Technologies for Proteome-Wide Discovery of Extracellular Host-Pathogen Interactions

Martinez-Martin, Nadia

Haploid cells, in turn, allow the study of recessive phenotypes that can be masked in diploid cells, due to the difficulties of creating true genetic knockouts in mammalian cells. Despite yeast being a useful tool due to the simplicity of obtaining relevant mutants at its haploid life stage, the majority of human pathogens do not replicate in yeast therefore limiting the applicability of this approach [129] . In recent years, human haploid cells have been increasingly utilized for genome-wide loss-of-function genetic screens using insertional mutagenesis [43, 44, 47] . In initial studies, Carette and colleagues took advantage of the KBM7 cell line, a derivative of the chronic myeloid leukemia cell line (CML) with a haploid karyotype except for chromosome 8 [130] . Using gene-trap retroviruses for efficient insertional mutagenesis, the authors generated a genomewide collection of null mutants for most nonessential genes [43] . This approach was successfully utilized to identify host factors essential for the functions of the distending toxins or CDTs, potent virulence factors secreted by a number of pathogenic bacteria. In particular, mutagenized KBM7 cells were treated with Escherichia coli-derived CDTs and resistant clones were isolated, leading to identification of insertions in the sphingomyelin synthase 1 and the putative G protein-coupled receptor TMEM181, suggesting that this molecule may serve as a surface receptor for the toxin [43, 44] . Similar haploid screens have identified novel receptors for a number of bacterial toxins, including the lipolysis-stimulated lipoprotein receptor for the Clostridium difficile transferase [45] , or the low-density lipoprotein receptor-related protein 1 as a host receptor of the Clostridium perfringens TpeL toxin [46] .