Migration of Founder Epithelial Cells Drives Proper Molar Tooth Positioning and Morphogenesis

Abstract

Summary The proper positioning of organs during development is essential, yet little is known about the regulation of this process in mammals. Using murine tooth development as a model, we have found that cell migration plays a central role in positioning of the organ primordium. By combining lineage tracing, genetic cell ablation, and confocal live imaging, we identified a migratory population of Fgf8-expressing epithelial cells in the embryonic mandible. These Fgf8-expressing progenitors furnish the epithelial cells required for tooth development, and the progenitor population migrates toward a Shh-expressing region in the mandible, where the tooth placode will initiate. Inhibition of Fgf and Shh signaling disrupted the oriented migration of cells, leading to a failure of tooth development. These results demonstrate the importance of intraepithelial cell migration in proper positioning of an initiating organ.

Epithelial cell behaviours during neurosensory organ formation

“…Invagination of the molar placode, by contrast, relies partially on Shh signalling; inhibition of Smo receptor activity by cyclopamine at E12.5 alters the shape of basal and suprabasal cells and nuclei from elongated to round, suggesting reduced cell motility, and mice with conditional Shh deletion in the oral ectoderm show wider and shallower molar buds (Dassule et al, 2000;Li et al, 2016). In line with this, it has been shown that Shh is expressed in the dental epithelium (Bitgood and McMahon, 1995;Dassule and McMahon, 1998;Keränen et al, 1998;Prochazka et al, 2015), and the Smo and Shh targets Ptch1 and Gli1 broadly in the dental epithelium and mesenchyme (Dassule and McMahon, 1998;Hardcastle et al, 1998;Prochazka et al, 2015). Yet, as SU5402 or cyclopamine treatments have tissue-wide (epithelium and mesenchyme) effects, and given that colocalization data of Fgf and Shh signalling components at the single-cell level are not available, it is difficult to assess precisely the direct target cells of Fgf and Shh signalling during molar tooth placodal thickening and invagination.…”

“…However, it has been shown that the conditional elimination of Fgf8-expressing epithelial cells (by diptheria toxin A) results in initiation of the molar placode but not further growth of the tooth bud (Prochazka et al, 2015), suggesting that Fgf8-expressing epithelial cells are required for the latter process. In addition, clonal analyses and in vivo imaging using a confetti multicolour reporter under the control of Fgf8 ires-cre , have shown that Fgf8-expressing cells and their descendants are dispersed in a mosaic manner in the E14.5 molar tooth, rather than retaining adjacent positions in clonal patches, suggesting a complex mode of displacement (Prochazka et al, 2015).…”

“…In addition, clonal analyses and in vivo imaging using a confetti multicolour reporter under the control of Fgf8 ires-cre , have shown that Fgf8-expressing cells and their descendants are dispersed in a mosaic manner in the E14.5 molar tooth, rather than retaining adjacent positions in clonal patches, suggesting a complex mode of displacement (Prochazka et al, 2015). In vivo imaging has also shown that, at E11.5, Fgf8-expressing cells form a centripetal-oriented structure (a rosette) at the surface of the epithelium (Fig.…”

“…Stratification of the molar dental epithelium occurs through cell divisions perpendicular to the basement membrane, generating basal and suprabasal cells, and the abrogation of Fgf receptor signalling (using SU5402) and complementary activation of Fgf signalling (using Fgf10-soaked beads applied to E11.5 mandible slice cultures) has shown that Fgf signalling activity is necessary and sufficient for this epithelial stratification (Li et al, 2016). Indeed, various Fgf pathway components are expressed during molar placode formation: Fgf8 and Fgfr2 are broadly expressed in the dental epithelium (Fgfr2 isoform IIIb; Kettunen et al, 1998;Laurikkala et al, 2006;Li et al, 2014;Prochazka et al, 2015), Fgfr1IIIc in the mesenchyme Li et al, 2014), and Etv4 broadly in the dental epithelium and mesenchyme (Porntaveetus et al, 2011;Prochazka et al, 2015). Invagination of the molar placode, by contrast, relies partially on Shh signalling; inhibition of Smo receptor activity by cyclopamine at E12.5 alters the shape of basal and suprabasal cells and nuclei from elongated to round, suggesting reduced cell motility, and mice with conditional Shh deletion in the oral ectoderm show wider and shallower molar buds (Dassule et al, 2000;Li et al, 2016).…”

“…1A,B). The centre of this rosette is located at a distance from Shh-expressing epithelial cells located at the level of the molar placode (Prochazka et al, 2015), and it is difficult to judge whether all Fgf8-expressing/derived and Shhexpressing cells are two entirely different and/or distant populations. Nevertheless, the rosette of Fgf8-expressing/derived cells has been shown to migrate anteriorly towards the placode's Shh-expressing cells (Fig.…”