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Restoration of miRNA-149 Expression by TmPyP4 Induced Unfolding of Quadruplex within Its Precursor

Author:
Ghosh, Arpita, Ekka, Mary Krishna, Tawani, Arpita, Kumar, Amit, Chakraborty, Debojyoti, Maiti, Souvik
Source:
Biochemistry 2018 v.58 no.6 pp. 514-525
ISSN:
1520-4995
Subject:
DNA, breast neoplasms, calorimetry, cell proliferation, cellular microenvironment, circular dichroism spectroscopy, gene expression, microRNA, neoplasm cells, non-coding RNA, nuclear magnetic resonance spectroscopy, nucleotide sequences, porphyrins, proteins, quantitative polymerase chain reaction, reverse transcriptase polymerase chain reaction, titration, transcription (genetics), translation (genetics), ultraviolet-visible spectroscopy
Abstract:
Noncoding RNAs are functional RNA molecules that get transcribed from DNA but are not translated into proteins; yet, they can regulate gene expression at transcriptional and post-transcriptional levels. Secondary structures present within these RNAs play a major role in determining their nature of function. In the case of miRNAs, the precursor miRNA have a hairpin stem loop structure which is required for Dicer recognition and further maturation. Alternately, it might assume a G-quadruplex structure. The transition from hairpin to G-quadruplex depends upon the nucleotide sequence as well as the cellular microenvironment, and this might affect the miRNA maturation and other downstream activity. Formation of the G-quadruplex within precursor miRNA-149 has been shown to inhibit Dicer processing activity followed by suppression of miRNA-149 maturation in cancer cells. In this report, we show that suppression of cell proliferation by the upregulated miRNA-149 could be rescued by unfolding the G-quadruplex present in pre-miRNA-149 by TmPyP4 (Porphyrin) treatment. Using UV–visible spectroscopy, circular dichroism, and isothermal titration calorimetry, we observed that TmPyP4 binds strongly to G-quadruplex and unfolds it, which was further verified by NMR spectroscopy. In cellulo, qRT-PCR measurements of miRNA-149 in MCF-7 breast cancer cells showed concentration dependent enhancement of mature miRNA-149 upon treatment of TmPyP4. As a consequence of enhanced miRNA-149 activity, we also observe the reduction in miRNA-149 target protein ZBTB2 that eventually leads to reduced cell proliferation.
Agid:
6305917