Project description:We performed RNA-seq on dTAG-NUDT21 HCT116 cells treated with DMSO+JTE-607 or dTAGV1+JTE-607 for 48 hours with 4 biological replicates
Project description:To investigate the function of NUDT21 in the regulation of progression of colorectal cancer, we established HCT116 cell line in which each target gene has been knocked down by shRNA. We then performed gene expression profiling analysis using data obtained from RNA-seq of 4 different cells at two time points.
Project description:To assess the impact of CPA inhibition by JTE-607 on intra-cell-line transcriptomic heterogeneity, we profiled two lung cancer cell lines treated with DMSO or JTE-607 by 3' tag-based single-cell RNA-seq with 10x Chromium. For data analysis, we performed transcriptome quantification at the PAS-based transcript level to generate a PAS-by-cell count matrix, and then investigate the effect of JTE-607 treatment on PAS usage pattern and cellular states.
Project description:HCC is the third leading cause of cancer-related deaths worldwide. However, the molecular mechanisms underlying the progression of HCC is still largely elusive. NUDT21 (CFIm25) is an important mediator of 3′ UTR APA and demonstrates a causal relationship between alternative polyadenylation and cancer cell proliferation. Although the function of NUDT21 has been explored in glioblastoma tumor, the functional significance of NUDT21 in solid tumors is not well understood. In this study, we observed that NUDT21 is suppressed in human HCC tissues, compared to adjacent noncancerous tissues. In addition we found that the HCC patients with suppressed NUDT21 are statistically associated with poor outcomes. These observations suggest NUDT21 possibly functions as tumor suppressor in hepatocellular carcinoma (HCC).
Project description:Stem cells continually self-renew and differentiate to sustain tissue homeostasis, yet the role of post-transcriptional mechanisms in guiding these processes remains incompletely understood. Here, we demonstrate that the regulation of 3’UTR length via alternative mRNA polyadenylation (APA) is essential for stem cell function across diverse tissues. Modulating the APA regulator Nudt21 reveals that stem cell self-renewal and differentiation depend on distinct dosage thresholds and thus can be uncoupled. Specifically, moderate Nudt21 suppression elicits a maturation arrest of stem cells due to 3’UTR-shortening of differentiation-associated mRNAs that escape miRNA regulation and perturb ceRNA networks. By contrast, complete Nudt21 suppression additionally shortens the 3’UTRs of mRNAs encoding essential multiprotein complexes, including the nuclear pore, leading to complex destabilization, proteotoxic stress, DNA damage, and cell cycle arrest. Critically, deletion of the alternative 3’UTRs of individual nucleoporins recapitulates defects observed with Nudt21 loss. We further demonstrate that the co-translational assembly of dozens of protein complexes is impaired in Nudt21-deficient cells, providing a mechanistic framework for compromised complex integrity. Collectively, our results show that APA plays distinct, dose-dependent roles in stem cell homeostasis by fine-tuning the expression of differentiation-associated genes and coordinating the biogenesis of multiprotein complexes essential for cell cycle progression.
Project description:Alternative polyadenylation (APA) generates mRNAs with distinct 3′-untranslated regions (3′ UTRs) to regulate post-transcriptional gene expression and is critical in many tumors. However, the role of alternative 3′ UTRs in neurolastoma (NB) remains elusive. Here, we show that NUDT21 knockdown inhibits NB cell proliferation, induces cycle arrest/apoptosis in vitro, and suppresses xenograft growth in nude mice. Mechanistically, NUDT21 silencing triggers global APA dynamics, with ∼85% of APA genes switching to proximal poly(A) sites, shortening 3′ UTRs. Integrative PAS-seq/RNA-seq analyses reveal that NUDT21 regulates E2F targets (e.g., MKI67) via APA. Additionally, NUDT21 occupies gene regulatory regions to directly activate E2F target transcription. Our findings demonstrate NUDT21 sustains NB proliferation through dual APA-dependent and APA-independent mechanisms, highlighting targeting NUDT21 or its downstream pathways as a promising therapeutic strategy.
Project description:Nuclear export factor 1 (NXF1) is the principal mRNA export receptor in eukaryotes. To characterize the role of NXF1 in regulating alternative polyadenylation (APA) dynamics, we employed the dTAG degradation system in a HeLa-NXF1-dTAG stable cell line. Cells were treated with 500 nM dTAG-13 or DMSO (vehicle control) for 6 hours to induce acute NXF1 depletion. Total RNA was subjected to 3' end sequencing (QuantSeq 3' mRNA-Seq Library Prep Kit V2, Lexogen) for APA analysis. Each condition was performed in biological duplicate.