Novel high throughput pooled shRNA screening identifies NQO1 as a potential drug target for host directed therapy for tuberculosis.
Chemical regulation of macrophage function is one key strategy for developing host-directed adjuvant therapies for tuberculosis (TB). A critical step to develop these therapies is the identification and characterization of specific macrophage molecules and pathways with a high potential to serve as drug targets. Using a barcoded lentivirus-based pooled short-hairpin RNA (shRNA) library combined with next generation sequencing, we identified 205 silenced host genes highly enriched in mycobacteria-resistant macrophages. Twenty-one of these "hits" belonged to the oxidoreductase functional category.



quinone oxidoreductase 1 (NQO1) was the top oxidoreductase "hit". NQO1 expression was increased after mycobacterial infection, and NQO1 knockdown increased macrophage differentiation, NF-κB activation, and the secretion of pro-inflammatory cytokines TNF-α and IL-1β in response to infection. This suggests that mycobacteria hijacks NQO1 to down-regulate pro-inflammatory and anti-bacterial functions. The competitive inhibitor of NQO1 dicoumarol synergized with rifampin to promote intracellular killing of mycobacteria. Thus, NQO1 is a new host target in mycobacterial infection that could potentially be exploited to increase antibiotic efficacy in vivo. Our findings also suggest that pooled shRNA libraries could be valuable tools for genome-wide screening in the search for novel druggable host targets for adjunctive TB therapies.
Department of Medicine, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Qing
Q
Li
Department of Molecular Biology and Microbiology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Ahmad F
AF
Karim
Department of Medicine, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Xuedong
X
Ding
Department of Molecular Biology and Microbiology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Biswajit
B
Das
Department of Molecular Biology and Microbiology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Curtis
C
Dobrowolski
University Hospitals Translational Laboratory, University Hospitals Case Medical Center, Cleveland, Ohio, United States of America.
Richard M
RM
Gibson
University Hospitals Translational Laboratory, University Hospitals Case Medical Center, Cleveland, Ohio, United States of America.|Center for AIDS Research (CFAR), Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.|Department of Pathology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Miguel E
ME
Quiñones-Mateu
Department of Molecular Biology and Microbiology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.|Center for AIDS Research (CFAR), Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Jonathan
J
Karn
Department of Molecular Biology and Microbiology, Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.|Center for AIDS Research (CFAR), Case Western Reserve University &University Hospitals, Cleveland, Ohio, United States of America.
Roxana E
RE
Rojas
D000995:Antitubercular Agents / Q000494:pharmacology*; D001728:Dicumarol / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D015723:Gene Library; D059014:High-Throughput Nucleotide Sequencing; D057166:High-Throughput Screening Assays; D054884:Host-Pathogen Interactions / Q000187:drug effects*; D006801:Humans; D053583:Interleukin-1beta / Q000819:agonists / Q000235:genetics / Q000276:immunology; D008264:Macrophages / Q000187:drug effects / Q000276:immunology* / Q000382:microbiology; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000472:pathogenicity / Q000502:physiology; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000276:immunology; D016328:NF-kappa B / Q000819:agonists / Q000235:genetics / Q000276:immunology; D061251:Primary Cell Culture; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012293:Rifampin / Q000494:pharmacology; D015398:Signal Transduction; D000074084:THP-1 Cells; D014409:Tumor Necrosis Factor-alpha / Q000819:agonists / Q000235:genetics / Q000276:immunology
D000995:Antitubercular Agents; D004791:Enzyme Inhibitors; C508585:IL1B protein, human; D053583:Interleukin-1beta; D016328:NF-kappa B; D034741:RNA, Small Interfering; D014409:Tumor Necrosis Factor-alpha; D001728:Dicumarol; D016660:NAD(P)H Dehydrogenase (Quinone); C447479:NQO1 protein, human; D012293:Rifampin