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Ryota Kurimoto
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P64 - Tumor Biology and Systems Biology - Basic and Translational Science - miRNA (ID 202)
- Event: WCLC 2020
- Type: Posters
- Track: Tumor Biology and Systems Biology - Basic and Translational Science
- Presentations: 1
- Moderators:
- Coordinates: 1/28/2021, 00:00 - 00:00, ePoster Hall
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P64.03 - RNA Modification Enzyme TruB1 Regulate Tumor Proliferation via MicroRNA let-7 (ID 2928)
00:00 - 00:00 | Presenting Author(s): Ryota Kurimoto
- Abstract
Introduction
The biogenies of microRNAs (miRNAs) is tightly regulated at multiple step by RNA-binding proteins. Indeed, let-7, which is involved in various pathophysiological events, is specifically regulated at its maturation step by multiple factors including Lin28A/B. To further understand the molecular mechanism of let-7 biogenesis, a gain-of-function cell-based screening with let-7 sensor luciferase reporter was performed (Kurimoto et al., EMBO J (2020)39:e104708). Moreover, we also evaluate the potential of this candidate as the novel tumor suppressor.
Methods
We performed cell-based, gain-of-function screening to search the novel let-7 regulators to whole RNA binding proteins (1469 genes) and picked up the several candidate genes. Next, qPCR and TaqMan array were performed to assess the specificity of the effect of the candidate genes on microRNAs. To elucidate the mechanism by which the candidate gene regulates let-7 maturation, we performed northern blots and in vitro reactions. Furthermore, binding of candidate genes to let-7 was confirmed in vitro and in vivo (these results in: Kurimoto et al., EMBO J (2020)39:e104708). We evaluated the effects of this candidate gene on cell proliferation and stemness in MTT assay, sphere formation assay and Xenograft model by using A549 cells.
Results
In resulted, TruB1, known as a pseudouridine synthase of tRNA, was identified. In overexpression, knockdown experiments and in vitro processing assay, TruB1 specifically enhanced microprocessing from pri- into pre- and mature-let-7 with its intrinsic enzymatic activity independent manner. HITS-CLIP (High throughput sequencing crosslinking immunoprecipitation) for endogenous TruB1 showed that TruB1 had been directly bound to the stem-loop structure of pri-let-7. TruB1 increased the affinity between DGCR-8 and let-7 by RIP (RNA immunoprecipitation) assay (these results in: Kurimoto et al., EMBO J (2020)39:e104708). TruB1 suppressed cell proliferation and sphere formation, which was mediated in part by let-7 in human lung adenocarcinoma cell line, A549.
Conclusion
These data reveal the novel function of TruB1 in specific miRNA biogenesis via its non-enzymatic manner. TruB1-let-7 axis might show the tumor suppressive effect in lung cancer.
