. Paraxial mesoderm development in the gastrulating embryo represents a crucial morphogenetic phase that is highly susceptible to the action of toxicants and teratogens resulting in skeletal developmental anomalies. Patterning of the paraxial mesoderm, followed by somitogenesis, us the first step in the formation of the embryonic axial skeletal structures. Somites are formed by the condensation and epithelialization of the paraxial mesoderm in a caudal-to-rostral, segmented. Upon further differentiation and maturation, somites undergo an epithelial-to-mesenchymal transformation at their ventral aspect to give rise to sclerotomal cells, which migrate medially into the perinotochordal area and resegment to produce individual prevertebrae. Thus, perturbations of this process, as a result of exposure to toxicants and teratogens, will lead to developmental defects of the vertebral column, e.g. fused, supernumerary and hemi-vertebrae. Understanding the mechanism and site of action of skeletal teratogens is thus of importance to the intervention and prevention of such birth defects. We have identified, cloned, and characterized two genes, Paraxis and Pax-1, and demonstrated their functional importance in chick embryonic development, i.e. Paraxis is involved in paraxial mesoderm patterning, whereas Pax-1 is required in maintenance and differentiation of somitic structures, specifically the sclerotome. In addition, Paraxis and Pax-1 related functions/pathways are likely to be targets of teratogenic agents and conditions, including hyperthermia, valproic acid, and carbon monoxide, that affect axial skeletal development. Interestingly, antisense-mediated perturbations of Paraxis and Pax-1 expression produce somitic anomalies similar to those seen with teratogen treatment. Our overall hypothesis is that patterning and cellular differentiation events of embryonic paraxial mesoderm development are crucially dependent on the expression and activities of paraxis and pax-1, and their perturbations are an underlying cause of somitic and axial skeletal anomalies. To test this hypothesis, we will identify the downstream cellular mechanisms and the upstream gene expression regulatory requirements of Paraxis and Pax-1 during normal paraxial mesoderm development, to develop a mechanistic framework for analyzing the targets of axial skeletal teratogens.
The specific aims are: 1) to analyze the importance of cell-cell/cell-matrix interactions as potential downstream functional targets of Paraxis and Pax-1 action, 2) to correlate Paraxis/Pax-1 development; 3) to identify the upstream cellular and molecular events that regulate Paraxis and Pax-1 expression; and 4) to elucidate the specific mechanistic steps in the Paraxis/Pax-1 expression and activity cascade affected by known axial skeletal teratogens.
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