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Stephen H. Devoto; Scott G. Holmes; Ann C. Burke


Molecular Biology and Biochemistry


Vertebrates derive their name from a distinguishing feature of their clade; the backbone, spine, or vertebral column. The repetitive vertebrae of the spine, which support the animal through adulthood, are the most apparent reflection of the segmental pattern of the vertebrate body plan and its development. The first visible metameric segments in the vertebrate trunk, however, are the somites. The transient rows of somites, embryonic structures key to body plan organization, are an earlier instance of segmented patterning observed adjacent to the notochord along the length of the developing animal. The vertebral column and the somite rows each contain identical units equally spaced and repeated invariantly in number in individuals of the same species. Although the progression of both of these developmental processes is paramount, the relationship between them has not been resolved. Namely, the source of information that regularly positions the vertebral bodies is not clear.

Much of the research on mechanisms of metameric patterning of the vertebral column has been performed using avian and mammalian models, in which the somites are accepted as the source of both the tissue and the patterning instructions in vertebrae development. Recent research has proposed that the teleost lineage independently evolved a segmented pattern intrinsic to the notochord, which drives vertebral column patterning. Models allow for the possibility of myotomal contribution to regular centra patterning in zebrafish, but the factors involved in myotomal influence on vertebral column segmentation are not yet understood.

This thesis explores the relationship between these two instances of segmented spatial patterning. Transcriptional regulators Tbx6, Mesp-b, and Ripply1 function in a feedback network essential for somite boundary formation. This study aims to elucidate how genes involved in this network influence vertebrae periodicity. In the zebrafish model, tbx6 homozygous mutants lack somite segmentation without an effect to the notochord. In this case, vertebral column segmentation occurs but uniformity in centra length is reduced. We show that genetic perturbations of the Tbx6-Mesp-b-Ripply1 regulatory network cause irregular vertebral patterning. We also show that targeted somite boundary disruption through heat-shock treatment reduces centra periodicity locally.

Additionally, we explore the role a subset of muscle fibers plays in communicating early established segmentation to the process of vertebrae development. Signals from the notochord induce somite cells to become slow muscle precursor cells. Of these, slow muscle pioneer cells at the dorsoventral midline maintain contact with the notochord through elongation. We show that preventing the development of muscle pioneer cells causes a significant increase in centra variability. This is an investigation of muscle fiber organization as an intermediary framework for the subsequent patterning of the vertebral column and a possible site of myotomal-notochordal interface. I endeavor to clarify the requirement for synthesis of myotomal and notochordal contributions in generating the characteristic periodicity of the zebrafish vertebral column.



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