Response of PRIMPOL-Knockout Human Lung Adenocarcinoma A549 Cells to Genotoxic Stress

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Anastasia S. Gromova^1,2#, Elizaveta O. Boldinova^1,2#, Daria V. Kim^3,4, Roman N. Chuprov-Netochin⁵, Sergey V. Leonov^5,6, Margarita V. Pustovalova⁵, Dmitry O. Zharkov^3,4,a*, and Alena V. Makarova^1,2,b*

¹Institute of Molecular Genetics, Kurchatov Institute National Research Center, 123182 Moscow, Russia

²Institute of Gene Biology, Russian Academy of Sciences, 119334 Moscow, Russia

³Institute of Chemical Biology and Fundamental Medicine, Siberian Branch of the Russian Academy of Sciences, 630090 Novosibirsk, Russia

⁴Novosibirsk State University, 630090 Novosibirsk, Russia

⁵School of Biological and Medical Physics, Moscow Institute of Physics and Technology, 141700 Dolgoprudny, Russia

⁶Institute of Cell Biophysics of the Russian Academy of Sciences, 142290 Pushchino, Russia

^* To whom correspondence should be addressed.

^# These authors contributed equally to the study.

Received December 9, 2022; Revised August 8, 2023; Accepted September 15, 2023

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Human DNA primase/polymerase PrimPol synthesizes DNA primers de novo after replication fork stalling at the sites of DNA damage, thus contributing to the DNA damage tolerance. The role of PrimPol in response to the different types of DNA damage is poorly understood. We knocked out the PRIMPOL gene in the lung carcinoma A549 cell line and characterized the response of the obtained cells to the DNA damage caused by hydrogen peroxide, methyl methanesulfonate (MMS), cisplatin, bleomycin, and ionizing radiation. The PRIMPOL knockout reduced the number of proliferating cells and cells in the G2 phase after treatment with MMS and caused a more pronounced delay of the S phase in the cisplatin-treated cells. Ionizing radiation at a dose of 10 Gy significantly increased the content of apoptotic cells among the PRIMPOL-deficient cells, while the proportion of cells undergoing necroptosis increased in both parental and knockout cells at any radiation dose. The viability of PRIMPOL-deficient cells upon the hydrogen peroxide-induced oxidative stress increased compared to the control cells, as determined by the methyl tetrazolium (MTT) assay. The obtained data indicate the involvement of PRIMPOL in the modulation of adaptive cell response to various types of genotoxic stress.

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KEY WORDS: replication, DNA damage, damage tolerance, apoptosis, PRIMPOL, primase, knockout cell lines

DOI: 10.1134/S0006297923110214

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