title: A Comprehensive, CRISPR-based Functional Analysis of Essential Genes in Bacteria.
abstract: Essential gene functions underpin the core reactions required for cell viability, but their contributions and relationships are poorly studied in vivo. Using CRISPR interference, we created knockdowns of every essential gene in Bacillus subtilis and probed their phenotypes. Our high-confidence essential gene network, established using chemical genomics, showed extensive interconnections among distantly related processes and identified modes of action for uncharacterized antibiotics. Importantly, mild knockdown of essential gene functions significantly reduced stationary-phase survival without affecting maximal growth rate, suggesting that essential protein levels are set to maximize outgrowth from stationary phase. Finally, high-throughput microscopy indicated that cell morphology is relatively insensitive to mild knockdown but profoundly affected by depletion of gene function, revealing intimate connections between cell growth and shape. Our results provide a framework for systematic investigation of essential gene functions in vivo broadly applicable to diverse microorganisms and amenable to comparative analysis.
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Jason M
authors.initials: JM
authors.lastname: Peters
authors.affiliation: Biophysics Program, Stanford University, Stanford, CA 94305, USA.
authors.forename: Alexandre
authors.initials: A
authors.lastname: Colavin
authors.affiliation: Department of Bioengineering, Stanford University, Stanford, CA 94305, USA.
authors.forename: Handuo
authors.initials: H
authors.lastname: Shi
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON L8S 3Z5, Canada; Michael G. DeGroote Institute of Infectious Disease Research, McMaster University, Hamilton, ON L8N 3Z5, Canada.
authors.forename: Tomasz L
authors.initials: TL
authors.lastname: Czarny
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
authors.forename: Matthew H
authors.initials: MH
authors.lastname: Larson
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Spencer
authors.initials: S
authors.lastname: Wong
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA; Biophysics Graduate Program, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: John S
authors.initials: JS
authors.lastname: Hawkins
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Candy H S
authors.initials: CHS
authors.lastname: Lu
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Byoung-Mo
authors.initials: BM
authors.lastname: Koo
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Elizabeth
authors.initials: E
authors.lastname: Marta
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA; Biophysics Graduate Program, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Anthony L
authors.initials: AL
authors.lastname: Shiver
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research, San Francisco, CA 94158, USA; UCSF Center for Systems and Synthetic Biology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Evan H
authors.initials: EH
authors.lastname: Whitehead
authors.affiliation: Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, CA 94158, USA; California Institute for Quantitative Biomedical Research, San Francisco, CA 94158, USA; Center for RNA Systems Biology, University of California, Berkeley, Berkeley, CA 94720, USA; UCSF Center for Systems and Synthetic Biology, University of California, San Francisco, San Francisco, CA 94158, USA.
authors.forename: Jonathan S
authors.initials: JS
authors.lastname: Weissman
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON L8S 3Z5, Canada; Michael G. DeGroote Institute of Infectious Disease Research, McMaster University, Hamilton, ON L8N 3Z5, Canada.
authors.forename: Eric D
authors.initials: ED
authors.lastname: Brown
authors.affiliation: Department of Bioengineering, Stanford University, Stanford, CA 94305, USA; Department of Chemical and Systems Biology, Stanford University, Stanford, CA 94305, USA; ChEM-H, Stanford University, Stanford, CA 94305, USA. Electronic address: stanley.qi@stanford.edu.
authors.forename: Lei S
authors.initials: LS
authors.lastname: Qi
authors.affiliation: Biophysics Program, Stanford University, Stanford, CA 94305, USA; Department of Bioengineering, Stanford University, Stanford, CA 94305, USA; School of Medicine, Department of Microbiology and Immunology, Stanford University, Stanford, CA 94305, USA. Electronic address: kchuang@stanford.edu.
authors.forename: Kerwyn Casey
authors.initials: KC
authors.lastname: Huang
authors.affiliation: Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94158, USA. Electronic address: cgrossucsf@gmail.com.
authors.forename: Carol A
authors.initials: CA
authors.lastname: Gross
mesh_terms: D001412:Bacillus subtilis / Q000235:genetics*; D064113:CRISPR-Cas Systems; D055785:Gene Knockdown Techniques; D015723:Gene Library; D053263:Gene Regulatory Networks; D005798:Genes, Bacterial; D020043:Genes, Essential; D058990:Molecular Targeted Therapy