5-Fluorouracil Treatment Alters the Efficiency of Translational Recoding

Abstract

5-fluorouracil (5-FU) is a chemotherapeutic agent that has been extensively studied since its initial development in the 1950s. It has been suggested that the mechanism of action of 5-FU involves both DNA- and RNA-directed processes, but this has remained controversial. In this study, using a series of in vivo reporter constructs capable of measuring translational recoding, we demonstrate that cells exposed to 5-FU display a reduced capacity to engage in a variety of translational recoding events, including +1 programmed frameshifting (PRF) and −1 PRF. In addition, 5-FU-treated cells are much less accurate at stop codon recognition, resulting in a significant increase in stop codon-readthrough. Remarkably, while the efficiency of cap-dependent translation appears to be unaffected by 5-FU, 5-FU-treated cells display a decreased ability to initiate cap-independent translation. We further show that knockdown of thymidylate synthase, an enzyme believed to be at the center of 5-FU-induced DNA damage, has no effect on the observed alterations in translational recoding. On the other hand, ribosomal RNA (rRNA) pseudouridylation, which plays an important role in translational recoding, is significantly inhibited. Taken together, our results suggest that the observed effect of 5-FU on recoding is an RNA-directed effect. Our results are the first to show definitely and quantitatively that translational recoding is affected by exposure to 5-FU. Thus, it is possible that a substantial portion of 5-FU cytotoxicity might possibly be the result of alterations in translational recoding efficiency.

Targeting the cell and non-cell autonomous regulation of 47S synthesis by GCN2 in colon cancer

“…In this context, gastrointestinal cancers may constitute interesting types of cancer models since they are classically treated using FOLFOX or FOLFIRI, two chemotherapeutic regimens containing drugs targeting the ribosomal function. Indeed, oxaliplatin was reported to kill cells through the induction of a ribosomal stress and 5-fluorouracil was described to impair 47S processing and recently to be integrated in the ribosome and promoting profound translational reprogramming (Bruno et al, 2017; Burger et al, 2010; Ge et al, 2017; Sutton and DeRose, 2021; Therizols et al, 2022). Nonetheless, these compounds are mainly effective on proliferative cells since metabolic stress limits their efficacy (Xu et al, 2019).…”

Translational readthrough of nonsense mutant TP53 by mRNA incorporation of 5-Fluorouridine

“…This efficient incorporation of FUr is most likely the reason for the time-dependent decrease in free intracellular FUr levels (Table S1 ). Stop codon readthrough by 5-FU has previously been reported in dual-luciferase reporter plasmids and suggested to be due to inhibition of rRNA pseudouridylation [ 39 ]. We also show that combination treatment with FUr and uridine causes a dose-dependent decrease in full-length p53 levels.…”

Diversity in Biological Function and Mechanism of the tRNA Methyltransferase Trm10

“…Importantly, the antimetabolite 5FU is a common chemotherapeutic drug used to treat several solid cancer tumors, including breast, colorectal, and neck cancers in humans, raising questions about similar synergies with tRNA modifications, including Trm10, in vertebrates that will be important to study in the future. Along these lines, 5FU treatment perturbs several translation-associated events in a human colorectal cancer cell line including decreased ability to perform ±1 programmed frameshifting and cap independent translation, and decreased recognition of STOP codons that might suggest such similarities. , The antimetabolite also directly affects RNA modification by inhibiting essential pseudouridine synthases and depleting the pseudouridine modification which is normally abundant in tRNA and other RNA molecules and is also important to pre-mRNA splicing and translational recoding. , …”

“…Along these lines, 5FU treatment perturbs several translation-associated events in a human colorectal cancer cell line including decreased ability to perform ±1 programmed frameshifting and cap independent translation, and decreased recognition of STOP codons that might suggest such similarities. 24,25 The antimetabolite also directly affects RNA modification by inhibiting essential pseudouridine synthases and depleting the pseudouridine modification which is normally abundant in tRNA and other RNA molecules and is also important to pre-mRNA splicing and translational recoding. 25,26 A 2010 study reported the genetic interaction profiles for 75% of genes in S. cerevisiae and revealed a strong synthetic interaction between PUS3 and TRM10 genes.…”