A FACS-Based Genome-wide CRISPR Screen Reveals a Requirement for COPI in Chlamydia trachomatis Invasion

Joseph S Park; Jennifer D Helble; Jacob E Lazarus; Guanhua Yang; Carlos J Blondel; John G Doench; Michael N Starnbach; Matthew K Waldor

doi:10.1016/j.isci.2018.12.011

A FACS-Based Genome-wide CRISPR Screen Reveals a Requirement for COPI in Chlamydia trachomatis Invasion

iScience. 2019 Jan 25:11:71-84. doi: 10.1016/j.isci.2018.12.011. Epub 2018 Dec 14.

Authors

Joseph S Park¹, Jennifer D Helble², Jacob E Lazarus³, Guanhua Yang⁴, Carlos J Blondel⁴, John G Doench⁵, Michael N Starnbach², Matthew K Waldor⁶

Affiliations

¹ Howard Hughes Medical Institute, Boston, MA 02215, USA; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA; Division of Infectious Diseases, Brigham & Women's Hospital, Boston 02115, MA, USA; Boston University School of Medicine, Boston, MA 02120, USA.
² Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA.
³ Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA; Division of Infectious Diseases, Brigham & Women's Hospital, Boston 02115, MA, USA; Division of Infectious Diseases, Massachusetts General Hospital, Boston, MA 02114, USA.
⁴ Division of Infectious Diseases, Brigham & Women's Hospital, Boston 02115, MA, USA.
⁵ Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA.
⁶ Howard Hughes Medical Institute, Boston, MA 02215, USA; Department of Microbiology and Immunobiology, Harvard Medical School, Boston, MA 02115, USA; Division of Infectious Diseases, Brigham & Women's Hospital, Boston 02115, MA, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA. Electronic address: mwaldor@research.bwh.harvard.edu.

Abstract

The invasion of Chlamydia trachomatis, an obligate intracellular bacterium, into epithelial cells is driven by a complex interplay of host and bacterial factors. To comprehensively define the host genes required for pathogen invasion, we undertook a fluorescence-activated cell sorting (FACS)-based CRISPR screen in human cells. A genome-wide loss-of-function library was infected with fluorescent C. trachomatis and then sorted to enrich for invasion-deficient mutants. The screen identified heparan sulfate, a known pathogen receptor, as well as coatomer complex I (COPI). We found that COPI, through a previously unappreciated role, promotes heparan sulfate cell surface presentation, thereby facilitating C. trachomatis attachment. The heparan sulfate defect does not fully account for the resistance of COPI mutants. COPI also promotes the activity of the pathogen's type III secretion system. Together, our findings establish the requirement for COPI in C. trachomatis invasion and the utility of FACS-based CRISPR screening for the elucidation of host factors required for pathogen invasion.

Keywords: Cell Biology; Genetic Engineering; Genetic Screens; Host-pathogen Interactions; Medical Microbiology; Methodology in Biological Sciences; Molecular Mechanism of Behavior; Molecular Microbiology.

Abstract

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