CIRB
Progenitor cell recruitment in adult
In recent years our understanding of stem cells, their origin and maintenance, has evolved and they are many examples of differentiated cells turning into progenitors for other cell types or tissues. A major issue is now to control the cellular milieu to promote the differentiated/progenitor transition. Epimorphic regeneration in amphibians and fishes provides powerful models of integrated regeneration. After the loss of an appendage, multipotent cells from different origins accumulate at the damaged surface to form a mass of proliferating progenitor cells: the blastema. Whereas it seems that any differentiated cell is able to participate to blastema formation, only a few of them will respond to injury by adopting a progenitor identity. The latter cells probably inform their neighbours that they should maintain their differentiated state, thus preserving tissue integrity.
Our studies mainly focus on the signals involved in the regeneration of the adult zebrafish caudal fin. Converging arguments support the idea that progenitor/stem cell recruitment and proliferation requires an interaction between the epidermis and the underlying mesenchyme. Recently we have shown that apoptosis in the epidermis plays a major role in mesenchymal progenitor cells mobilization. The next step is to study apoptosis regulation and to which signals induced by apoptosis participate in progenitor cell recruitment.
Photocontrol of protein activity
The in vivo control, in space and time, of gene expression is an important issue in integrative biology. Inducible gene expression is still highly limited by inducer delivery. Ideally, inducers should be noninvasive with a rapid, local and tunable activity. To fulfill these criteria, we developed an approach based on two-photon driven photo-activation of a non active precursor distributed in the organism. We presently use this technology to study, in the live zebrafish and at the single cell level, different developmental and regenerative processes. This project that combines biological, chemical and physical approaches, is collaborative with Pr Ludovic Jullien (ENS, Paris) and Dr David Bensimon (ENS, Paris).
We have produced transgenic fish strains to study (i) cell lineage decisions during regeneration and (ii) the dynamics of transcription factors (Engrailed, Pax6) and kinases (p38a). These proteins (fused to a steroid receptor) are activated at the single cell level by uncaging a nonendogeneous tamoxifen that binds the receptor and induces the nuclear addressing of the fusion proteins. These transgenic strains will also express a fluorescent inducible construct allowing us to identify the “activated cells”.
Photo-control of CreERT2 recombinase activity in Tg reporter line.
Tg (ef1a : loxP-GFP-loxP-dsRed2) embryos were injected with 3 pg of CreERT2 mRNA at the one cell stage and further incubated with 3mM of caged inducer (clnd). Photoactivation of clnd at the 3/6-somite stage resulted in dsRed expression observed here at 2 dpf. In epifluorescence microscopy, the non-UV illuminated embryos (not shown) do not express dsRed whereas they do express dsRed in few cells upon two-photon excitation in a single cell in the forming retinaor in a just formed somite. Scale bars: 100mm. (Sinha et al. Zebrafish, 2010).
SELECTED PUBLICATIONS 2006-2010
- Dufourcq P. & Vriz S. (2006), The chemokine SDF-1 regulates blastema formation during fin regeneration. Dev Genes Evol, 216, 635-9.
- Dufourcq P., Roussigné M., Blader P., Rosa F., Peyrieras N. & Vriz S. (2006), Mechano-sensory organ regeneration in adult : the zebrafish lateral line as a model. Mol Cell Neurosci, 33, 180-7.
- Neveu P., Aujard I., Benbrahim C., Le Saux T., Allemand J.F., Vriz S., Bensimon D. & Julien L. (2008), A caged retinoic acid for use with one and two photon excitation in zebrafish embryos. Angewandte Chemie Int Ed Engl, 47, 3744-46.
- Bouzaffour M., Dufourcq P., Lecaudey V., Haas P. & Vriz S. (2009), Fgf and Sdf-1 pathways interact during zebrafish fin regeneration. PLoS ONE, 4(6): e5824.
- Rampon C., Bouzaffour M., Ostuni M., Dufourcq P., Girard C., Freyssinet J.M., Lacapere J.J., Schweizer-Groyer G. & Vriz S. (2009), Translocator protein, TSPO (18kDa) is involved in primitive erythropoiesis in zebrafish. FASEB J, 23(12) : 4181-92.
- Sinha D., Neveu P., Gagey N., Aujard I., Benbrahim-Bouzidi C., Le Saux T., Rampon C., Gauron C., Goetz B., Dubruille S., Baaden M., Volovitch M., Bensimon D., Vriz S. & Jullien L. (2010), Photocontrol of protein activity in cultured cells and zebrafish with one- and two-photon. ChemBioChem, 11(5) : 653-663.
- Sinha D., Neveu P., Gagey N., Aujard I., Le Saux T., Rampon C., Gauron C., Kawakami K., Leucht C., Bally-Cuif L., Volovitch M., Bensimon D., Jullien L. & Vriz S. (2010), Photo-activation of the CreERT2 recombinase conditional site-specific recombination with hogh spatio-temporal resolution. Zebrafish, in Press.
- Bouzzafour M., Rampon C., Ramaugé M., Courtin F. & Vriz S. (2010) Differential effect of thyroid hormones during regeneration in zebrafish. Gen Comp Endocrinol, in Press.