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EXAMINING THE POST-TRANSCRIPTIONAL REGULATION OF LUNATIC FRINGE (Lfng) IN THE MOUSE SEGMENTATION CLOCK

Wahi, Kanu, Wahi
http://orcid.org/0000-0003-1114-6075
http://rave.ohiolink.edu/etdc/view?acc_num=osu1469108634

Abstract Details

2016, Doctor of Philosophy, Ohio State University, Molecular Genetics.
Somitogenesis is a developmental process in vertebrates involving periodic formation of somites that bud from an unsegmented region known as the pre-somitic mesoderm (PSM) and give rise to the axial skeleton and skeletal muscle in the developed organism. The process of somitogenesis is regulated by a "segmentation clock" that times somite formation and is evolutionarily conserved among vertebrates. Genes, such as Lunatic fringe (Lfng), are required for normal clock function in chickens, mice and humans and exhibit rapid cyclic expression in the PSM with a period that matches the rate of somite formation. To maintain rapid oscillations, especially in the posterior PSM, it is hypothesized that the Lfng transcript should be promptly degraded to ensure its clearance of from cells before the next round of oscillation begins. The 3’UTR of Lfng contains number of conserved sequences that could regulate Lfng expression at the post transcriptional level. In this study we explore the post-transcriptional regulation of the Lfng transcript by the Lfng 3’UTR in the context of the segmentation clock. The Lfng 3'UTR has binding sites for microRNAs (miRNAs), such as miR-125a and miR-200b that are known to affect transcript stability via the 3’UTR. We tested the effect of mutating the mir-125a binding sites in the mouse Lfng 3’UTR on mRNA stability of Venus reporter constructs in transgenic mice and observed constitutive Venus expression in the posterior PSM, as opposed to the oscillatory Venus expression when the miR-125a binding sites are not mutated. This implies that the miR-125a binding sites in the Lfng 3'UTR influence RNA turnover in the posterior region of the mouse PSM. Interestingly, when we generated mutant mouse lines that do not express miR-125a we found no effect on somitogenesis and skeletal formation. This indicates that either a compensatory mechanism may be in effect or that miR-125a does not play a role in the mouse segmentation clock. We also examined if another highly conserved region of the Lfng 3'UTR, that is 120 bp in length and has a miR-200 binding site within it, can sufficiently destabilize the transcript. We examined this in two contexts. We found that blocking the miR-200b binding site in the Lfng 3’UTR in chicken embryos has no effect on somitogenesis. Similarly, in mouse myoblast cells the 120 bp sequence of the mouse Lfng 3'UTR is not sufficient to destabilize the transcript. Our results indicate that the 120 bp sequence and the miR-200b binding site within it, are not sufficient to destabilize the transcript and it is likely that this regulation operates in a tissue specific manner. Future work in the lab will examine the function of the endogenous miR-125a binding site in the Lfng 3'UTR on mouse segmentation and skeletal formation. Understanding the post transcriptional regulation of clock genes like Lfng would shed light on mechanisms that maintain oscillatory gene expression in the PSM and ensure timely somite formation.
Susan Cole (Advisor)
Robin Wharton (Committee Member)
Keith Slotkin (Committee Member)
Dawn Chandler (Committee Member)
171 p.
Developmental Biology; Genetics; Molecular Biology

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Citations

  • Wahi, Wahi, K. (2016). EXAMINING THE POST-TRANSCRIPTIONAL REGULATION OF LUNATIC FRINGE (Lfng) IN THE MOUSE SEGMENTATION CLOCK [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1469108634

    APA Style (7th edition)

  • Wahi, Wahi, Kanu. EXAMINING THE POST-TRANSCRIPTIONAL REGULATION OF LUNATIC FRINGE (Lfng) IN THE MOUSE SEGMENTATION CLOCK. 2016. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1469108634.

    MLA Style (8th edition)

  • Wahi, Wahi, Kanu. "EXAMINING THE POST-TRANSCRIPTIONAL REGULATION OF LUNATIC FRINGE (Lfng) IN THE MOUSE SEGMENTATION CLOCK." Doctoral dissertation, Ohio State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=osu1469108634

    Chicago Manual of Style (17th edition)

osu1469108634
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© 2016, some rights reserved.Creative Commons License EXAMINING THE POST-TRANSCRIPTIONAL REGULATION OF LUNATIC FRINGE (Lfng) IN THE MOUSE SEGMENTATION CLOCK by Kanu Wahi Wahi is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License. Based on a work at etd.ohiolink.edu.
This open access ETD is published by The Ohio State University and OhioLINK.