Date of Award

8-2007

Level of Access

Campus-Only Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biochemistry and Molecular Biology

Advisor

Douglas B. Spicer

Second Committee Member

Carol Kim

Third Committee Member

Robert Friesel

Abstract

The cranial sutures are composed of two edges of calvarial bones called osteogenic fronts (OF) and a narrow space in between, the mid-suture mesenchyme (MSM). These sutures must remain patent while the underlying brain grows. Therefore, the premature closure (craniosynostosis) of these structures results in craniofacial dysmorphism, and increased intracranial pressure. Craniosynostosis results from activating mutations of FGFR1-3 and from haploinsufficiency of Twistland although the mutations are diverse the resulting phenotype is the premature fusion of sutures. Twisti is a member of the basic Helix-Loop-Helix family of transcription factors which forms both homodimers with another Twisti protein (T/T) or heterodimers with E protein (T/E). Saethre-Chotzen is the syndrome associated with haploinsufficiency of Twisti which presents with an extension of T/T homodimer toward the mid-suture mesenchyme resulting in craniosynostosis. Therefore in order to recover the phenotype of premature fusion we shifted the dimer balance to form more T/E relative to T/T. We took three approaches: First, we modified the relative concentrations of two other basic Helix-Loop-Helix transcription factors which regulate Twistl dimer formation: E12 and Id protein. Therefore, by increasing E12 and decreasing Id we favored T/E production in Twistl+/- mice which resulted in patent sutures. Second: We inhibited FGF signaling regulated by T/T and over-expressed the gene up-regulated by T/E crossing Twistl+/- mice with Spryl overexpressing mice which recovered the phenotype. Third: We over-expressed tethered TT and TE dimers to characterize the behavior of the sutures after FGF2 stimulation. We observed that only TT overexpressing calvarias resembled those of Twistl+/-. In conclusion, changes in the relative levels of Twistl dimers T/T and T/E result in differentially expressed genes, which regulate the behavior of the sutures in such a way that it can prevent craniosynostosis in Twistl+/- mice.

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