Faculty Bibliography

The membranous bones of the mammalian skull vault arise from discrete condensations of neural crest- and mesodermally-derived cells. Recently, a number of homeodomain transcription factors have been identified as critical regulators of this process. Here, we show that the homeoprotein engrailed 1 (EN1) is expressed during embryonic and perinatal craniofacial bone development, where it localizes to the skeletogenic mesenchyme, and, subsequently, to calvarial osteoblasts and osteoprogenitors. Mice lacking En1 exhibit generalized calvarial bone hypoplasia and persistent widening of the sutural joints. A reduction in calvarial membranous bone deposition and mineralization (osteopenia) is coupled to enhanced osteolytic resorption in En1 mutants. Consistent with these observations, expression of established osteoblast differentiation markers reveals that En1 function is required for both early and late phases of calvarial osteogenesis. Further analysis shows that EN1 regulates FGF signaling in calvarial osteoblasts. Moreover, EN1 indirectly influences calvarial osteoclast recruitment and bone resorption by regulating the expression of receptor activator of NFkappaB ligand (RANKL) in osteoblasts. Thus, during intramembranous bone formation, EN1 acts both cell autonomously and non-cell autonomously. In summary, this study identifies EN1 as a novel modulator of calvarial osteoblast differentiation and proliferation, processes that must be exquisitely balanced to ensure proper skull vault formation

The apical ectodermal ridge (AER) is a critical signaling center at the tip of the limb that promotes outgrowth. In mouse, formation of the AER involves a gradual restriction of AER gene expression from a broad ventral preAER domain to the tip of the limb, as well as progressive thickening of cells to form a multilayered epithelium. The AER is visible from embryonic day 10.5 to 13.5 (E10.5-E13.5) in the mouse forelimb. Previous short-term fate mapping studies indicated that, once a cell is incorporated into the AER, its descendents remain within the AER. In addition, some preAER cells appear to become incorporated into the ventral ectoderm. In the present study, we used an inducible CreER/loxP fate mapping approach in mouse to examine the long-term contribution of preAER cells to limb ventral ectoderm, as well as the ultimate fate of the mature AER cells. We used a CreER transgene that contains Msx2 regulatory sequences specific to the developing AER, and demonstrate by marking preAER cells that, at stage 2 of mouse limb bud development, the majority of the ventral ectoderm that protrudes from the body wall later covers only the paw. Furthermore, when Msx2-CreER-expressing preAER cells are marked after the onset of preAER gene expression, a similar domain of paw ventral ectoderm is marked at E16.5, in addition to the AER. Strikingly, mapping the long-term fate of cells that form the mature AER showed that, although this structure is indeed a distinct compartment, AER-derived cells are gradually lost after E12.5 and no cells remain by birth. A distinct dorsal/ventral border nevertheless is maintained in the ectoderm of the paw, with the distal-most border being located at the edge of the nail bed. These studies have uncovered new aspects of the cellular mechanisms involved in AER formation and in partitioning the ventral ectoderm in mouse limb