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Objective: To develop a multi‐step workflow for the isolation of circulating extravillous trophoblasts (cEVTs) by describing the key steps enabling a semi‐automated process, including a proprietary algorithm for fetal cell origin genetic confirmation and copy number variant (CNV) detection. Methods: Determination of the limit of detection (LoD) for submicroscopic CNV was performed by serial experiments with genomic DNA and single cells from Coriell cell line biobank with known imbalances of different sizes. A pregnancy population of 372 women was prospectively enrolled and blindly analyzed to evaluate the current workflow. Results: An LoD of 800 Kb was demonstrated with Coriell cell lines. This level of resolution was confirmed in the clinical cohort with the identification of a pathogenic CNV of 800 Kb, also detected by chromosomal microarray. The mean number of recovered cEVTs was 3.5 cells per sample with a significant reverse linear trend between gestational age and cEVT recovery rate and number of recovered cEVTs. In twin pregnanices, evaluation of zygosity, fetal sex and copy number profiling was performed in each individual cell. Conclusion: Our semi‐automated methodology for the isolation and single‐cell analysis of cEVTS supports the feasibility of a cell‐based noninvasive prenatal test for fetal genomic profiling. Key points: What's already known about this topic?Fetal circulating extravillous trophoblasts (cEVTs) have been isolated from maternal blood for both aneuploidy and copy number variant (CNV) classification.Previous reported methodologies require mostly manual low throughput workflows and identify putative fetal cells based on immunophenotype. What does this study add?We present a novel semi‐automated workflow for the isolation of circulating fetal trophoblasts and single‐cell genomic profiling of submicroscopic variants.With this methodology, the feasibility of fetal genetic confirmation of each individual cell, a limit of detection (LoD) for CNVs down to 800Kb and the assessment of zygosity, fetal sex and genomic profiling in the same cell in twin pregnancies was demonstrated. [ABSTRACT FROM AUTHOR]

Oncogene-induced DNA damage elicits genomic instability in epithelial cancer cells, but apoptosis is blocked through inactivation of the tumor suppressor p53. In hematological cancers, the relevance of ongoing DNA damage and the mechanisms by which apoptosis is suppressed are largely unknown. We found pervasive DNA damage in hematologic malignancies, including multiple myeloma, lymphoma and leukemia, which leads to activation of a p53-independent, proapoptotic network centered on nuclear relocalization of ABL1 kinase. Although nuclear ABL1 triggers cell death through its interaction with the Hippo pathway coactivator YAP1 in normal cells, we show that low YAP1 levels prevent nuclear ABL1-induced apoptosis in these hematologic malignancies. YAP1 is under the control of a serine-threonine kinase, STK4. Notably, genetic inactivation of STK4 restores YAP1 levels, triggering cell death in vitro and in vivo. Our data therefore identify a new synthetic-lethal strategy to selectively target cancer cells presenting with endogenous DNA damage and low YAP1 levels., In epithelial cancers, rampant DNA double-strand break (DSB) formation followed by activation of the DNA damage response (DDR) occurs in both premalignant and malignant conditions. However, in precancerous settings, senescence [...]

Structural integrity and cellular homeostasis of the embryonic stem cell niche are critical for normal tissue development. In the telencephalic neuroepithelium, this is controlled in part by cell adhesion molecules and regulators of progenitor cell lineage, but the specific orchestration of these processes remains unknown. Here, we studied the role of microRNAs in the embryonic telencephalon as key regulators of gene expression. By using the early recombiner Rx‐Cre mouse, we identify novel and critical roles of miRNAs in early brain development, demonstrating they are essential to preserve the cellular homeostasis and structural integrity of the telencephalic neuroepithelium. We show that Rx‐Cre;DicerF/F mouse embryos have a severe disruption of the telencephalic apical junction belt, followed by invagination of the ventricular surface and formation of hyperproliferative rosettes. Transcriptome analyses and functional experiments in vivo show that these defects result from upregulation of Irs2 upon loss of let‐7 miRNAs in an apoptosis‐independent manner. Our results reveal an unprecedented relevance of miRNAs in early forebrain development, with potential mechanistic implications in pediatric brain cancer. Synopsis: Dicer1‐dependent let‐7 microRNAs are critical to repress p53 pathway activity and Irs2 expression levels in the early telencephalic neuroepithelium, which otherwise enters into hyperproliferation and loss of adhesion of progenitor cells, leading to the formation of true rosettes reminiscent of pediatric cancers. Loss of miRNAs in the early embryonic telencephalon of Rx‐Cre;Dicer1flox mutant mice produces massive transient apoptosis of progenitor cells, loss of adherens junctions and increased proliferation, leading to the formation of true hyperproliferative rosettes.Rx‐Dicer mutants have a significant loss of let‐7 family microRNAs, and increased p53 signaling and Irs2 expression. Loss of p53 in Rx‐Dicer mutants rescues their phenotype in a partially‐penetrant manner.Increased Irs2 expression or blockade of let‐7 activity in early wild‐type embryos, induce rosette formation in the rostral telencephalon, rescued by overexpression of let‐7 and downregulation of Irs‐2, respectively.Let‐7 miRNAs downregulate the amplification of human cortical progenitor cells by repressing Irs2 expression, consistent with a role of this signaling axis in pediatric cancers with true rosettes. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are short (~22 nt) single-stranded non-coding RNAs that regulate gene expression at the post-transcriptional level. Over the past years, many studies have extensively characterized the involvement of miRNA-mediated regulation in neurogenesis and brain development. However, a comprehensive catalog of cortical miRNAs cell-specifically expressed in progenitor types of the developing mammalian cortex is still missing. Overcoming this limitation, here we exploited a double reporter mouse line previously validated by our group to allow the identification of the transcriptional signature to neurogenic commitment and provide the field with the complete atlas of miRNAs expression in proliferating neural stem cells, neurogenic progenitors and newborn neurons during corticogenesis. By extending the currently known list of miRNAs expressed in the mouse brain by over two fold, our study highlights the power of cell type-specific analyses for the detection of transcripts that would otherwise be diluted out when studying bulk tissues. We further exploited our data by predicting putative novel miRNAs and validated the power of our approach by providing novel evidence for the involvement of miR-486 as a novel player in brain development. [ABSTRACT FROM AUTHOR]

A letter to the editor is presented related to the article that studies that association between Complement Factor H-related protein 1 (CFHR1) gene homozygous deletion and acute myelogenous leukemia.

HiChIP is a novel method for the analysis of chromatin interactions based on in situ Hi-C that adds an immuno-precipitation (ChIP) step for the investigation of chromatin structures driven by specific proteins. This approach has been shown to be very efficient as it reliably reproduces Hi-C results and displays a higher rate of informative reads with a required lower amount of input cells when compared with other ChIP-based techniques (as ChIA-PET). Although HiChIP data preprocessing can be performed with the same methods developed for other Hi-C techniques, the identification of chromatin interactions needs to take into account specific biases introduced by the ChIP step. In this chapter we describe a computational pipeline for the analysis of HiChIP data obtained with the immuno-precipitation of Rad21 (part of the cohesin complex) in human embryonic stem cells before and after heat-shock treatment. We provide a detailed description of the preprocessing of raw data, the identification of chromatin interactions, the evaluation of the alterations induced by treatment, and, finally, the visualization of differential loops., (© 2022. Springer Science+Business Media, LLC, part of Springer Nature.)