Sociosexual behavior requires both activating and repressive roles of Tfap2e/AP-2ε in vomeronasal sensory neurons

Neuronal identity dictates the position in an epithelium, and the ability to detect, process, and transmit specific signals to specified targets. Transcription factors (TFs) determine cellular identity via direct modulation of genetic transcription and recruiting chromatin modifiers. However, our understanding of the mechanisms that define neuronal identity and their magnitude remains a critical barrier to elucidate the etiology of congenital and neurodegenerative disorders. The rodent vomeronasal organ provides a unique system to examine in detail the molecular mechanisms underlying the differentiation and maturation of chemosensory neurons. Here we demonstrated that the identity of postmitotic/maturing VSNs and vomeronasal dependent behaviors can be reprogrammed through the rescue of AP-2ε expression in the AP-2ε Null mice and by inducing ectopic AP-2ε expression in mature apical VSNs. We suggest that the transcription factor AP-2ε can reprogram VSNs bypassing cellular plasticity restrictions, and that it directly controls the expression of batteries of vomeronasal genes.

B1 SINE-binding ZFP266 impedes reprogramming through suppression of chromatin opening by pioneering factors

Successful generation of induced pluripotent stem cells (iPSCs) via the overexpression of Oct4 (Pou5f1), Sox2, Klf4 and c-Myc (OSKM) highlights the power of transcription factor (TF)-mediated cellular conversions. Nevertheless, iPSC reprogramming is inherently inefficient and understanding the molecular mechanisms underlying this inefficiency holds the key to control cellular identity successfully. Here, we report 16 novel reprogramming roadblock genes identified by CRISPR/Cas9-mediated genome-wide knockout (KO) screening. Of these, disruption of KRAB zinc finger protein (KRAB-ZFP) Zfp266 strongly and consistently enhanced iPSC generation in several iPSC reprogramming settings, emerging as the most robust roadblock. Further analyses revealed that ZFP266 bound Short Interspersed Nuclear Elements (SINEs) adjacent to OSK binding sites and impedes chromatin opening. This work serves as a resource for better understanding reprogramming mechanisms and proposes SINEs as a critical genetic element that regulates chromatin accessibility at enhancers for efficient pluripotency induction.

RNA polymerase II pausing in development: orchestrating transcription

The coordinated regulation of transcriptional networks underpins cellular identity and developmental progression. RNA polymerase II promoter-proximal pausing (Pol II pausing) is a prevalent mechanism by which cells can control and synchronize transcription. Pol II pausing regulates the productive elongation step of transcription at key genes downstream of a variety of signalling pathways, such as FGF and Nodal. Recent advances in our understanding of the Pol II pausing machinery and its role in transcription call for an assessment of these findings within the context of development. In this review, we discuss our current understanding of the molecular basis of Pol II pausing and its function during organismal development. By critically assessing the tools used to study this process we conclude that combining recently developed genomics approaches with refined perturbation systems has the potential to expand our understanding of Pol II pausing mechanistically and functionally in the context of development and beyond.

Long-term culture expanded alveolar macrophages restore full epigenetic identity in vivo

Alveolar macrophages (AM) are tissue resident macrophages of the lung that can be expanded in culture, but it is unknown to what extent culture affects their in vivo identity. Here we show that long-term ex vivo expanded mouse AM (exAM) maintain core AM gene expression but show culture adaptations related to adhesion, metabolism and proliferation. Strikingly, even after several months in culture exAM reacquired full transcriptional and epigenetic identity upon transplantation into the lung and could self-maintain in the natural niche long-term. Changes in open chromatin regions (OCR) observed in culture were fully reversible in transplanted exAM (texAM) and resulted in a gene expression profile indistinguishable from resident AM. Our results demonstrate that long-term proliferation of AM in culture does not compromise cellular identity in vivo. The demonstrated robustness of exAM identity provides new opportunities for mechanistic analysis and highlights the therapeutic potential of ex vivo expanded macrophages.

Chromatin accessibility differences between alpha, beta, and delta cells identifies common and cell type-specific enhancers

High throughput sequencing has enabled the interrogation of the transcriptomic landscape of glucagon-secreting alpha cells, insulin-secreting beta cells, and somatostatin-secreting delta cells. These approaches have furthered our understanding of expression patterns that define healthy or diseased islet cell types and helped explicate some of the intricacies between major islet cell crosstalk and glucose regulation. All three endocrine cell types derive from a common pancreatic progenitor, yet alpha and beta cells have partially opposing functions, and delta cells modulate and control insulin and glucagon release. While gene signatures that define and maintain cellular identity have been widely explored, the underlying epigenetic components are incompletely characterized and understood. Chromatin accessibility and remodeling is a dynamic attribute that plays a critical role to determine and maintain cellular identity. Here, we compare and contrast the chromatin landscape between mouse alpha, beta, and delta cells using ATAC-Seq to evaluate the significant differences in chromatin accessibility. The similarities and differences in chromatin accessibility between these related islet endocrine cells help define their fate in support of their distinct functional roles. We identify patterns that suggest that both alpha and delta cells are poised, but repressed, from becoming beta-like. We also identify patterns in differentially enriched chromatin that have transcription factor motifs preferentially associated with different regions of the genome. Finally, we identify and visualize both novel and previously discovered common endocrine- and cell specific- enhancer regions across differentially enriched chromatin.

Cell-specific alterations in Pitx1 regulatory landscape activation caused by the loss of a single enhancer

Developmental genes are frequently controlled by multiple enhancers sharing similar specificities. As a result, deletions of such regulatory elements have often failed to reveal their full function. Here, we use the Pitx1 testbed locus to characterize in detail the regulatory and cellular identity alterations following the deletion of one of its enhancers (Pen). By combining single cell transcriptomics and an in-embryo cell tracing approach, we observe an increased fraction of Pitx1 non/low-expressing cells and a decreased fraction of Pitx1 high-expressing cells. We find that the over-representation of Pitx1 non/low-expressing cells originates from a failure of the Pitx1 locus to coordinate enhancer activities and 3D chromatin changes. This locus mis-activation induces a localized heterochrony and a concurrent loss of irregular connective tissue, eventually leading to a clubfoot phenotype. This data suggests that, in some cases, redundant enhancers may be used to locally enforce a robust activation of their host regulatory landscapes.

Loss of Polycomb Repressive Complex 2 Function Alters Digestive Organ Homeostasis and Neuronal Differentiation in Zebrafish

Polycomb repressive complex 2 (PRC2) mediates histone H3K27me3 methylation and the stable transcriptional repression of a number of gene expression programs involved in the control of cellular identity during development and differentiation. Here, we report on the generation and on the characterization of a zebrafish line harboring a null allele of eed, a gene coding for an essential component of the PRC2. Homozygous eed-deficient mutants present a normal body plan development but display strong defects at the level of the digestive organs, such as reduced size of the pancreas, hepatic steatosis, and a loss of the intestinal structures, to die finally at around 10–12 days post fertilization. In addition, we found that PRC2 loss of function impairs neuronal differentiation in very specific and discrete areas of the brain and increases larval activity in locomotor assays. Our work highlights that zebrafish is a suited model to study human pathologies associated with PRC2 loss of function and H3K27me3 decrease.

Cancer CRC: A Comprehensive Cancer Core Transcriptional Regulatory Circuit Resource and Analysis Platform

A core transcriptional regulatory circuit (CRC) is a group of interconnected auto-regulating transcription factors (TFs) that form loops and can be identified by super-enhancers (SEs). Studies have indicated that CRCs play an important role in defining cellular identity and determining cellular fate. Additionally, core TFs in CRCs are regulators of cell-type-specific transcriptional regulation. However, a global view of CRC properties across various cancer types has not been generated. Thus, we integrated paired cancer ATAC-seq and H3K27ac ChIP-seq data for specific cell lines to develop the Cancer CRC (http://bio.liclab.net/Cancer_crc/index.html). This platform documented 94,108 cancer CRCs, including 325 core TFs. The cancer CRC also provided the “SE active core TFs analysis” and “TF enrichment analysis” tools to identify potentially key TFs in cancer. In addition, we performed a comprehensive analysis of core TFs in various cancer types to reveal conserved and cancer-specific TFs.