Dissertation
MicroRNA mediated mechanisms of stem cell specification
University of Iowa
Doctor of Philosophy (PhD), University of Iowa
Spring 2020
DOI: 10.17077/etd.005400
Abstract
The role of extracellular signaling pathways and transcription factor cascades have been well studied during organogenesis but microRNAs (miRs) remain poorly investigated, due to their complex genomic arrangement and redundancy. In this dissertation, I begin to fill this gap in knowledge with my examination of the miR-200 family, which was inhibited using a Plasmid based miR Inhibitor System (PMIS) in developing murine embryos. We found that the miR-200 family directly regulated the differentiation of Dental Epithelial Stem Cells (DESCs), which failed to exit the cell cycle when the miR-200 family was inhibited. Two transcription factors, Sox2 and Pitx2, have been shown by our lab and others to both be required for tooth development, and to be direct targets of miR-200c (Sox2) or miR-200a(Pitx2). Upon inhibition of the miR-200 family, these factors were ectopically expressed in expanded regions of the lower incisor. We show that the miR-200c functional group regulates the level of autophagy which occurs at low levels normally in the lower incisor. Inhibition of miR-200c resulted in an increase in autophagy accompanied by epithelial cell death, in a caspase independent manner.
The embryonic lethality of the PMIS-miR-200 double inhibitor embryos suggested that the miR-200 family functions in the development of other organs as well. We examined heart and lung development and found defects in progenitor cell differentiation, and an increase in proliferation, in PMIS-miR inhibitor embryos. We noted defects in the expression of proteins required for contractility in the heart, and hyperproliferation of cells in the lung. We found that increased levels of Pitx2 in the heart promoted the expression of other transcription factors, including Gata4, which promoted cardiomyocyte proliferation and prevented differentiation. In the lung, we found a miR-200c-Irx1-Sox9 regulatory loop that promotes the differentiation of distal airway epithelial cells.
Details
- Title: Subtitle
- MicroRNA mediated mechanisms of stem cell specification
- Creators
- Mason E. Sweat
- Contributors
- Brad A Amendt (Advisor)Huojun Cao (Committee Member)John F Engelhardt (Committee Member)C. Andrew Frank (Committee Member)Liu Hong (Committee Member)
- Resource Type
- Dissertation
- Degree Awarded
- Doctor of Philosophy (PhD), University of Iowa
- Degree in
- Anatomy and Cell Biology
- Date degree season
- Spring 2020
- DOI
- 10.17077/etd.005400
- Publisher
- University of Iowa
- Number of pages
- xvi, 129 pages
- Copyright
- Copyright 2020 Mason E. Sweat
- Comment
- This thesis has been optimized for improved web viewing. If you require the original version, contact the University Archives at the University of Iowa: https://www.lib.uiowa.edu/sc/contact/
- Language
- English
- Description illustrations
- color illustrations
- Description bibliographic
- Includes bibliographical references (pages 125-129).
- Public Abstract (ETD)
MicroRNA are small RNA molecules that are expressed in cells to regulate gene expression. These small, 20-23 nucleotide molecules usually function to prevent the translation of mRNA molecules by the ribosome, and while they have been extensively studied in diseases including cancer and obesity, little is known about what role they play in development.
The Amendt lab has developed a system to allow for the specific inhibition of microRNA in mice, and we used this system to study the function of the miR-200 family. The miR-200 family is composed of five different microRNA that share one of two different target recognition sequences. Therefore, in order to study the function of the entire miR-200 family, two mouse lines were created to inhibit each target recognition sequence. By crossing the two separate mouse lines, we could functionally inhibit the entire miR-200 family, and investigate the role for the family in tooth, heart and lung development.
After inhibiting the miR-200 family, we found that proteins normally repressed by the family were present in developing hearts, lungs, and teeth in higher-than-normal amounts. Furthermore, these proteins, which are called transcription factors and govern cell behavior, seemed to prevent the cells from reaching maturity. Thus, we had an increase in the number of dividing stem and progenitor cells in these organs. The findings of the study might be useful for future therapeutic applications.
- Academic Unit
- Anatomy and Cell Biology; Craniofacial Anomalies Research Center
- Record Identifier
- 9983949695502771
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