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Influence and regulation of PCBP2 and YTHDF2 RNA-binding proteins during self-renewal and differentiation of human induced pluripotent stem cells

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dc.contributor.authorHeck, Adam M., author
dc.contributor.authorWilusz, Carol J., advisor
dc.contributor.authorWilusz, Jeffrey, advisor
dc.contributor.authorOsborne Nishimura, Erin, committee member
dc.contributor.authorMontgomery, Tai, committee member
dc.contributor.authorZhou, Wen, committee member
dc.date.accessioned2020-01-13T16:42:00Z
dc.date.available2021-01-07T16:41:53Z
dc.date.issued2019
dc.description.abstractEmbryonic stem cells (ESCs) are able to self-renew or differentiate into any cell type in the body, a property known as pluripotency that enables them to initiate early growth and development. However, the ethical implications of harvesting and manipulating ESCs hinders their use in basic research and the clinical applications. Thus, the discovery that somatic cells can be exogenously reprogrammed into induced pluripotent stem cells (iPSCs) offers new and exciting possibilities for gene therapy, personalized medicine and basic research. However, more research is needed into the mechanisms involved in regulating pluripotency in order for iPSCs to reach their full potential in the research lab and clinic. To maintain a state of self-renewal, yet also be able to rapidly differentiate in response to external signals, pluripotent stem cells need to exert tight control over gene expression through transcriptional and post-transcriptional mechanisms. There are several notable transcriptional networks that regulate pluripotency, but the post-transcriptional mechanisms remain poorly characterized. mRNA decay is one form of post-transcriptional regulation that can help to both maintain the steady-state of a transcriptome or facilitate its rapid remodeling. To this end, degradation rates are influenced by the elements contained in an mRNA and the RNA-binding proteins (RBPs) they associate with. Previous reports have indicated the RNA modification N6-methyladenosine (m6A) and C-rich sequence elements (CREs) can affect mRNA decay in pluripotent stem cells. Therefore, we sought to further understand the roles of m6A and CREs in mRNA decay in stem cells by characterizing the expression and mRNA targets of two RBPs that recognize these elements, YTHDF2 and PCBP2, respectively. In this thesis, I report YTHDF2 is differentially regulated in pluripotent and differentiated cells and that YTHDF2 contributes to pluripotency by targeting a group of mRNAs encoding factors important for neural development. The down-regulation of YTHDF2 during neural differentiation is consistent with increased expression of neural factors during this time. Moreover, YTHDF2 expression is regulated at the level of translation via elements located in the first 300 nucleotides of the 3' untranslated region of YTHDF2 mRNA. Based on these results, I propose that stem cells are primed for rapid differentiation by transcribing low levels of mRNAs encoding neural factors that are subsequently targeted for degradation, in part by YTHDF2, until differentiation is induced. On the other hand, PCBP2 is up-regulated upon differentiation of pluripotent stem cells and regulates several mRNAs associated with pluripotency and development, including LIN28B. Notably, expression of long non-coding RNAs (lncRNAs) that contain human endogenous retrovirus element H (HERV-H) is influenced by PCBP2. HERV-H lncRNAs are almost exclusively expressed in stem cells and play a role in maintaining a pluripotent state, although their functions are not fully understood. Intriguingly, some HERV-lncRNAs can also regulate PCBP2 expression, as altering the expression of LINC01356 or LINC00458 effects PCBP2 protein levels. Based on these results, I propose the reciprocal regulation of PCBP2 and HERV-H lncRNAs influences whether stem cells maintain a state of self-renewal or differentiate. Taken together, these findings demonstrate that YTHDF2 and PCBP2 post-transcriptionally regulate gene expression in stem cells and influence pluripotency.
dc.format.mediumborn digital
dc.format.mediumdoctoral dissertations
dc.identifierHeck_colostate_0053A_15801.pdf
dc.identifier.urihttps://hdl.handle.net/10217/199825
dc.languageEnglish
dc.language.isoeng
dc.publisherColorado State University. Libraries
dc.relation.ispartof2000-2019
dc.rightsCopyright and other restrictions may apply. User is responsible for compliance with all applicable laws. For information about copyright law, please see https://libguides.colostate.edu/copyright.
dc.subjectinduced pluripotent stem cells
dc.subjectRNA binding protein
dc.subjectRNA modification
dc.subjectpost-transcriptional regulation
dc.subjectC-rich element
dc.subjectRNA decay
dc.titleInfluence and regulation of PCBP2 and YTHDF2 RNA-binding proteins during self-renewal and differentiation of human induced pluripotent stem cells
dc.typeText
dcterms.embargo.expires2021-01-07
dcterms.embargo.terms2021-01-07
dcterms.rights.dplaThis Item is protected by copyright and/or related rights (https://rightsstatements.org/vocab/InC/1.0/). You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s).
thesis.degree.disciplineCell and Molecular Biology
thesis.degree.grantorColorado State University
thesis.degree.levelDoctoral
thesis.degree.nameDoctor of Philosophy (Ph.D.)

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