Hypoxia induces an early primitive streak signature, enhancing spontaneous elongation and lineage representation in gastruloids.

The cellular microenvironment, together with intrinsic regulators, shapes stem cell identity and differentiation capacity. Mammalian early embryos are exposed to hypoxia in vivo and appear to benefit from hypoxic culture in vitro. Yet, how hypoxia influences stem cell transcriptional networks and lineage choices remain poorly understood. Here, we investigated the molecular effects of acute and prolonged hypoxia on embryonic and extra-embryonic stem cells as well as the functional impact on differentiation potential. We find a temporal and cell type-specific transcriptional response including an early primitive streak signature in hypoxic embryonic stem cells mediated by HIF1α. Using a 3D gastruloid differentiation model, we show that hypoxia-induced T expression enables symmetry breaking and axial elongation in the absence of exogenous WNT activation. When combined with exogenous WNT activation, hypoxia enhances lineage representation in gastruloids, as demonstrated by highly enriched signatures of gut endoderm, notochord, neuromesodermal progenitors and somites. Our findings directly link the microenvironment to stem cell function and provide a rationale supportive of applying physiological conditions in models of embryo development.

Identifying a novel role for the master regulator Tal1 in the Endothelial to Hematopoietic Transition.

Progress in the generation of Hematopoietic Stem and Progenitor Cells (HSPCs) in vitro and ex vivo has been built on the knowledge of developmental hematopoiesis, underscoring the importance of understanding this process. HSPCs emerge within the embryonic vasculature through an Endothelial-to-Hematopoietic Transition (EHT). The transcriptional regulator Tal1 exerts essential functions in the earliest stages of blood development, but is considered dispensable for the EHT. Nevertheless, Tal1 is expressed with its binding partner Lmo2 and it homologous Lyl1 in endothelial and transitioning cells at the time of EHT. Here, we investigated the function of these genes using a mouse embryonic-stem cell (mESC)-based differentiation system to model hematopoietic development. We showed for the first time that the expression of TAL1 in endothelial cells is crucial to ensure the efficiency of the EHT process and a sustained hematopoietic output. Our findings uncover an important function of Tal1 during the EHT, thus filling the current gap in the knowledge of the role of this master gene throughout the whole process of hematopoietic development.

TADs enriched in histone H1.2 strongly overlap with the B compartment, inaccessible chromatin, and AT-rich Giemsa bands.

Giemsa staining of metaphase chromosomes results in a characteristic banding useful for identification of chromosomes and its alterations. We have investigated in silico whether Giemsa bands (G bands) correlate with epigenetic and topological features of the interphase genome. Staining of G-positive bands decreases with GC content; nonetheless, G-negative bands are GC heterogeneous. High GC bands are enriched in active histone marks, RNA polymerase II, and SINEs and associate with gene richness, gene expression, and early replication. Low GC bands are enriched in repressive marks, lamina-associated domains, and LINEs. Histone H1 variants distribute heterogeneously among G bands: H1X is enriched at high GC bands and H1.2 is abundant at low GC, compacted bands. According to epigenetic features and H1 content, G bands can be organized in clusters useful to compartmentalize the genome. Indeed, we have obtained Hi-C chromosome interaction maps and compared topologically associating domains (TADs) and A/B compartments to G banding. TADs with high H1.2/H1X ratio strongly overlap with B compartment, late replicating, and inaccessible chromatin and low GC bands. We propose that GC content is a strong driver of chromatin compaction and 3D genome organization, that Giemsa staining recapitulates this organization denoted by high-throughput techniques, and that H1 variants distribute at distinct chromatin domains. DATABASES: Hi-C data on T47D breast cancer cells have been deposited in NCBI's Gene Expression Omnibus and are accessible through GEO Series accession number GSE147627.

Single-cell transcriptomics reveals a new dynamical function of transcription factors during embryonic hematopoiesis.

Recent advances in single-cell transcriptomics techniques have opened the door to the study of gene regulatory networks (GRNs) at the single-cell level. Here, we studied the GRNs controlling the emergence of hematopoietic stem and progenitor cells from mouse embryonic endothelium using a combination of single-cell transcriptome assays. We found that a heptad of transcription factors (Runx1, Gata2, Tal1, Fli1, Lyl1, Erg and Lmo2) is specifically co-expressed in an intermediate population expressing both endothelial and hematopoietic markers. Within the heptad, we identified two sets of factors of opposing functions: one (Erg/Fli1) promoting the endothelial cell fate, the other (Runx1/Gata2) promoting the hematopoietic fate. Surprisingly, our data suggest that even though Fli1 initially supports the endothelial cell fate, it acquires a pro-hematopoietic role when co-expressed with Runx1. This work demonstrates the power of single-cell RNA-sequencing for characterizing complex transcription factor dynamics.