Diseases of the vertebrae include severe malformations such as spondylocostal dysostosis, and milder ones including scoliosis, or curvature of the spine. About 1 in 1000 live births has a congenital form of one of these diseases. The genes implicated in these diseases also affect segmentation of the embryonic mesoderm, strongly suggesting that embryonic segmentation defects underlie vertebral malformations. We propose to explore the link between embryonic segmentation and vertebral development, focusing on three genes that are at the center of the segmentation, and have not been extensively characterized for their role in spine development. One of these, mespb, is regulated by Notch signaling. The other two are tbx6 and ripply1, which together regulate mespb. We will do all of our work in zebrafish, because its advantages of external development, high fecundity, transparency, and genetic tools will allow for rapid advances in understanding normal and pathological vertebral development. We will use mutants and anti-sense morpholinos to reduce, and already established transgenic animals to increase, the dosage of each of the three genes. We will assay both embryonic segmentation and vertebral shapes, using a quantitative assay to map the location and number of defects. The quantitative approach will allow us to test for correlations between levels of genes and number of defects. We will also characterize the development of vertebrae following perturbations in embryonic segmentation, to probe the relationship between embryonic segments and adult vertebral shapes. We will use gene expression, histological observations, and live imaging to characterize the development of vertebrae and ribs in animals in which embryonic segmentation was previously characterized. Finally, we will modify gene expression and examine the expression of sclerotome genes to begin to understand the relationship between segmentation defects and vertebral defects our research will be in the areas of molecular, cell, and developmental biology of vertebral development. The research will be carried out by a Ph.D. graduate student who will receive training in these research areas. S/he will be assisted by two undergraduates at a time, who will receive one to two years of training each. The total number of participating undergraduate trainees will be six.
Malformations of the vertebrae, including both severe forms and scoliosis, have been linked to the genes that regulate segmentation in the early embryo. The proposed research examines multiple genes in this network, to identify targets for therapeutic approaches to prevent congenital spinal diseases.