Denis Duboule

The Orphan Nuclear Receptor REV-ERBα Controls Circadian Transcription within the Positive Limb of the Mammalian Circadian Oscillator

Mammalian circadian rhythms are generated by a feedback loop in which BMAL1 and CLOCK, players of the positive limb, activate transcription of the cryptochrome and period genes, components of the negative limb. Bmal1 and Per transcription cycles display nearly opposite phases and are thus governed by different mechanisms. Here, we identify the orphan nuclear receptor REV-ERBalpha as the major regulator of cyclic Bmal1 transcription. Circadian Rev-erbalpha expression is controlled by components of the general feedback loop. Thus, REV-ERBalpha constitutes a molecular link through which components of the negative limb drive antiphasic expression of components of the positive limb. While REV-ERBalpha influences the period length and affects the phase-shifting properties of the clock, it is not required for circadian rhythm generation.

The structural and functional organization of the murine HOX gene family resembles that of Drosophila homeotic genes

This paper reports the cloning of the fourth major murine homeogene complex, HOX‐5. The partial characterization of this gene cluster revealed the presence of two novel genes (Hox‐5.2, Hox‐5.3) located at the 5′ extremity of this complex. In situ hybridization experiments showed that these two genes are transcribed in very posterior domains during embryonic and foetal development. We also show that Hox‐1.6, the gene located at the 3′ most position in the HOX‐1 complex, has a very anterior expression boundary during early development. These results clearly support the recently proposed hypothesis that the expression of murine Antp‐like homeobox‐containing genes along the antero‐posterior developing body axis follows a positional hierarchy which reflects their respective physical positions within the HOX clusters, similar to that which is found for the Drosophila homeotic genes. Such a structural and functional organization is likely conserved in most vertebrates. Moreover, on the basis of sequence comparisons, we propose that the ordering of homeobox‐containing genes within clusters has been conserved between Drosophila and the house mouse. Thus, very different body plans might be achieved, both in insects and vertebrates, by evolutionarily conserved gene networks possibly displaying similar regulatory interactions.

A Global Control Region Defines a Chromosomal Regulatory Landscape Containing the HoxD Cluster

During limb development, coordinated expression of several Hoxd genes is required in presumptive digits. We searched for the underlying control sequences upstream from the cluster and found Lunapark (Lnp), a gene which shares limb and CNS expression specificities with both Hoxd genes and Evx2, another gene located nearby. We used a targeted enhancer-trap approach to identify a DNA segment capable of directing reporter gene expression in both digits and CNS, following Lnp, Evx2, and Hoxd-specific patterns. This DNA region showed an unusual interspecies conservation, including with its pufferfish counterpart. It contains a cluster of global enhancers capable of controlling transcription of several genes unrelated in structure or function, thus defining large regulatory domains. These domains were interrupted in the Ulnaless mutation, a balanced inversion that modified the topography of the locus. We discuss the heuristic value of these results in term of locus specific versus gene-specific regulation.

Disruption of the Hoxd-13 gene induces localized heterochrony leading to mice with neotenic limbs

Vertebrate Hoxd genes are sequentially activated during the morphogenesis and pattern formation of the limb. Using the approach of gene disruption via homologous recombination in embryonic stem cells, we have assessed the function of the last gene of the complex, Hoxd-13. Mutant mice displayed skeletal alterations along all body axes suggesting the existence of a general multiaxial patterning system. In limbs, abnormalities such as a reduction in the length of some bony elements, loss of phalanges, bone fusions, and the presence of an extra element were observed. We propose that the mutation induces local heterochrony, as illustrated by an important retardation in limb morphogenesis. The relevance of these observations to our understanding of the development and evolution of the tetrapod limb is discussed.

Impaired skin wound healing in peroxisome proliferator–activated receptor (PPAR)α and PPARβ mutant mice

We show here that the α, β, and γ isotypes of peroxisome proliferator–activated receptor (PPAR) are expressed in the mouse epidermis during fetal development and that they disappear progressively from the interfollicular epithelium after birth. Interestingly, PPARα and β expression is reactivated in the adult epidermis after various stimuli, resulting in keratinocyte proliferation and differentiation such as tetradecanoylphorbol acetate topical application, hair plucking, or skin wound healing. Using PPARα, β, and γ mutant mice, we demonstrate that PPARα and β are important for the rapid epithelialization of a skin wound and that each of them plays a specific role in this process. PPARα is mainly involved in the early inflammation phase of the healing, whereas PPARβ is implicated in the control of keratinocyte proliferation. In addition and very interestingly, PPARβ mutant primary keratinocytes show impaired adhesion and migration properties. Thus, the findings presented here reveal unpredicted roles for PPARα and β in adult mouse epidermal repair.

Colinearity and functional hierarchy among genes of the homeotic complexes

Homeotic genes identify structures along the anterior to posterior axis during the development of most animals. These genes are clustered into complexes, and their positions within the cluster correlates with their time of expression and the positions of the anterioposterior boundaries of their expression domains. Functional analyses have revealed that this specific genetic order also coincides with a functional hierarchy among members of these complexes, so that the products of more posterior genes in the cluster tend to be prevalent over those of more anterior genes.

Of fingers, toes and penises

Vertebrate Hox genes are essential for limb development. The posterior-most Hoxd and Hoxa genes are required for growth and patterning of digits and are also strongly expressed in the genital bud, which gives rise to the urogenital system, including the penis. Here, we show that removal of posterior Hox gene function results in a concomitant loss of digits and genital bud-derivatives, illustrating that similar developmental mechanisms are at work in these different buds.

Murine genes related to the Drosophila AbdB homeotic genes are sequentially expressed during development of the posterior part of the body

The cloning, characterization and developmental expression patterns of two novel murine Hox genes, Hox‐4.6 and Hox‐4.7, are reported. Structural data allow us to classify the four Hox‐4 genes located in the most upstream (5′) position in the HOX‐4 complex as members of a large family of homeogenes related to the Drosophila homeotic gene Abdominal B (AbdB). It therefore appears that these vertebrate genes are derived from a selective amplification of an ancestral gene which gave rise, during evolution, to the most posterior of the insect homeotic genes so far described. In agreement with the structural colinearity, these genes have very posteriorly restricted expression profiles. In addition, their developmental expression is temporally regulated according to a cranio‐caudal sequence which parallels the physical ordering of these genes along the chromosome. We discuss the phylogenetic alternative in the evolution of genetic complexity by amplifying either genes or regulatory sequences, as exemplified by this system in the mouse and Drosophila. Furthermore, the possible role of ‘temporal colinearity’ in the ontogeny of all coelomic (metamerized) metazoans showing a temporal anteroposterior morphogenetic progression is addressed.

Topology of mammalian developmental enhancers and their regulatory landscapes

How a complex animal can arise from a fertilized egg is one of the oldest and most fascinating questions of biology, the answer to which is encoded in the genome. Body shape and organ development, and their integration into a functional organism all depend on the precise expression of genes in space and time. The orchestration of transcription relies mostly on surrounding control sequences such as enhancers, millions of which form complex regulatory landscapes in the non-coding genome. Recent research shows that high-order chromosome structures make an important contribution to enhancer functionality by triggering their physical interactions with target genes.

The Dynamic Architecture of Hox Gene Clusters

Sequential activation of Hox genes correlates with a transition of negative to positive three-dimensional chromosome structure. The spatial and temporal control of Hox gene transcription is essential for patterning the vertebrate body axis. Although this process involves changes in histone posttranslational modifications, the existence of particular three-dimensional (3D) architectures remained to be assessed in vivo. Using high-resolution chromatin conformation capture methodology, we examined the spatial configuration of Hox clusters in embryonic mouse tissues where different Hox genes are active. When the cluster is transcriptionally inactive, Hox genes associate into a single 3D structure delimited from flanking regions. Once transcription starts, Hox clusters switch to a bimodal 3D organization where newly activated genes progressively cluster into a transcriptionally active compartment. This transition in spatial configurations coincides with the dynamics of chromatin marks, which label the progression of the gene clusters from a negative to a positive transcription status. This spatial compartmentalization may be key to process the colinear activation of these compact gene clusters.