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Whole transcriptome sequencing of HCT116 cells with altered DNA repair capacities treated with RTX or 5FdUR

ABSTRACT: Thymidylate synthase (TS) is a key enzyme in thymidylate biosynthesis and an established target of chemotherapeutics such as 5-fluoro-2’-deoxyuridine (5FdUR) and raltitrexed (RTX). Inhibition of TS disrupts the dUTP:dTTP balance, leading to uracil misincorporation, futile base excision repair cycles, DNA strand breaks, and ultimately cell death. However, active U-DNA repair is apparently not required for the cytotoxic effect of these drugs, indicating that alternative pathways contribute to the observed S-phase arrest. To dissect these mechanisms, we investigated the transcriptomic effects of TS inhibition in HCT116 colon cancer cell lines with altered DNA repair capacities treated with 5-fluoro-2′-deoxyuridine (5FdUR) or raltitrexed (RTX). Both drugs induced similar DNA damage responses and S-phase arrest, yet displayed distinct transcriptional signatures. Our findings underscore the multifaceted impact of TS inhibition and reveal drug-specific differences in cellular responses.

ORGANISM(S): Homo sapiens

PROVIDER: GSE318306 | GEO | 2026/07/08

REPOSITORIES: GEO

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Thymidylate synthase RIP-seq from DNA-repair-deficient HCT116 cells treated with RTX or 5FdUR [RIP-Seq]

Project description:Thymidylate synthase (TS) is a key enzyme in thymidylate biosynthesis and an established target of chemotherapeutics such as 5-fluoro-2’-deoxyuridine (5FdUR) and raltitrexed (RTX). Inhibition of TS disrupts the dUTP:dTTP balance, leading to uracil misincorporation, futile base excision repair cycles, DNA strand breaks, and ultimately cell death. Beyond its catalytic role, TS also binds RNA, autoregulating its own translation and interacting with transcripts such as p53 and c-myc, thereby linking TS activity to broader post-transcriptional regulatory networks. To better understand how TS inhibitory drugs influence this network, we performed TS-coupled RNA-coimmunoprecipitation (TS-RIP) and sequencing from mismatch repair-deficient, UNG-inhibited HCT116 colon cancer cells treated with 5FdUR or RTX. TS-RIP revealed RNA-binding targets of TS, in addition to the known p53 and TS mRNAs. Our findings underscore the multifaceted impact of TS inhibition, linking enzymatic disruption to RNA-level regulation and revealing drug-specific differences in cellular responses.

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Whole transcriptome sequencing of HCT116 cells with altered DNA repair capacities treated with RTX or 5FdUR

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