Biology subjects

Barral, A.

Publications and source records attributed to Barral, A..

6 recordsLinked to original sources

bioRxiv2026

DNA replication is dispensable for developmental progression, but required for heterochromatin organization at mouse zygotic genome activation

Early mammalian development progresses through a small number of cleavage divisions with extensive transcriptional and epigenetic reprogramming that result in the very first lineage decisions, leading to blastocyst formation. Although classical embryological studies suggested that developmental progression can occur independently of normal cell division, the extent to which DNA replication contributes to these processes remains unclear. In particular, whether replication is required simply for proliferation or also for establishing the cellular states that regulate stage-specific gene expression is unknown. Here we show that mouse preimplantation development can proceed without DNA replication, but it is required to maintain proper heterochromatin organization at the 2-cell stage, the time of zygotic genome activation. The inhibition of DNA replication with aphidicolin blocked cleavage divisions without preventing zygotic genome activation, embryo morphogenesis, activation of lineage-specification factors, or the establishment of stage-specific transcriptional programs. However, replication arrest selectively altered the expression of developmental genes associated with late-replicating, Polycomb-enriched chromatin domains, and also reduced global levels of H3K27me3 and H3K9me3. These findings indicate that developmental timing in the early embryo is largely uncoupled from DNA replication and cell-cycle progression. Instead, DNA replication contributes to transcriptional fidelity by establishing or maintaining repressive chromatin states during the extensive genome reprogramming that accompanies zygotic genome activation.

developmental biology↗

bioRxiv2025

NR6A1 is essential for neural crest cell specification, formation and survival

Neural crest cells (NCC) are a migratory progenitor cell population considered unique to vertebrates. Derived from the neuroepithelium during early embryogenesis, NCC contribute to nearly every tissue and organ system throughout the body, and disruptions in NCC development can result in congenital disorders, termed neurocristopathies. Despite decades of research, we have a poor understanding of the cellular mechanisms and signals that govern mammalian NCC formation. We discovered nuclear receptor superfamily 6 group member 1 (NR6A1/GCNF/RTR), is a novel, critical regulator of mammalian NCC specification, formation and survival. Nr6a1 is expressed throughout the neuroepithelium in mouse embryos from E8.0 to E9.5 and briefly in newly delaminated NCC. Nr6a1 loss-of-function perturbs anterior cranial NCC formation and survival, and results in the complete agenesis of migratory NCC caudal to the first pharyngeal arch. Using mouse ESC and human iPSC differentiation into NCC, chromatin immunoprecipitation, and multiomic approaches, we demonstrate that these phenotypes are associated with perturbation of NCC specification (Foxd3, Sox9, Sox10) and epithelial-mesenchymal transition (EMT; Snail1, Zeb2), in concert with persistent expression of pluripotency-associated factors (Oct4 and Nanog) in the neuroepithelium. Conditional deletion revealed that Nr6a1 is required during mid-late gastrulation, demonstrating that murine NCC specification occurs earlier than previously thought. Consistent with these observations, in vivo overexpression of Oct4 in gastrulating mouse embryos disrupts NCC specification and formation. Therefore, Nr6a1 is essential for mammalian NCC formation and moreover, may function as a bimodal switch repressing pluripotency-associated factors in the neuroepithelium, while concomitantly activating NCC specifiers and regulators of EMT.

developmental biology↗

bioRxiv2020

Enhancer priming by H3K4 methylation safeguards germline competence

Germline specification in mammals occurs through an inductive process whereby competent cells in the post-implantation epiblast differentiate into primordial germ cells (PGC). The intrinsic factors that endow epiblast cells with the competence to respond to germline inductive signals remain unknown. Single-cell RNA sequencing across multiple stages of an in vitro PGC-like cells (PGCLC) differentiation system shows that PGCLC genes initially expressed in the naive pluripotent stage become homogeneously dismantled in germline competent epiblast like-cells (EpiLC). In contrast, the decommissioning of enhancers associated with these germline genes is incomplete. Namely, a subset of these enhancers partly retain H3K4me1, accumulate less heterochromatic marks and remain accessible and responsive to transcriptional activators. Subsequently, as in vitro germline competence is lost, these enhancers get further decommissioned and lose their responsiveness to transcriptional activators. Importantly, using H3K4me1 deficient cells, we show that the loss of this histone modification reduces the germline competence of EpiLC and decreases PGCLC differentiation efficiency. Our work suggests that, although H3K4me1 might not be essential for enhancer function, it can facilitate the (re)activation of enhancers and the establishment of gene expression programs during specific developmental transitions.

genomics↗

bioRxiv2019

Nanog regulates Pou3f1 expression and represses anterior fate at the exit from pluripotency.

Pluripotency is regulated by a network of transcription factors that maintains early embryonic cells in an undifferentiated state while allowing them to proliferate. NANOG is a critical factor for maintaining pluripotency and its role in primordial germ cell differentiation has been well described. However, Nanog is expressed during gastrulation across all the posterior epiblast, and only later in development its expression is restricted to primordial germ cells. In this work, we unveiled a previously unknown mechanism by which Nanog specifically represses the anterior epiblast lineage. Analysis of transcriptional data from both embryonic stem cells and gastrulating mouse embryos revealed Pou3f1 expression to be negatively correlated with that of Nanog during the early stages of differentiation. We have functionally demonstrated Pou3f1 to be a direct target of NANOG by using a dual transgene system for the controlled expression of Nanog. Use of Nanog null ES cells further demonstrated a role for Nanog in repressing anterior neural genes. Deletion of a NANOG binding site (BS) located nine kilobases downstream of the transcription start site of Pou3f1 revealed this BS to have a specific role in the regionalization of the expression of this gene in the embryo. Our results indicate an active role of Nanog inhibiting the neural fate by repressing Pou3f1 at the onset of gastrulation.

developmental biology↗

bioRxiv2019

The Long Pentraxin 3 (PTX3) Suppresses Immunity to Cutaneous Leishmaniasis by Negatively Regulating Th17 Response

The long Pentraxin 3 (PTX3), a soluble pattern recognition molecule, plays a critical role in inflammation, tissue repair and wound healing. Here, we show that PTX3 regulates disease pathogenesis in cutaneous leishmaniasis (CL). PTX3 expression is increased in active skin lesions in patients and mice during CL, with higher levels being expressed in individuals with severe disease. PTX3 deficient (PTX3-/-) mice were highly resistant to L. major infection and the enhanced resistance was associated with increased IL-17 response. Neutralization of IL-17A abolished this enhanced resistance while treatment with recombinant PTX3 resulted in reduced IL-17A response and increased susceptibility to L. major infection. Naive CD4+ T cells from PTX3-/- mice displayed increased differentiation into Th17 cells, which was reversed in the presence of recombinant PTX3. The enhanced Th17 response observed in PTX3-/- cells was associated with increased Leishmania specific IL-6 production from dendritic cells along with enhanced expression of Th17-specific transcription factors including ROR{gamma}t, AhR and STAT3. Addition of recombinant PTX3 significantly inhibited the expression of Th17-specific transcription factors and dramatically reduced the frequency of Th17 cells in Th17-polarizing cultures of PTX3-/- CD4+ T cells. Collectively, our results show that PTX3 contributes to the pathogenesis of CL by suppressing Th17 differentiation and IL-17A production.\n\nAuthor SummaryCutaneous leishmaniasis (CL) is caused by several species of Leishmania. Currently, there is no approved vaccine against human CL because of the poor understanding of the mechanisms that regulate disease pathogenesis and correlates of protective immunity. Because the long pentraxin 3 (PTX3, a soluble pattern recognition molecule that forms an integral part of the host innate immunity), regulates inflammation and tissue repair, which are critical physiological events associated with resolution of skin lesions during CL, we investigated its role in disease pathogenesis.\n\nHere, we show that PTX3 levels were elevated in skin-lesions in patients and mice during CL. Using a loss of function approach, we showed that PTX3 contributes to pathogenesis, and this was associated with increased IL-17A responses. Neutralization and recombinant cytokine treatment studies showed that the increased resistance of PTX3 deficient mice to L. major is due to enhanced Th17 response in these mice. We further show that PTX3 negatively regulates IL-6 production by dendritic cells and the expression of IL-17A-specific transcription factors (including ROR{gamma}T, STAT3, IRF4, BATF and AhR) in CD4+ T cells. Collectively, these findings show that PTX3 is a negative regulator of Th17 response and protective immunity during L. major infection.

immunology↗

bioRxiv2019

Pluripotency factors regulate the onset of Hox cluster activation in the early embryo

Pluripotent cells are a transient population present in the early mammalian embryo dependent on transcription factors, such as OCT4 and NANOG, which maintain pluripotency while simultaneously suppressing lineage specification. Interestingly, these factors are not exclusive to uncommitted cells, but are also expressed during early phases of differentiation. However, their role in the transition from pluripotency to lineage specification is largely unknown. Using genetic models for controlled Oct4 or Nanog expression during postimplantation stages, we found that pluripotency factors play a dual role in regulating key lineage specifiers, initially repressing their expression and later being required for their proper activation. We show that the HoxB cluster is coordinately regulated in this way by OCT4 binding sites located at the 3 end of the cluster. Our results show that core pluripotency factors are not limited to maintaining the pre-committed epiblast, but are also necessary for the proper deployment of subsequent developmental programs.

developmental biology↗