Abstract
The development of forward genetic screens in human haploid cells has the potential to transform our understanding of the genetic basis of cellular processes unique to man. So far, this approach has been limited mostly to the identification of genes that mediate cell death in response to a lethal agent, likely due to the ease with which this phenotype can be observed. Here, we perform the first reporter screen in the near-haploid KBM7 cell line to identify constitutive inhibitors of NF-κB. CYLD was the only currently known negative regulator of NF-κB to be identified, thus uniquely distinguishing this gene. Also identified were three genes with no previous known connection to NF-κB. Our results demonstrate that reporter screens in haploid human cells can be applied to investigate the many complex signaling pathways that converge upon transcription factors.
Highlights
Forward genetic screens are a powerful means to decipher a biological process without any prior knowledge or assumptions
All screens in human haploid cells performed to date have relied on intrinsic phenotypes, such as sensitivity to toxins or protein surface expression, both of which can be observed at a cellular level
Most genetic screens performed in human haploid cells have sought to identify components in pathways required for cell death in response to a lethal insult
Summary
Forward genetic screens are a powerful means to decipher a biological process without any prior knowledge or assumptions. Such screens are performed in yeast, Drosophila, Caenorhabditis elegans and other genetic model organisms to identify new gene functions. Application of this method to human cultured cells allows the dissection of pathways that are dissimilar or even absent in other model organisms. It may enable the discovery of novel drug targets to treat disease. The recent isolation of human cells lines that are nearly or completely haploid (KBM7 and HAP1, respectively) has revolutionized human forward genetic screens and led to the identification of numerous human host factors required for infection by pathogens and intoxication by bacterial toxins [1,2,3,4,5,6,7]
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