Defense Date

5-5-2018

Graduation Date

Summer 8-11-2018

Availability

One-year Embargo

Submission Type

dissertation

Degree Name

PhD

Department

Chemistry and Biochemistry

Committee Chair

Mihaela-Rita Mihailescu

Committee Member

Michael Cascio

Committee Member

Ellen Gawalt

Committee Member

Yue Feng

Committee Member

Philip Reeder

Keywords

DNA, RNA, microRNA, FMRP, G-quadruplex, gene regulation

Abstract

This study investigates protein nucleic acid interactions, focusing on G-quadruplex (GQ) forming DNA/RNA in human disease. GQ structures are formed in DNA/RNA, when four guanine residues form planar tetrads stabilized by Hoogsteen base pairing, that stack forming a GQ structure stabilized by potassium ions. These GQ structures are targeted by the arginine glycine-glycine (RGG) RNA-binding domain. Fragile X mental retardation protein (FMRP), a translation regulator protein implicated in the fragile X syndrome, has an RGG domain and has been previously shown to interact with neuronal GQ forming messenger RNA (mRNA). We have investigated three neuronal FMRP mRNA targets that we predicted form GQ structures and analyzed their interactions with FMRP. One mRNA target was NR2B, an important protein in the synaptic density. Our work shows that v a GQ forms in the 3’-UTR of NR2B and that this structure is sufficient for the NR2B mRNA recognition by FMRP. The second neuronal FMRP target we investigated was PSD-95, which is post synaptic density protein 95, a protein important to healthy synapse functioning. We showed that two GQ structures are forming in the PSD-95 3’-UTR, encompassing the binding site of the miRNA 125a, a non-coding microRNA that regulates the translation of PSD-95 mRNA. Moreover, depending on its phosphorylation status, FMRP acts as a switch, mediating the binding of miRNA-125a and modulating the activation or deactivation of the PSD-95 translation. Further investigation of noncanonical roles for GQ in translation regulation led us to investigate a GQ structure in close proximity to an alternative polyadenylation site in the 3’-UTR of the brain derived neurotrophic factor (BDNF) gene. Our results suggest that the GQ structure works to preferentially choose one poly(A) over another, ultimately affecting the translation of BDNF. Lastly, we investigated GQ formation in pre-miRNA-149 and how the GQ leads to a disruption of the canonical long extended hairpin and eventually maturation of the microRNA miR-149, which will lead to a disruption of mRNA’s translational regulation. These four projects demonstrate unique roles for the GQ structure, contributing to our understanding of GQ role in translational regulation.

Language

English

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