Synchrotron-based FTIR microspectroscopy reveals DNA methylation profile in DNA-HALO structure
Abstract:
Fourier transform infrared (FTIR) spectroscopy is a rapid, non-destructive and label-free technique for identifying subtle changes in all bio-macromolecules, and has been used as a method of choice for studying DNA conformation, secondary DNA structure transition and DNA damage. In addition, the specific level of chromatin complexity is introduced via epigenetic modifications forcing the technological upgrade in the analysis of such an intricacy. As the most studied epigenetic mechanism, DNA methylation is a major regulator of transcriptional activity, involved in the suppression of a broad spectrum of genes and its deregulation is involved in all non-communicable diseases. The present study was designed to explore the use of synchrotron-based FTIR analysis to monitor the subtle changes in molecule bases regarding the DNA methylation status of cytosine in the whole genome. In order to reveal the conformation-related best sample for FTIR-based DNA methylation analysis in situ, we used methodology for nuclear HALO preparations and slightly modified it to isolated DNA in HALO formations. Nuclear DNA-HALOs represent samples with preserved higher-order chromatin structure liberated of any protein residues that are closer to native DNA conformation than genomic DNA (gDNA) isolated by the standard batch procedure. Using FTIR spectroscopy we analyzed the DNA methylation profile of isolated gDNA and compared it with the DNA-HALOs. This study demonstrated the potential of FTIR microspectroscopy to detect DNA methylation marks in analyzed DNA-HALO specimens more precisely in comparison with classical DNA extraction procedures that yield unstructured whole genomic DNA. In addition, we used different cell types to assess their global DNA methylation profile, as well as defined specific infrared peaks that can be used for screening DNA methylation.
Note:
This is accepted manuscript version of the article: A. Sarić, J. Rajić, A. Tolić, T. Dučić, M.Vidaković, Synchrotron-based FTIR microspectroscopy reveals DNA methylation profile in DNA-HALO structure, Spectrochim. Acta A Mol. Biomol. Spectrosc. (2023):, 123090. https://doi.org/10.1016/j.saa.2023.123090
Keywords:
Chromatin; HALO structure; Epigenetic marks; DNA methylation; FTIR microspectroscopySource:
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy, 2023, 302, 123090Funding / projects:
- ALBA In-house grant: “Towards FTIR spectroscopy of single living cells” PI T. Ducic, 2018093034
- COST Action CA1406
- COST Action CA16119
- Ministry of Science, Technological Development and Innovation of the Republic of Serbia, institutional funding - 200007 (University of Belgrade, Institute for Biological Research 'Siniša Stanković') (RS-MESTD-inst-2020-200007)
Related info:
- Version of
https://radar.ibiss.bg.ac.rs/handle/123456789/5947 - Referenced by
https://radar.ibiss.bg.ac.rs/handle/123456789/5960
DOI: 10.1016/j.saa.2023.123090
ISSN: 1386-1425