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Regulation of Notch Activation by Lunatic Fringe During Somitogenesis

Williams, Dustin R

Abstract Details

2014, Doctor of Philosophy, Ohio State University, Molecular Genetics.
During somitogenesis, paired somites periodically bud from the presomitic mesoderm (PSM) located at the caudal end of the embryo. These somites will give rise to the axial skeleton and musculature of the back. The regulation of this process is complex and occurs at multiple levels. In the posterior PSM, Notch activity levels oscillate as part of a clock that controls the timing of somite formation. In the anterior PSM, the Notch pathway is involved in somite patterning. In the clock, cyclic Notch activation is dependent upon periodic repression by the glycosyltransferase Lunatic fringe (LFNG). Lfng mRNA levels cycle over a two-hour period in the clock, facilitating oscillatory Notch activity. Lfng is also expressed in the anterior PSM, where it may regulate Notch activity during somite patterning. We previously found that mice lacking overt oscillatory Lfng expression in the posterior PSM (Lfng¿FCE) exhibit abnormal anterior development but relatively normal posterior development, suggesting distinct requirements for segmentation clock activity during the formation of the anterior skeleton compared to the posterior skeleton and tail. To further test this idea, we created an allelic series that progressively lowers Lfng levels in the PSM. We find that further reduction of Lfng expression levels in the PSM does not increase disruption of anterior development. However tail development is increasingly compromised as Lfng levels are reduced, suggesting that primary body formation is more sensitive to Lfng dosage than is secondary body formation. Further, we find that low levels of oscillatory Lfng are expressed in the posterior PSM of Lfng¿FCE mutants. This reduced expression if sufficient to support relatively normal posterior development, however, we find that the period of the segmentation clock is increased when the amplitude of Lfng oscillations are low. These data support the hypothesis that there are differential requirements for oscillatory Lfng during primary and secondary body formation and that posterior development is less sensitive to overall Lfng levels as long as some oscillatory Lfng is present. For oscillatory Lfng transcription to result in the cyclic Notch activation that is required for somitogenesis, there must be mechanisms in place to rapidly eliminate LFNG protein during the short “off” phase of the clock. We hypothesized that secretion of LFNG is a mechanism for terminating its activity in the clock when it is no longer needed. To test the in vivo relevance of LFNG secretion, we generated knock-in mice expressing a Golgi-tethered LFNG variant that cannot be secreted (LfngtLFNG). Mice carrying a single copy of the LfngtLFNG allele exhibit severe skeletal abnormalities, supporting our hypothesis that tethering LFNG in the Golgi generates a dominant, hyperactive fringe protein that perturbs the segmentation clock. Somites in these mutants do not exhibit proper rostro-caudal patterning and form irregular boundaries. Expression of clock genes is perturbed, and Notch activity levels no longer oscillate. These results support our hypothesis that LFNG processing and secretion play important roles in its function in the segmentation clock, and provide further evidence that post-transcriptional regulation of the segmentation clock is critical during somitogenesis.
Susan Cole (Advisor)
Mark Seeger (Committee Member)
Amanda Simcox (Committee Member)
Heithem El-Hodiri (Committee Member)
111 p.

Recommended Citations

Citations

  • Williams, D. R. (2014). Regulation of Notch Activation by Lunatic Fringe During Somitogenesis [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1397756440

    APA Style (7th edition)

  • Williams, Dustin. Regulation of Notch Activation by Lunatic Fringe During Somitogenesis. 2014. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1397756440.

    MLA Style (8th edition)

  • Williams, Dustin. "Regulation of Notch Activation by Lunatic Fringe During Somitogenesis." Doctoral dissertation, Ohio State University, 2014. http://rave.ohiolink.edu/etdc/view?acc_num=osu1397756440

    Chicago Manual of Style (17th edition)