We are interested in the regulatory mechanisms underlying vertebrate pattern formation, in both developmental and evolutionary contexts. For the past years, we have focused on Hox genes, a family of transcription factors that display special paradigmatic values, regarding their regulatory strategies, their functional organization and their key roles in morphological evolution. Over the past 20 years, we implemented a large program aimed at genetically dissecting these various aspects of Hox gene biology, either using the potential of established mouse genetic manipulations or by designing and implementing powerful strategies relying upon in vivo chromosome engineering via LoxP based recombinations to generate either subtle or large genomic rearrangements (TAMERE, STRING and PANTHERE). This large allelic series is currently used to analyse the regulation of this gene family with the help of molecular and biochemical tools.
Short- and long-range regulation of Hox gene expression ; the colinearity enigma
We pursue our efforts to understand the mechanistic basis of colinearity, i.e. the mechanism whereby neighboring Hox genes are activated one after the other in overlapping anterior to posterior domains, in the developing trunk axis. The nature of this process at work during gastrulation is still poorly understood due to the scarcity of material available for molecular studies at this stage. A distinct version of colinearity was observed during the development of the appendicular axes (the arms and legs) and, in this case, our systematic approach has recently started to provide a conceptual framework to account for this enigmatic phenomenon. The understanding of this process in limbs was made possible by analysing the 3D structure of the surrounding chromatin, which revealed to Topologically Associating Domains (TADs) on either sides on the gene cluster. These two TADs implement distinct regulations and are globally activated in a time sequence that leads to a colinear transcriptional output. We currently try to isolate the factors activating these TADs at the proper time and in the proper cells during limb development.
We also plan to characterize in some details the various global regulatory controls which were shown to direct expression of groups of Hox genes (shared enhancers) in a variety of structures such as the digits, the external genitalia, the intestinal hernia, the metanephric kidneys, the whisker pads or the emerging somites. These enhancer elements are of critical mechanistic and evolutionary importance and are looked for by using both a genetic approach in vivo and a large-scale transgenic program using bacterial artificial chromosomes (BACs) tagged with reporter transgenes. We are currently narrowing down such global enhancers and try and isolate those factors binding to them.
The function of Homeobox genes
In parallel, we continue our attempts to unravel the functions of Hox genes by combining our allelic series with cutting-edge molecular and biochemical approaches. In this way, various combinations of Hoxd gene gain of function and loss of function produced as a result of our TAMERE approach complemented by CRISPR-cas9 based modifications are currently being evaluated. We are particularly interested to look at the functional relationships existing between genes members of the HoxD cluster and some of the LncRNAs claimed in the literature to directly regulate their potential functions. In this context, functional approaches are targeted to the developping limbs, the uro-genital system as well as to the intestinal tractus.
Our work naturally interfaces with an evolutionnary context and we thus continue to explore the potential involvement of Hox genes in morphological evolution, at the levels of both the modification of regulatory controls and the resulting variations in the functional deployment of HOX proteins and their impacts upon the emergence of novel structures in the course of vertebrate phylogeny. We are particularly interested in the relationship between the emergence of novel enhancers and the evolution of the structures where they are active. In this context, we focus on birds and snakes, in addition to mammals.