we discovered genes that are repression targets that unbiasedly increased autotrophic growth rates fourfold and acetoin production 1.5-fold compared to the wild-type strain under CO
-H
conditions. These results provide insight for understanding acetogenic metabolism and genome engineering in acetogenic bacteria.</description><subject>Acetogenesis</subject><subject>Acetoin</subject><subject>Autotrophic Processes</subject><subject>Bacteria</subject><subject>Bioconversion</subject><subject>Biological Sciences</subject><subject>Biosynthesis</subject><subject>Carbon dioxide</subject><subject>Carbon Dioxide - metabolism</subject><subject>CRISPR</subject><subject>Eubacterium - genetics</subject><subject>Eubacterium - metabolism</subject><subject>Eurytium limosum</subject><subject>Fitness</subject><subject>Gene silencing</subject><subject>Genes</subject><subject>Genome, Bacterial</subject><subject>Genomes</subject><subject>Genotypes</subject><subject>Growth rate</subject><subject>Membrane proteins</subject><subject>Membranes</subject><subject>Metabolic engineering</subject><subject>Phenotypes</subject><subject>Protein biosynthesis</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2023</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpdkc9rFTEQx4Mo9lk9e5MFL162za-XbC6CPGotFJSq55Amk27KbrIm2Zb-9-bR2qqnGWY-82W-fBF6S_ARwZIdL9GUI0qJoJwTip-hDcGK9IIr_BxtMKayHzjlB-hVKdcYY7Ud8Et0wIQkgkm2QTenENMM_W1w0O0uzr5_uwhdsRkgdm0Ua_ABSgdxNNGC68xa0xTqmGpOyxhst4xNoN4tDQqxMxZquoLYFpfGVshhnbuT9amfwpzKOr9GL7yZCrx5qIfo5-eTH7sv_fnX07Pdp_Peckpqz4D7gbutAgIgsJBceIWtG6wHiSUmHBiTfnASmjHnmSNb6x0esIJBCMUO0cd73WW9nMHZZiibSS85zCbf6WSC_ncTw6iv0o1WSjClhibw4UEgp18rlKrnUCxMk4mQ1qKplIQxJgRt6Pv_0Ou05tjs7SmuON0q1qjje8rmVEoG__gMwXqfqd5nqp8ybRfv_vbwyP8Jkf0Gk0ugfg</recordid><startdate>20230207</startdate><enddate>20230207</enddate><creator>Shin, Jongoh</creator><creator>Bae, Jiyun</creator><creator>Lee, Hyeonsik</creator><creator>Kang, Seulgi</creator><creator>Jin, Sangrak</creator><creator>Song, Yoseb</creator><creator>Cho, Suhyung</creator><creator>Cho, Byung-Kwan</creator><general>National Academy of Sciences</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>7QG</scope><scope>7QL</scope><scope>7QP</scope><scope>7QR</scope><scope>7SN</scope><scope>7SS</scope><scope>7T5</scope><scope>7TK</scope><scope>7TM</scope><scope>7TO</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope><scope>5PM</scope><orcidid>https://orcid.org/0000-0003-4788-4184</orcidid><orcidid>https://orcid.org/0000-0003-4363-5148</orcidid></search><sort><creationdate>20230207</creationdate><title>Genome-wide CRISPRi screen identifies enhanced autolithotrophic phenotypes in acetogenic bacterium Eubacterium limosum</title><author>Shin, Jongoh ; Bae, Jiyun ; Lee, Hyeonsik ; Kang, Seulgi ; Jin, Sangrak ; Song, Yoseb ; Cho, Suhyung ; Cho, Byung-Kwan</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c421t-3e4f84d59e1ee606746f90cd8cfe707014e337f8d7e580df3d15cfd0809e86693</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2023</creationdate><topic>Acetogenesis</topic><topic>Acetoin</topic><topic>Autotrophic Processes</topic><topic>Bacteria</topic><topic>Bioconversion</topic><topic>Biological Sciences</topic><topic>Biosynthesis</topic><topic>Carbon dioxide</topic><topic>Carbon Dioxide - metabolism</topic><topic>CRISPR</topic><topic>Eubacterium - genetics</topic><topic>Eubacterium - metabolism</topic><topic>Eurytium limosum</topic><topic>Fitness</topic><topic>Gene silencing</topic><topic>Genes</topic><topic>Genome, Bacterial</topic><topic>Genomes</topic><topic>Genotypes</topic><topic>Growth rate</topic><topic>Membrane proteins</topic><topic>Membranes</topic><topic>Metabolic engineering</topic><topic>Phenotypes</topic><topic>Protein biosynthesis</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Shin, Jongoh</creatorcontrib><creatorcontrib>Bae, Jiyun</creatorcontrib><creatorcontrib>Lee, Hyeonsik</creatorcontrib><creatorcontrib>Kang, Seulgi</creatorcontrib><creatorcontrib>Jin, Sangrak</creatorcontrib><creatorcontrib>Song, Yoseb</creatorcontrib><creatorcontrib>Cho, Suhyung</creatorcontrib><creatorcontrib>Cho, Byung-Kwan</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Animal Behavior Abstracts</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Calcium & Calcified Tissue Abstracts</collection><collection>Chemoreception Abstracts</collection><collection>Ecology Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Immunology Abstracts</collection><collection>Neurosciences Abstracts</collection><collection>Nucleic Acids Abstracts</collection><collection>Oncogenes and Growth Factors Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Shin, Jongoh</au><au>Bae, Jiyun</au><au>Lee, Hyeonsik</au><au>Kang, Seulgi</au><au>Jin, Sangrak</au><au>Song, Yoseb</au><au>Cho, Suhyung</au><au>Cho, Byung-Kwan</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Genome-wide CRISPRi screen identifies enhanced autolithotrophic phenotypes in acetogenic bacterium Eubacterium limosum</atitle><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle><addtitle>Proc Natl Acad Sci U S A</addtitle><date>2023-02-07</date><risdate>2023</risdate><volume>120</volume><issue>6</issue><spage>e2216244120</spage><epage>e2216244120</epage><pages>e2216244120-e2216244120</pages><issn>0027-8424</issn><eissn>1091-6490</eissn><abstract>Acetogenic bacteria are a unique biocatalyst that highly promises to develop the sustainable bioconversion of carbon oxides (e.g., CO and CO
) into multicarbon biochemicals. Genotype-phenotype relationships are important for engineering their metabolic capability to enhance their biocatalytic performance; however, systemic investigation on the fitness contribution of individual gene has been limited. Here, we report genome-scale CRISPR interference screening using 41,939 guide RNAs designed from the
genome, one of the model acetogenic species, where all genes were targeted for transcriptional suppression. We investigated the fitness contributions of 96% of the total genes identified, revealing the gene fitness and essentiality for heterotrophic and autotrophic metabolisms. Our data show that the Wood-Ljungdahl pathway, membrane regeneration, membrane protein biosynthesis, and butyrate synthesis are essential for autotrophic acetogenesis in
. Furthermore, we discovered genes that are repression targets that unbiasedly increased autotrophic growth rates fourfold and acetoin production 1.5-fold compared to the wild-type strain under CO
-H
conditions. These results provide insight for understanding acetogenic metabolism and genome engineering in acetogenic bacteria.</abstract><cop>United States</cop><pub>National Academy of Sciences</pub><pmid>36716373</pmid><doi>10.1073/pnas.2216244120</doi><orcidid>https://orcid.org/0000-0003-4788-4184</orcidid><orcidid>https://orcid.org/0000-0003-4363-5148</orcidid><oa>free_for_read</oa></addata></record> |
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| ispartof | Proceedings of the National Academy of Sciences - PNAS, 2023-02, Vol.120 (6), p.e2216244120-e2216244120 |
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| source | MEDLINE; PubMed Central; Alma/SFX Local Collection; Free Full-Text Journals in Chemistry |
| subjects | Acetogenesis Acetoin Autotrophic Processes Bacteria Bioconversion Biological Sciences Biosynthesis Carbon dioxide Carbon Dioxide - metabolism CRISPR Eubacterium - genetics Eubacterium - metabolism Eurytium limosum Fitness Gene silencing Genes Genome, Bacterial Genomes Genotypes Growth rate Membrane proteins Membranes Metabolic engineering Phenotypes Protein biosynthesis |
| title | Genome-wide CRISPRi screen identifies enhanced autolithotrophic phenotypes in acetogenic bacterium Eubacterium limosum |
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