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DIFFERENTIAL REGULATION OF TYROSINE HYDROXYLASE TRANSCRIPTION THROUGH HIGHLY CONSERVED G- QUADRUPLEX FORMING SEQUENCE IN THE PROMOTER

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2016, PHD, Kent State University, College of Arts and Sciences / Department of Chemistry.
Nucleic acids are extremely flexible biopolymer which can assume diverse conformational structures. Although in living systems, DNA remains largely as double helical structure; guanine-rich sequences can exist in unconventional structural forms known as G-quadruplex (GQ). G-quadruplex structures are composed of G-quartets stacked on top of each other, stabilized by monovalent cations such as K+ and Na+. It has been reported that the promoter regions of genes are rich in GQ motifs relative to the rest of the genome and more than 40% of human gene promoters contain at least one or more GQ motifs implying that they might play important role in gene expression especially at the transcriptional level. Interestingly we identified a highly conserved potential GQ forming sequence (wtTH49) in the human TH promoter close to the transcription starting site (TSS). The close proximity of this sequence to transcription binding sites, TATA box and CRE further emphasized a potential regulatory role it may play in TH transcription. Tyrosine hydroxylase (TH) catalyzes the first and rate-limiting step of the catecholamine (CA) biosynthetic pathway which produces dopamine, norepinephrine and epinephrine. Dysregulation of this gene is related to several common neurological disorders such as Parkinson’s and Schizophrenia. Biochemical and biophysical studies such as DMS footprinting, Circular Dichroism (CD) and polymerase stop assay confirmed that the TH49 sequence adopts two major GQ structures (5'GQ and 3'GQ). The reporter functional assay revealed that these distinct GQ structures regulate the TH transcription in a contrasting manner, albeit the 3'GQ reduces TH transcription while the 5'GQ enhances TH transcription. The transcription augmentation by the 5'GQ is the premier example of a GQ in the promoter that acts as an enhancing element. This observation is further clarified using GQ interacting small molecule, TMPyP4. Then, we rationally designed a DNA oligonucleotide based approach, DNA GQ Clip, in order to favor formation of specific GQ structure by wtTH49. This strategy allowed us not only to modulate the endogenous TH expression but also cellular dopamine level which is synthesized in the subsequent step of the catecholamine biosynthesis.
Soumitra Basu, PhD (Advisor)
190 p.

Recommended Citations

Citations

  • Farhath, M. (2016). DIFFERENTIAL REGULATION OF TYROSINE HYDROXYLASE TRANSCRIPTION THROUGH HIGHLY CONSERVED G- QUADRUPLEX FORMING SEQUENCE IN THE PROMOTER [Doctoral dissertation, Kent State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=kent1478722999704543

    APA Style (7th edition)

  • Farhath, Mohamed . DIFFERENTIAL REGULATION OF TYROSINE HYDROXYLASE TRANSCRIPTION THROUGH HIGHLY CONSERVED G- QUADRUPLEX FORMING SEQUENCE IN THE PROMOTER. 2016. Kent State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=kent1478722999704543.

    MLA Style (8th edition)

  • Farhath, Mohamed . "DIFFERENTIAL REGULATION OF TYROSINE HYDROXYLASE TRANSCRIPTION THROUGH HIGHLY CONSERVED G- QUADRUPLEX FORMING SEQUENCE IN THE PROMOTER." Doctoral dissertation, Kent State University, 2016. http://rave.ohiolink.edu/etdc/view?acc_num=kent1478722999704543

    Chicago Manual of Style (17th edition)