Long-range enhancer-promoter interactions in the HoxD complex

Collinear expression of 5′ Hoxd genes during mammalian limb development is required to properly determine digits number and identity. We recently proposed a two-step model to account for the underlying regulatory mechanism, which involves an initial looping and recognition of the cluster by a complex of enhancer sequences, followed by a micro-scanning of nearby-located genes. This model could account for both the wild-type dosage of the various Hoxd transcripts, as well as for their quantitative variations in several mutant strains. Here, using the Chromosome Conformation Capture (3C) technique, we confirm the initial looping step by showing that the 5′ extremity of the HoxD cluster physically interacts with two previously identified, remote regulatory sequences displaying enhancer activity in developing digits. This spatial conformation is specific to presumptive digits and is not observed in tissues devoid of Hox gene transcription. The flexibility of these long-range interactions is illustrated by the ability of the enhancers to activate reporter transgenes inserted at various positions along the locus. To further define this regulatory landscape, we produced a large chromosomal inversion, separating the reporter transgene from the digit enhancers and HoxD cluster. Surprisingly, this configuration fails to abolish the transgene expression in developing digits, but leads to a major down-regulation of Hoxd genes in this domain, and concurrent morphological defects. These results suggest that the transcriptional activation of Hoxd genes requires the activity of additional and distant regulatory sequences, in addition to the enhancers identified so far by transgenic analysis

1992 års uppföljning av CG-provvägar - Lambohovsleden del 2, Linköping - Östra Ringvägen, Kristinehamn - Lambohovsleden del 3, Linköping

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Heterogeneous combinatorial expression of Hoxd genes in single cells during limb development

Abstract Background Global analyses of gene expression during development reveal specific transcription patterns associated with the emergence of various cell types, tissues, and organs. These heterogeneous patterns are instrumental to ensure the proper formation of the different parts of our body, as shown by the phenotypic effects generated by functional genetic approaches. However, variations at the cellular level can be observed within each structure or organ. In the developing mammalian limbs, expression of Hox genes from the HoxD cluster is differentially controlled in space and time, in cells that will pattern the digits and the forearms. While the Hoxd genes broadly share a common regulatory landscape and large-scale analyses have suggested a homogenous Hox gene transcriptional program, it has not previously been clear whether Hoxd genes are expressed together at the same levels in the same cells. Results We report a high degree of heterogeneity in the expression of the Hoxd11 and Hoxd13 genes. We analyzed single-limb bud cell transcriptomes and show that Hox genes are expressed in specific combinations that appear to match particular cell types. In cells giving rise to digits, we find that the expression of the five relevant Hoxd genes (Hoxd9 to Hoxd13) is unbalanced, despite their control by known global enhancers. We also report that specific combinatorial expression follows a pseudo-time sequence, which is established based on the transcriptional diversity of limb progenitors. Conclusions Our observations reveal the existence of distinct combinations of Hoxd genes at the single-cell level during limb development. In addition, we document that the increasing combinatorial expression of Hoxd genes in this developing structure is associated with specific transcriptional signatures and that these signatures illustrate a temporal progression in the differentiation of these cells

Ligand-dependent contribution of RXRβ to cholesterol homeostasis in Sertoli cells

We show that mice expressing retinoid X receptor β (RXRβ) impaired in its transcriptional activation function AF-2 (Rxrb(af20) mutation) do not display the spermatid release defects observed in RXRβ-null mutants, indicating that the role of RXRβ in spermatid release is ligand-independent. In contrast, like RXRβ-null mutants, Rxrb(af20) mice accumulate cholesteryl esters in Sertoli cells (SCs) due to reduced ABCA1 transporter-mediated cholesterol efflux. We provide genetic and molecular evidence that cholesterol homeostasis in SCs does not require PPARα and β, but depends upon the TIF2 coactivator and RXRβ/LXRβ heterodimers, in which RXRβ AF-2 is transcriptionally active. Our results also indicate that RXRβ may be activated by a ligand distinct from 9-cis retinoic acid