Much of the basic information about individual organ development comes from studies using model species. Whereas conservation of gene regulatory networks across higher taxa supports generalizations made from a limited number of species, generality of mechanistic inferences remains to be tested in tissue culture systems. Here, using mammalian tooth explants cultured in isolation, we investigate self-regulation of patterning by comparing developing molars of the mouse, the model species of mammalian research, and the bank vole. A distinct patterning difference between the vole and the mouse molars is the alternate cusp offset present in the vole. Analyses of both species using 3D reconstructions of developing molars and jaws, computational modeling of cusp patterning, and tooth explants cultured with small braces show that correct cusp offset requires constraints on the lateral expansion of the developing tooth. Vole molars cultured without the braces lose their cusp offset, and mouse molars cultured with the braces develop a cusp offset. Our results suggest that cusp offset, which changes frequently in mammalian evolution, is more dependent on the 3D support of the developing jaw than other aspects of tooth shape. This jaw-tooth integration of a specific aspect of the tooth phenotype indicates that organs may outsource specific aspects of their morphology to be regulated by adjacent body parts or organs. Comparative studies of morphologically different species are needed to infer the principles of organogenesis.T he conservation of gene regulatory networks in the development of homologous organs is well exemplified by dentitions. Teeth in fish, reptiles, and mammals have been found to use largely shared developmental gene networks (1, 2). Furthermore, tooth development, similar to organ development in general, results from sequential events that appear to be relatively independent from the development of other organs. In particular, mammalian teeth have been considered highly selfregulatory, as they form inside the dental follicle buffered from the surrounding environment (3-5). The self-regulatory aspect of teeth is further supported by the reiterative use of the same signaling pathways and signaling centers, called enamel knots, present within each growing tooth (2). Experiments on growing teeth have revealed how an activator-inhibitor balance of signaling molecules within a developing tooth and between adjacent teeth regulate dental patterning (6-12).Despite the evidence from mice indicating self-regulation of teeth, several observations suggest an intriguing link between jaw bone and tooth crown formation (3,13,14). In humans, abnormal crown patterns in patients with congenital syphilis have been ascribed to result from infection of the tooth follicle (3). Likewise, loss of function of parathyroid hormone-related protein (PTHrP) leads to loss of alveolar bone resorption and deformation of the tooth crown in the mouse (15)(16)(17)(18)(19). A lethal human syndrome called Blomstrand chondrodysplasia, which invol...