An Investigation of RNA Methylations with Biophysical Approaches in a Cervical Cancer Cell Model

An Investigation of RNA Methylations with Biophysical Approaches in a Cervical Cancer Cell Model

RNA methylation adds a second layer of genetic information that dictates the post-transcriptional fate of RNAs. Although various methods exist that enable the analysis of RNA methylation in a site-specific or transcriptome-wide manner, whether biophysical approaches can be employed to such analyses...

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Bibliographic Details
Main Authors: Buket Sağlam, Onur Akkuş, Azime Akçaöz-Alasar, Çağatay Ceylan, Günnur Güler, Bünyamin Akgül
Format: Article
Language:English
Published: MDPI AG 2024-11-01
Series:Cells
Subjects:
m<sup>6</sup>A
<i>N</i><sup>6</sup>-methyladenosine
FT-IR
circular dichroism
apoptosis
TNF-α
Online Access:https://www.mdpi.com/2073-4409/13/22/1832
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Summary:RNA methylation adds a second layer of genetic information that dictates the post-transcriptional fate of RNAs. Although various methods exist that enable the analysis of RNA methylation in a site-specific or transcriptome-wide manner, whether biophysical approaches can be employed to such analyses is unexplored. In this study, Fourier-transform infrared (FT-IR) and circular dichroism (CD) spectroscopy are employed to examine the methylation status of both synthetic and cellular RNAs. The results show that FT-IR spectroscopy is perfectly capable of quantitatively distinguishing synthetic m<sup>6</sup>A-methylated RNAs from un-methylated ones. Subsequently, FT-IR spectroscopy is successfully employed to assess the changes in the extent of total RNA methylation upon the knockdown of the m<sup>6</sup>A writer, METTL3, in HeLa cells. In addition, the same approach is shown to accurately detect reduction in total RNA methylation upon the treatment of HeLa cells with tumor necrosis factor alpha (TNF-α). It is also demonstrated that m<sup>1</sup>A and m<sup>6</sup>A methylation induce quite a distinct secondary structure on RNAs, as evident from CD spectra. These results strongly suggest that both FT-IR and CD spectroscopy methods can be exploited to uncover biophysical properties impinged on RNAs by methyl moieties, providing a fast, convenient and cheap alternative to the existing methods.
ISSN:2073-4409

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