AFM imaging of the transcriptionally active chromatin in mammalian cells' nuclei.

Autor: Bairamukov VY; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia. Electronic address: bayramukov_vy@pnpi.nrcki.ru., Filatov MV; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Kovalev RA; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Fedorova ND; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Pantina RA; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Ankudinov AV; The Ioffe Physical-Technical Institute of the Russian Academy of Sciences, 26, Politekhnicheskaya, 194021 Saint Petersburg, Russia., Iashina EG; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Grigoriev SV; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia., Varfolomeeva EY; Petersburg Nuclear Physics Institute Named by B.P. Konstantinov of NRC 'Kurchatov Institute', 1, Orlova Roshcha, 188300 Gatchina, Russia.
Jazyk: angličtina
Zdroj: Biochimica et biophysica acta. General subjects [Biochim Biophys Acta Gen Subj] 2022 Dec; Vol. 1866 (12), pp. 130234. Date of Electronic Publication: 2022 Aug 22.
DOI: 10.1016/j.bbagen.2022.130234
Abstrakt: Background: Nuclear rigidity is traditionally associated with lamina and densely packed heterochromatin. Actively transcribed DNA is thought to be less densely packed. Currently, approaches for direct measurements of the transcriptionally active chromatin rigidity are quite limited.
Methods: Isolated nuclei were subjected to mechanical stress at 60 g and analyzed by Atomic Force Microscopy (AFM).
Results: Nuclei of the normal fibroblast cells were completely flattened under mechanical stress, whereas nuclei of the cancerous HeLa were extremely resistant. In the deformed HeLa nuclei, AFM revealed a highly-branched landscape assembled of ~400 nm closed-packed globules and their structure was changing in response to external influence. Normal and cancerous cells' isolated nuclei were strikingly different by DNA resistance to applied mechanical stress. Paradoxically, more transcriptionally active and less optically dense chromatin of the nuclei of the cancerous cells demonstrated higher physical rigidity. A high concentration of the transcription inhibitor actinomycin D led to complete flattening of HeLa nuclei, that might be related to the relaxation of supercoiled DNA tending to deformation. At a low concentration of actinomycin D, we observed the intermediary formation of stochastically distributed nanoloops and nanofilaments with different shapes but constant width ~ 180 nm. We related this phenomenon with partial DNA relaxation, while non-relaxed DNA still remained rigid.
Conclusions: The resistance to deformation of nuclear chromatin correlates with fundamental biological processes in the cell nucleus, such as transcription, as assessed by AFM.
General Significance: A new outlook to studying internal nuclei structure is proposed.
Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(Copyright © 2022 Elsevier B.V. All rights reserved.)
Databáze: MEDLINE
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