The effect of a G-quadruplex structure in the Oct4 3′-untranslated region (UTR) on its binding interactions with miR-145-5p and FMRP

Cancer is the second leading cause of death worldwide, due to its narrow treatment window and the chances of remission and resistance. A major area of interest are chemo-resistance cells known as cancer stem cells (CSCs), due to their prevalence in cancer relapse cases. Oct4 (octamer-binding transcription factor 4), a master transcription factor, has been found to be aberrantly expressed in cancer cells and has been linked to the growth of CSCs. However, the molecular mechanism of Oct4 translation regulation in relation to cancer stemness has yet to be explored fully. We determined that the Oct4 mRNA has a guanine rich sequence in its 3′-untranslated region (UTR) which we postulated folds into a G quadruplex (GQ) structure. Thus, this study focuses on the characterization of this predicted GQ in the Oct4 3′-UTR, and its binding interactions with miR-145-5p, a microRNA shown to regulate the translation of Oct4. We used native polyacrylamide gel electrophoresis (PAGE), circular dichroism (CD) spectroscopy, and 1H nuclear magnetic resonance (NMR) spectroscopy to demonstrate the formation of the Oct4 3′-UTR GQ structure and its modulation of the miR-145-5p binding site. The characterization of miR-145-5p binding to the Oct4 mRNA 3′-UTR, along with its overlap with the GQ structure, indicates a potential molecular switch for Oct4 translation regulation.