Abstract
Mammalian development is regulated by the interplay of tissue-specific and ubiquitously expressed transcription factors, such as Sp1. Sp1 knockout mice die in utero with multiple phenotypic aberrations, but the underlying molecular mechanism of this differentiation failure has been elusive. Here, we have used conditional knockout mice as well as the differentiation of mouse ES cells as a model with which to address this issue. To this end, we examined differentiation potential, global gene expression patterns and Sp1 target regions in Sp1 wild-type and Sp1-deficient cells representing different stages of hematopoiesis. Sp1−/− cells progress through most embryonic stages of blood cell development but cannot complete terminal differentiation. This failure to fully differentiate is not seen when Sp1 is knocked out at later developmental stages. For most Sp1 target and non-target genes, gene expression is unaffected by Sp1 inactivation. However, Cdx genes and multiple Hox genes are stage-specific targets of Sp1 and are downregulated at an early stage. As a consequence, expression of genes involved in hematopoietic specification is progressively deregulated. Our work demonstrates that the early absence of active Sp1 sets a cascade in motion that culminates in a failure of terminal hematopoietic differentiation and emphasizes the role of ubiquitously expressed transcription factors for tissue-specific gene regulation. In addition, our global side-by-side analysis of the response of the transcriptional network to perturbation sheds a new light on the regulatory hierarchy of hematopoietic specification.
Highlights
Changes in gene expression programs during cell differentiation are regulated by the interplay of ubiquitous and tissue-specific transcription factors, and the epigenetic regulatory machinery
The absence of Sp1 DNA binding activity affects multiple hematopoietic lineages In the past decade, a number of attempts have been made to dissect the molecular mechanism of the developmental arrest caused by lack of Sp1 DNA-binding activity, using conditional knockout mice and CRE-recombinase enzyme expressed from different types of tissue-specific promoters
This opened the possibility of using the differentiation of such cells in vitro to obtain molecular insights into the molecular mechanisms of differentiation perturbed by the lack of Sp1 activity
Summary
Changes in gene expression programs during cell differentiation are regulated by the interplay of ubiquitous and tissue-specific transcription factors, and the epigenetic regulatory machinery. The effect of removing tissue-specific regulators for a specific pathway leads to dramatic alterations in gene expression and often a block in differentiation at the point where this factor is crucially needed The reason for this could be that such factors nucleate shifts in transcription factor assemblies to alter cell fates (Heinz et al, 2010; Lichtinger et al, 2012). Sp1-deficient embryos die very early in development with variable phenotypes that range from developmental arrest as an amorphous cell mass to retarded embryos with a number of recognizable structures (Marin et al, 1997) Such a pleiotropic phenotype that affects multiple cell types is seen in a number of knockout mice carrying mutations in global regulators, but to investigate the molecular mechanisms that underlie how such factors are involved in the regulation of multiple genes in specific tissues is an extremely difficult task.
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