CRISPR screens identify gene targets at breast cancer risk loci

Background: Genome-wide association studies (GWAS) have identified > 200 loci associated with breast cancer risk. The majority of candidate causal variants are in noncoding regions and likely modulate cancer risk by regulating gene expression. However, pinpointing the exact target of the association, and identifying the phenotype it mediates, is a major challenge in the interpretation and translation of GWAS. Results: Here, we show that pooled CRISPR screens are highly effective at identifying GWAS target genes and defining the cancer phenotypes they mediate. Following CRISPR mediated gene activation or suppression, we measure proliferation in 2D, 3D, and in immune-deficient mice, as well as the effect on DNA repair. We perform 60 CRISPR screens and identify 20 genes predicted with high confidence to be GWAS targets that promote cancer by driving proliferation or modulating the DNA damage response in breast cells. We validate the regulation of a subset of these genes by breast cancer risk variants. Conclusions: We demonstrate that phenotypic CRISPR screens can accurately pinpoint the gene target of a risk locus. In addition to defining gene targets of risk loci associated with increased breast cancer risk, we provide a platform for identifying gene targets and phenotypes mediated by risk variants.Natasha K. Tuano, Jonathan Beesley, Murray Manning, Wei Shi, Laura Perlaza, Jimenez, Luis F. Malaver, Ortega, Jacob M. Paynter, Debra Black, Andrew Civitarese, Karen McCue, Aaron Hatzipantelis, Kristine Hillman, Susanne Kaufmann, Haran Sivakumaran, Jose M. Polo, Roger R. Reddel, Vimla Band, Juliet D. French, Stacey L. Edwards, David R. Powell, Georgia Chenevix, Trench, and Joseph Rosenblu

Excavation of an early 17th-century glassmaking site at Glasshouse, Shinrone, Co. Offaly, Ireland

An archaeological research excavation was conducted in the area immediately surrounding an upstanding glassmaking furnace near Shinrone, Co. Offaly, Ireland. It dates to the early to mid 17th century and was built and operated by French Huguenots, probably de Hennezells (de Hennezel/Henzeys/Hensie) who had settled in this region as part of the Crown plantation of King’s County (now Co. Offaly). This furnace, which employed wood rather than coal as a fuel, is a very rare survival, with no other upstanding examples known in Ireland, Britain or the Lorraine region of France where the form probably originated

Comprehensive characterization of distinct states of human naive pluripotency generated by reprogramming

Recent reports on the characteristics of naive human pluripotent stem cells (hPSCs) obtained using independent methods differ. Naive hPSCs have been mainly derived by conversion from primed hPSCs or by direct derivation from human embryos rather than by somatic cell reprogramming. To provide an unbiased molecular and functional reference, we derived genetically matched naive hPSCs by direct reprogramming of fibroblasts and by primed-to-naive conversion using different naive conditions (NHSM, RSeT, 5iLAF and t2iLGöY). Our results show that hPSCs obtained in these different conditions display a spectrum of naive characteristics. Furthermore, our characterization identifies KLF4 as sufficient for conversion of primed hPSCs into naive t2iLGöY hPSCs, underscoring the role that reprogramming factors can play for the derivation of bona fide naive hPSCs.Xiaodong Liu, Christian M Nefzger, Fernando J Rossello, Joseph Chen, Anja S Knaupp, Jaber Firas ... et al

Intestinal stem cell aging signature reveals a reprogramming strategy to enhance regenerative potential

The impact of aging on intestinal stem cells (ISCs) has not been fully elucidated. In this study, we identified widespread epigenetic and transcriptional alterations in old ISCs. Using a reprogramming algorithm, we identified a set of key transcription factors (Egr1, Irf1, FosB) that drives molecular and functional differences between old and young states. Overall, by dissecting the molecular signature of aged ISCs, our study identified transcription factors that enhance the regenerative capacity of ISCs.Christian M. Nefzger, Thierry Jardé, Akanksha Srivastava, Jan Schroeder, Fernando J. Rossello, Katja Horvay, Mirsada Prasko, Jacob M. Paynter, Joseph Chen, Chen-Fang Weng, Yu B. Y. Sun, Xiaodong Liu, Eva Chan, Nikita Deshpande, Xiaoli Chen, Y. Jinhua Li, Jahnvi Pflueger, Rebekah M. Engel, Anja S. Knaupp, Kirill Tsyganov, Susan K. Nilsson, Ryan Lister, Owen J. L. Rackham, Helen E. Abud, and Jose M. Pol

Reprogramming roadmap reveals route to human induced trophoblast stem cells

The reprogramming of human somatic cells to primed or naive induced pluripotent stem cells recapitulates the stages of early embryonic development¹⁻⁶. The molecular mechanism that underpins these reprogramming processes remains largely unexplored, which impedes our understanding and limits rational improvements to reprogramming protocols. Here, to address these issues, we reconstruct molecular reprogramming trajectories of human dermal fibroblasts using single-cell transcriptomics. This revealed that reprogramming into primed and naive pluripotency follows diverging and distinct trajectories. Moreover, genome-wide analyses of accessible chromatin showed key changes in the regulatory elements of core pluripotency genes, and orchestrated global changes in chromatin accessibility over time. Integrated analysis of these datasets revealed a role for transcription factors associated with the trophectoderm lineage, and the existence of a subpopulation of cells that enter a trophectoderm-like state during reprogramming. Furthermore, this trophectoderm-like state could be captured, which enabled the derivation of induced trophoblast stem cells. Induced trophoblast stem cells are molecularly and functionally similar to trophoblast stem cells derived from human blastocysts or first-trimester placentas7. Our results provide a high-resolution roadmap for the transcription-factor-mediated reprogramming of human somatic cells, indicate a role for the trophectoderm-lineage-specific regulatory program during this process, and facilitate the direct reprogramming of somatic cells into induced trophoblast stem cells.Xiaodong Liu … Jose M. Polo … et al