Project description:We undertook a unbiased genome-wide haploid genetic screen to identify new components in interferon lambda signaling. In addition, we performed a genome-wide screen to identify genes that repress spontaneous activation of interferon stimulated genes in the absence of interferon. Both of these screens were performed using a HAP1 cell line containing GFP reporter under the transcriptional regulation of the Interferon-Stimulated Response Element from IFIT2. We also overexpressed IL28RA (IFNLR1) in this cell line, in order to sensitize the cells to type III interferon
Project description:To search for factors regulating neuronal differentiation, we performed a genome-wide loss-of-function CRISPR/Cas9 screen in haploid human ESCs. The regulators were identified by the quantification of depletion of their mutant clones within a pooled loss-of-function library upon neuronal differentiation.
Project description:To search for factors regulating paternally imprinted genes (PEGs), we performed a genome-wide loss-of-function CRISPR/Cas9 screen in haploid parthenogenetic ESCs. This by staining a pooled CRISPR library with a PEG10 antibody and next FACS-sorted for cells that presented de-novo PEG10 expression.
Project description:Whole chromosome losses resulting in near-haploid karyotypes are found in a rare subgroup of treatment-refractory acute lymphoblastic leukemia. To systematically dissect the unique physiology and uncover susceptibilities that can be exploited in near-haploid leukemia, we leveraged single-cell transcriptomics and computational inference of cell cycle stages to pinpoint key differences between near-haploid and diploid leukemia cells. Combining cell cycle stage-specific differential expression with gene essentiality metrics obtained from a genome-wide CRISPR-Cas9-mediated knockout screen, we identified the homologous recombination pathway component RAD51B as an essential gene in near-haploid leukemia. DNA damage analyses revealed significantly increased sensitivity of RAD51-mediated repair to RAD51B loss in the G2/M stage of near-haploid cells, suggesting a unique role of RAD51B in the homologous recombination pathway. We observed elevated G2/M checkpoint signaling as well as a RAD51B signature expression program in response to chemotherapy in a xenograft model of human near-haploid B-ALL, as well as overexpression of RAD51B and its associated programs in a large panel of near-haploid B-ALL patients. These data highlight a unique genetic dependency on DNA repair machinery in near-haploid leukemia and demarcate RAD51B as a promising candidate for targeted therapy in this treatment-resistant disease.
