The lack of functional DNMT2/TRDMT1 gene modulates cancer cell responses during drug-induced senescence

Abstract

Cellular senescence may be a side effect of chemotherapy and other anti-cancer treatments that may promote inflammation and paracrine secondary senescence in healthy tissues. DNMT2/TRDMT1 methyltransferase is implicated in the regulation of cellular lifespan and DNA damage response (DDR). In the present study, the responses to senescence inducing concentrations of doxorubicin and etoposide in different cancer cells with DNMT2/TRDMT1 gene knockout were evaluated, namely changes in the cell cycle, apoptosis, autophagy, interleukin levels, genetic stability and DDR, and 5-mC and NSUN1-6 levels. Moreover, the effect of azacytidine post-treatment was considered. Diverse responses were revealed that was based on type of cancer cells (breast and cervical cancer, osteosarcoma and glioblastoma cells) and anti-cancer drugs. DNMT2/TRDMT1 gene knockout in drug-treated glioblastoma cells resulted in decreased number of apoptotic and senescent cells, IL-8 levels and autophagy, and increased number of necrotic cells, DNA damage and affected DDR compared to drug-treated glioblastoma cells with unmodified levels of DNMT2/TRDMT1. We suggest that DNMT2/TRDMT1 gene knockout in selected experimental settings may potentiate some adverse effects associated with chemotherapy-induced senescence.

5-Azacytidine Inhibits the Activation of Senescence Program and Promotes Cytotoxic Autophagy during Trdmt1-Mediated Oxidative Stress Response in Insulinoma β-TC-6 Cells

“…We have previously shown that 2 h treatment with 100 µM HP promoted SIPS in normal human fibroblasts such as WI-38 and BJ cells that were accompanied by elevated levels of reactive oxygen species (ROS) [ 17 ]. In the present study, the same concentration of HP was used, but insulinoma cells were stimulated with HP for 24 h. 5-AzaC post-treatment (1 µM, 24 h) was applied as previously reported using a doxorubicin- and etoposide-induced senescence model using four genetically different cancer cell lines [ 11 ]. HP treatment reduced the cell number of both insulinoma cell lines, but the cytostatic effects of HP were more pronounced in β-TC-6 cells compared to NIT-1 cells ( Figure 1 A).…”

“…Cells were passaged by detaching with the trypsin-EDTA solution (Corning, Tewksbury, MA, USA, 25-053-CI). Both cell lines were seeded at the total number of 1 × 10 5 cells per well in a 6-well plate and cultured until approximately 70–80% confluency was reached (3 to 4 days) and then treated with 100 µM hydrogen peroxide (H 2 O 2 ) for 24 h (HP, Merck KGaA, Darmstadt, Germany) for the induction of oxidative stress, or with 1 µM 5-azacitidine for 24 h (5-azaC, Merck KGaA, Darmstadt, Germany) for the inhibition of methylation or cells were treated with 100 µM HP for 24 h and then the medium was discarded and cells were post-treated with 1 µM 5-azaC for 24 h. The concentrations of HP and 5-azaC were selected on the basis of our previously published protocols [ 11 , 17 ]. Except for the assay of senescence-associated β-galactosidase activity, all parameters were analyzed immediately after 5-azaC post-treatment.…”

“…After HP and 5-azaC treatments, phosphatidylserine externalization (an apoptotic biomarker) was analyzed using a Muse ® Cell Analyzer and Muse ® Annexin V and Dead Cell Assay Kit (Luminex Corporation, Austin, TX, USA) as described elsewhere [ 11 ].…”

“…A side effect of chemotherapy may be stress-induced premature senescence (SIPS) observed in both treated cancer cells as well as normal adjacent cells [ 10 , 11 ]. Normal and neoplastic senescent cells can secrete numerous proinflammatory agents into their microenvironment (senescence-associated secretory phenotype, SASP) that can promote proliferation, migration, invasiveness of cancer cells, angiogenesis, epithelial-mesenchymal transition (EMT), and metastasis [ 12 , 13 , 14 ].…”

Comprehensive Analysis of the Prognostic Value and Immune Infiltrates of the Three-m5C Signature in Colon Carcinoma

“… 33 TRDMT1 (tRNA aspartic acid methyltransferase 1), also known as DNMT2, promoting tRNA stability and protein synthesis, participates in cell cycle, apoptosis, autophagy and interleukin levels of many tumors. 34 , 35 However, there are few reports on the correlation between TRDMT1 and colon cancer. NSUN6 (NOP2/Sun RNA methyltransferase 6), is a member of the family of NSUN proteins, which is associated with tRNAs and acts as a tRNA methyltransferase.…”