Haploid genetic screens in human cells identify host factors used by pathogens.
Loss-of-function genetic screens in model organisms have elucidated numerous biological processes, but the diploid genome of mammalian cells has precluded large-scale gene disruption. We used insertional mutagenesis to develop a screening method to generate null alleles in a human cell line haploid for all chromosomes except chromosome 8. Using this approach, we identified host factors essential for infection with influenza and genes encoding important elements of the biosynthetic pathway of diphthamide, which are required for the cytotoxic effects of diphtheria toxin and exotoxin A. We also identified genes needed for the action of cytolethal distending toxin, including a cell-surface protein that interacts with the toxin. This approach has both conceptual and practical parallels with genetic approaches in haploid yeast.
Carette
Jan E
JE
Whitehead Institute for Biomedical Research, 9 Cambridge Center, Cambridge, MA 02142, USA.
Guimaraes
Carla P
CP
Varadarajan
Malini
M
Park
Annie S
AS
Wuethrich
Irene
I
Godarova
Alzbeta
A
Kotecki
Maciej
M
Cochran
Brent H
BH
Spooner
Eric
E
Ploegh
Hidde L
HL
Brummelkamp
Thijn R
TR
D036002:ADP Ribose Transferases / Q000378:metabolism / Q000633:toxicity; D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000595:Amino Acid Sequence; D000942:Antigens, Bacterial / Q000378:metabolism / Q000633:toxicity; D001427:Bacterial Toxins / Q000378:metabolism* / Q000633:toxicity; D053898:Biosynthetic Pathways; D002265:Carboxylic Ester Hydrolases; D045744:Cell Line, Tumor; D004167:Diphtheria Toxin / Q000378:metabolism / Q000633:toxicity; D005098:Exotoxins / Q000378:metabolism / Q000633:toxicity; D005796:Genes; D005820:Genetic Testing; D006238:Haploidy; D006639:Histidine / Q000031:analogs & derivatives / Q000096:biosynthesis; D054884:Host-Pathogen Interactions; D006801:Humans; D053118:Influenza A Virus, H1N1 Subtype / Q000472:pathogenicity*; D008780:Methyltransferases; D008969:Molecular Sequence Data; D009004:Monosaccharide Transport Proteins / Q000235:genetics / Q000378:metabolism; D016254:Mutagenesis, Insertional; D000216:N-Acylneuraminate Cytidylyltransferase / Q000235:genetics / Q000378:metabolism; D020652:Peptide Elongation Factor 2 / Q000378:metabolism; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D037521:Virulence Factors / Q000378:metabolism / Q000633:toxicity; D000097668:Pseudomonas aeruginosa Exotoxin A; D000099237:UDP-Galactose Translocators
D036002:ADP Ribose Transferases; D000246:Adenosine Diphosphate Ribose; D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D002265:Carboxylic Ester Hydrolases; D004167:Diphtheria Toxin; D005098:Exotoxins; D006639:Histidine; D008780:Methyltransferases; D009004:Monosaccharide Transport Proteins; D000216:N-Acylneuraminate Cytidylyltransferase; D020652:Peptide Elongation Factor 2; D011506:Proteins; D037521:Virulence Factors; D000097668:Pseudomonas aeruginosa Exotoxin A; D000099237:UDP-Galactose Translocators; C030325:anthrax toxin; C067044:cytolethal distending toxin; C027527:diphthamide; C000706694:DPH7 protein, human