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Various species of the intestinal microbiota have been associated with the development of colorectal cancer1,2, but it has not been demonstrated that bacteria have a direct role in the occurrence of oncogenic mutations. Escherichia coli can carry the pathogenicity island pks, which encodes a set of enzymes that synthesize colibactin3. This compound is believed to alkylate DNA on adenine residues4,5 and induces double-strand breaks in cultured cells3. Here we expose human intestinal organoids to genotoxic pks+E. coli by repeated luminal injection over five months. Whole-genome sequencing of clonal organoids before and after this exposure revealed a distinct mutational signature that was absent from organoids injected with isogenic pks-mutant bacteria. The same mutational signature was detected in a subset of 5,876 human cancer genomes from two independent cohorts, predominantly in colorectal cancer. Our study describes a distinct mutational signature in colorectal cancer and implies that the underlying mutational process results directly from past exposure to bacteria carrying the colibactin-producing pks pathogenicity island. Organoids derived from human intestinal cells that are co-cultured with bacteria carrying the genotoxic pks+ island develop a distinct mutational signature associated with colorectal cancer. [ABSTRACT FROM AUTHOR]

The ordered assembly of a functional preinitiation complex (PIC), composed of general transcription factors (GTFs), is a prerequisite for the transcription of protein-coding genes by RNA polymerase II. TFIID, comprised of the TATA binding protein (TBP) and 13 TBP-associated factors (TAFs), is the GTF that is thought to recognize the promoter sequences allowing site-specific PIC assembly. Transcriptional cofactors, such as SAGA, are also necessary for tightly regulated transcription initiation. The contribution of the two TAF10-containing complexes (TFIID, SAGA) to erythropoiesis remains elusive. By ablating TAF10 specifically in erythroid cells in vivo, we observed a differentiation block accompanied by deregulated GATA1 target genes, including Gata1 itself, suggesting functional cross talk between GATA1 and TAF10. Additionally, we analyzed by mass spectrometry the composition of TFIID and SAGA complexes in mouse and human cells and found that their global integrity is maintained, with minor changes, during erythroid cell differentiation and development. In agreement with our functional data, we show that TAF10 interacts directly with GATA1 and that TAF10 is enriched on the GATA1 locus in human fetal erythroid cells. Thus, our findings demonstrate a cross talk between canonical TFIID and SAGA complexes and cell-specific transcription activators during development and differentiation. [ABSTRACT FROM AUTHOR]

Hematopoietic stem cell (HSC) generation in the aorta-gonad-mesonephros region requires HSC specification signals from the surrounding microenvironment. In zebrafish, PDGF-B/PDGFRβ signaling controls hematopoietic stem/progenitor cell (HSPC) generation and is required in the HSC specification niche. Little is known about murine HSPC specification in vivo and whether PDGF-B/PDGFRβ is involved. Here, we show that PDGFRβ is expressed in distinct perivascular stromal cell layers surrounding the mid-gestation dorsal aorta, and its deletion impairs hematopoiesis. We demonstrate that PDGFRβ+ cells play a dual role in murine hematopoiesis. They act in the aortic niche to support HSPCs, and in addition, PDGFRβ+ embryonic precursors give rise to a subset of HSPCs that persist into adulthood. These findings provide crucial information for the controlled production of HSPCs in vitro. [Display omitted] • PDGFRβ deletion affects hematopoietic development in the AGM in vivo • The transcriptome and hematopoietic support of the PDGFRβ-KO niche are altered • The osteogenic gene profile and differentiation of KO AGM MSCs are affected • PDGFRβ+ early embryonic precursors contribute to EC and HSPC lineages in vivo Sá da Bandeira et al. find that PDGFRβ, expressed in the AGM hematopoietic microenvironment, controls HSPCs. By RNA sequencing, hematopoietic assays, and lineage tracing, they show that PDGFRβ has a dual role: it is required in the AGM hematopoietic niche and marks early precursors of a subset of ECs and HSPCs. [ABSTRACT FROM AUTHOR]

Rad23a and Rad23b proteins are linked to nucleotide excision DNA repair (NER) via association with the DNA damage recognition protein xeroderma pigmentosum group C (XPC) are and known to be implicated in protein turnover by the 26S proteasome. Rad23b-null mice are NER proficient, likely due to the redundant function of the Rad23b paralogue, Rad23a. However, Rad23b-null midgestation embryos are anemic, and most embryos die before birth. Using an unbiased proteomics approach, we found that the majority of Rad23b-interacting partners are associated with the ubiquitin-proteasome system (UPS). We tested the requirement for Rad23b-dependent UPS activity in cellular proliferation and more specifically in the process of erythropoiesis. In cultured fibroblasts derived from embryos lacking Rad23b, proliferation rates were reduced. In fetal livers of Rad23b-null embryos, we observed reduced proliferation, accumulation of early erythroid progenitors, and a block during erythroid maturation. In primary wild-type (WT) erythroid cells, knockdown of Rad23b or chemical inhibition of the proteasome reduced survival and differentiation capability. Finally, the defects linked to Rad23b loss specifically affected fetal definitive erythropoiesis and stress erythropoiesis in adult mice. Together, these data indicate a previously unappreciated requirement for Rad23b and the UPS in regulation of proliferation in different cell types. [ABSTRACT FROM AUTHOR]

The human β-globin locus contains the β-like globin genes (i.e. fetal γ-globin and adult β-globin), which heterotetramerize with α-globin subunits to form fetal or adult hemoglobin. Thalassemia is one of the commonest inherited disorders in the world, which results in quantitative defects of the globins, based on a number of genome variations found in the globin gene clusters. Hereditary persistence of fetal hemoglobin (HPFH) also caused by similar types of genomic alterations can compensate for the loss of adult hemoglobin. Understanding the regulation of the human γ-globin gene expression is a challenge for the treatment of thalassemia. A mouse model that facilitates high-throughput assays would simplify such studies. We have generated a transgenic dual reporter mouse model by tagging the γ- and β-globin genes with GFP and DsRed fluorescent proteins respectively in the endogenous human β-globin locus. Erythroid cell lines derived from this mouse model were tested for their capacity to reactivate the γ-globin gene. Here, we discuss the applications and limitations of this fluorescent reporter model to study the genetic basis of red blood cell disorders and the potential use of such model systems in high-throughput screens for hemoglobinopathies therapeutics. [ABSTRACT FROM AUTHOR]

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