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Mechanics and Genetics of Embryonic and Tumoral Development

Group leader : Emmanuel Farge

Read the scientific activity report. (pdf 438Ko, last update 26th, march 2010) 

Embryogenesis involves two main types of morphogenetic process: genetic patterning of the body plan and mechanical movements that create the physical shape of the embryo (movie 1). We know that morphogenetic movements are controlled by expression of patterned developmental genes but, conversely, might the expression of some patterning genes be modulated by mechanical forces in the developing embryo? (Fig. 1)

Fig 1: Mechanical induction of twist gene expression. Expression of the twist gene reporter complex, Twi-lacZ (green), normally restricted to the ventral side of an unconstrained Drosophila embryo (left), is induced all around the embryo when it is constrained by a uni-axial global deformation (right).Fig 1: Mechanical induction of twist gene expression. Expression of the twist gene reporter complex, Twi-lacZ (green), normally restricted to the ventral side of an unconstrained Drosophila embryo (left), is induced all around the embryo when it is constrained by a uni-axial global deformation (right).

We are investigating whether the morphogenetic movements of Drosophila embryos influence expression of genes that control their development. We have discovered that, during gastrulation, the process of convergent extension - in which layers of cells intercalate (converge) and become longer (extend) - compresses the future anterior gut cells of the embryo and so induces expression of twist in these cells, which is necessary for proper formation of the anterior gut (Fig. 2).

fig2: Mechanical compression induces Twist expression in Drosophila embryos. (A) In stage 6 Drosophila embryos, stomodeal cells (red arrows) are not compressed; they are compressed at stage 7 by the anterior convergence-extension movement of ventral and, possibly, dorsal tissues (yellow arrows). (B) Twist is expressed weakly in the stomodeal cells early in stage 6 (red arrows); 10 mins after compression begins in stage 7, Twist starts to be expressed strongly. (C) Mutants in the genes bcd, nos and tsl, which do not undergo convergence-extension and so do not compress stomodeal cells, do not induce strong expression of Twist in these cells. (D) Strong expression of Twist in the stomodeal cells of mutant embryos is induced by mechanical compression of the cells by applying a 50µm point.fig2: Mechanical compression induces Twist expression in Drosophila embryos. (A) In stage 6 Drosophila embryos, stomodeal cells (red arrows) are not compressed; they are compressed at stage 7 by the anterior convergence-extension movement of ventral and, possibly, dorsal tissues (yellow arrows). (B) Twist is expressed weakly in the stomodeal cells early in stage 6 (red arrows); 10 mins after compression begins in stage 7, Twist starts to be expressed strongly. (C) Mutants in the genes bcd, nos and tsl, which do not undergo convergence-extension and so do not compress stomodeal cells, do not induce strong expression of Twist in these cells. (D) Strong expression of Twist in the stomodeal cells of mutant embryos is induced by mechanical compression of the cells by applying a 50µm point.

We have investigated the physiological function of twist mechanical induction in controlling expression of genes that govern anterior gut differentiation in living embryos, as well as the mechano-transduction mechanism involved in this specific case. We also are investigating whether mechanical forces regulate developmental genes in the embryos of other species and whether they regulate homeostatic genes in adult organs.

Specifically, we have observed the mechanical activation of the expression of twist-1 and c-myc genes that initiate the programm of tumoral progression in colon cancer, in genetcially predisposed APC+/-pre-tumoral mice tisues (Fig. 3)

fig3: Hydrodynamic simulation of drosophila embryo mesoderm invagination. a At time zero, the acto-myosin apical tension is increased at apical membranes (external membranes) in ventral mesoderm cells (the 7 upper cells in the simulation), in response to the increase of Myosin-II apical concentration regulated by the transduction pathway involving Dorsal, Twist and Snail. b The mechanical and hydrodynamics response of the all embryo is mesoderm invagination. The simulation describes individual cells, with membranes and junctions as sub-cellular mechanical structures. Subcellulalr movements of apical membrane flattening, apical movements of adherens junctions, apical constriction, invaginating cells elongation and subsequent shortering, experimentally observed, are predicted by the simulation. As well as the formation of a ventro-dorsal thickness gradient. Such multi-scale simulation suggests that mesoderm invagination, with the associated cascade of sub-cellular movements experimentally observed, are the consequences of a unique genetically controlled event: the increase of ventral cells apical tension.fig3: Hydrodynamic simulation of drosophila embryo mesoderm invagination. a At time zero, the acto-myosin apical tension is increased at apical membranes (external membranes) in ventral mesoderm cells (the 7 upper cells in the simulation), in response to the increase of Myosin-II apical concentration regulated by the transduction pathway involving Dorsal, Twist and Snail. b The mechanical and hydrodynamics response of the all embryo is mesoderm invagination. The simulation describes individual cells, with membranes and junctions as sub-cellular mechanical structures. Subcellulalr movements of apical membrane flattening, apical movements of adherens junctions, apical constriction, invaginating cells elongation and subsequent shortering, experimentally observed, are predicted by the simulation. As well as the formation of a ventro-dorsal thickness gradient. Such multi-scale simulation suggests that mesoderm invagination, with the associated cascade of sub-cellular movements experimentally observed, are the consequences of a unique genetically controlled event: the increase of ventral cells apical tension.

Initially, we used several complementary approaches, including cell biology, which led us to propose a mechanism for mechano-transduction in which membrane tension modulates the endocytosis of signalling proteins, causing strong modulation of downstream gene expression (Fig. 4).

fig4: snail mutant embryos (left) lack the apical redistribution of Myo-II that generates mesoderm invagination in the wild type. Both Myo-II apical redistribution and mesoderm invagination can be rescued by a soft indent in the mesoderm (right), in a Fog dependent mechano-transduction process.fig4: snail mutant embryos (left) lack the apical redistribution of Myo-II that generates mesoderm invagination in the wild type. Both Myo-II apical redistribution and mesoderm invagination can be rescued by a soft indent in the mesoderm (right), in a Fog dependent mechano-transduction process.

We are working on mechanical signaling in the activation of Myosine-II dependent mesoderm invagination in early Drosophila embryos, and its dependence of Fog endocytosis mechanical inhibition (Fig. 5).


fig 5: Uni-axial deformation quantitatively mimicking intestinal pressure induces c-Myc (cell division) and Twist (cell invasivity) expression in APC+/- heterozygous pre-tumourous tissue (i.e in tissues genetically altered, but without tumour formation).fig 5: Uni-axial deformation quantitatively mimicking intestinal pressure induces c-Myc (cell division) and Twist (cell invasivity) expression in APC+/- heterozygous pre-tumourous tissue (i.e in tissues genetically altered, but without tumour formation).

We are now studying other mechanisms by which mechanical cues from gastrulation might activate master genes protein product that control active multi-cellular morphogenetic movements and the formation of primitive organs. Our goal is to investigate if, and how, the macroscopic mechanics of a tissue contribute to the regulation of genes involved at the microscopic level in the morphogenesis of the tissue. In parallel, we develop numerical simulations of the Drosophila embryo gastrulation, to characterize in a quantitative way the biomechanical parameters of morphogenetic movements (Fig. 6 and movie 2).

fig 6: Enhancement of JunB expression in response to membrane tension inhibits BMP2 endocytosis. The images show immunofluorescence microscopy of BMP2 (a and b) and JunB (c and d) in a mouse myoblast cell line, C2C12. (a) After 10 mins of endocytosis, the signalling protein BMP2 has accumulated inside the cell. (b) Increased membrane tension induced by osmotic shock blocks BMP2 endocytosis and, after 10 mins, the protein remains on the cell surface. (c) BMP2-dependent expression of JunB in cell nuclei 30 mins after incubation of the cells with BMP2. (d) Increased expression of JunB in cell nuclei 30 mins after inhibiting endocytosis by osmotic shock and incubating the cells with BMP2. Blocking endocytosis prevents the internalisation and degradation of BMP2 and thus increases signalling by the BMP2-bound receptor on the cell surface, so stimulating JunB expression.fig 6: Enhancement of JunB expression in response to membrane tension inhibits BMP2 endocytosis. The images show immunofluorescence microscopy of BMP2 (a and b) and JunB (c and d) in a mouse myoblast cell line, C2C12. (a) After 10 mins of endocytosis, the signalling protein BMP2 has accumulated inside the cell. (b) Increased membrane tension induced by osmotic shock blocks BMP2 endocytosis and, after 10 mins, the protein remains on the cell surface. (c) BMP2-dependent expression of JunB in cell nuclei 30 mins after incubation of the cells with BMP2. (d) Increased expression of JunB in cell nuclei 30 mins after inhibiting endocytosis by osmotic shock and incubating the cells with BMP2. Blocking endocytosis prevents the internalisation and degradation of BMP2 and thus increases signalling by the BMP2-bound receptor on the cell surface, so stimulating JunB expression.