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Background Long noncoding RNAs (lncRNAs) play crucial roles in tumorigenesis, and lncRNA taurine-upregulated gene 1 ( TUG1 ) has been proven to be associated with several human cancers. However, the mechanisms of TUG1 -involved regulation remain largely unknown. Methods We examined the expressions of TUG1 in a cohort of 89 patients with non-small cell lung cancer (NSCLC) to determine the association between TUG1 expression and clinical parameters. We used circular chromosome conformation capture (4C) coupled with next-generation sequencing to explore the genome regions that interact with TUG1 and the TUG1 -mediated regulation. Results TUG1 was significantly downregulated, and the TUG1 downregulation correlated with sex ( p  = 0.006), smoking status ( p  = 0.016), and tumor differentiation grade ( p  = 0.001). Knockdown of TUG1 significantly promoted the proliferation of NSCLC cells. According to the bioinformatic analysis result of TUG1 4C sequencing data, 83 candidate genes and their interaction regions were identified. Among these candidate genes, CUGBP and Elav-like family member 1 ( CELF1 ) are potential targets of TUG1 in-trans regulation. To confirm the interaction between TUG1 and CELF1 , relative expressions of CELF1 were examined in TUG1 knockdown H520 cells; results showed that CELF1 was significantly upregulated in TUG1 knockdown H520 cells. RNA immunoprecipitation was then performed to examine whether TUG1 RNA was bound to PRC2, a TUG1 -involved regulation mechanism reported in previous studies. The results demonstrated that TUG1 RNA was bound to enhancer of zeste protein 2/embryonic ectoderm development (EZH2/EED), which is essential for PRC2. Finally, our designed ChIP assay revealed that the EZH2/EED was bound to the promotor region of CELF1 within 992 bp upstream of the transcript start site. Conclusion TUG1 is downregulated in NSCLC. Using TUG1 4C sequencing and bioinformatic analysis, we found CELF1 to be a potential target of TUG1 RNA in in-trans regulation. Moreover, subsequent experiments showed that TUG1 RNA could bind to PRC2 in the promotor region of CELF1 and negatively regulate CELF1 expressions in H520 cells. Our results may facilitate developing new treatment modalities targeting TUG1 /PRC2/ CELF1 interactions in patients with NSCLC.

Intron retention (IR) is one of the cellular mechanisms to perform alternative splicing and thus control gene expression in several mammalian cellular pathways. IR in PI-PLC γ1 mRNA was observed in some primary synoviocyte samples from osteoarthritis (OA) patients, likely due to inter-patient variability. The aim of the present manuscript was to explore the PI-PLC γ1 IR molecular mechanism as a consequence of nutraceutical treatment of synoviocytes and the molecular basis of individual response. To evaluate the gene expression modulation of molecules involved in mRNA splicing, an RNA-seq analysis was performed, and the transcription modulation of six differentially expressed genes was validated by RT-PCR. Moreover, through a silencing experiment, the relationship between PI-PLC γ1 IR and the six modulated genes was explored. Finally, two of them, the RNA-binding proteins CELF1 and PTBP3, whose mRNA levels were elevated in samples exhibiting IR, were analyzed in detail. CELF1 and PTBP3 were overexpressed in synoviocytes lacking PI-PLC γ1 IR, and we found that CELF1 was responsible for IR, whereas PTBP3 did not seem to be involved. In conclusion, in our experimental model, the role of CELF1 protein in PI-PLC γ1 IR was explored, opening new scenarios for understanding the molecular mechanisms underlying the IR phenomenon present in several kinds of diseases.

Melanomas are well-known for their altered mRNA expression profiles. Yet, the specific contribution of mRNA binding proteins (mRBPs) to melanoma development remains unclear. Here we identify a cluster of melanoma-enriched genes under the control of CUGBP Elav-like family member 1 (CELF1). CELF1 was discovered with a distinct prognostic value in melanoma after mining the genomic landscape of the 692 known mRBPs across different cancer types. Genome-wide transcriptomic, proteomic, and RNA-immunoprecipitation studies, together with loss-of-function analyses in cell lines, and histopathological evaluation in clinical biopsies, revealed an intricate repertoire of CELF1-RNA interactors with minimal overlap with other malignancies. This systems approach uncovered the oncogene DEK as an unexpected target and downstream effector of CELF1. Importantly, CELF1 and DEK were found to represent early-induced melanoma genes and adverse indicators of overall patient survival. These results underscore novel roles of CELF1 in melanoma, illustrating tumor type-restricted functions of RBPs in cancer. Hundreds of mRNA binding proteins (mRBPs) have been described in mammalian cells. Here, the authors identify RBPs differentially regulated in melanoma, and show the RBP CELF1 controlling a distinct set of protumorigenic factors.

RNA-binding proteins (RBPs) regulate diverse post-transcriptional processes and play roles in adipocyte development; however, their role in white fat beiging remains unclear. Here we identify CUG-BP Elav-like family member 1 (CELF1) as a key RBP promoting beiging of inguinal white adipose tissue in response to cold. Adipocyte-specific Celf1 deficiency impairs cold-induced thermogenic gene expression and reduces energy expenditure. Mechanistically, CELF1 binds to the 3′UTR of Dio2 mRNA and enhances its stability, promoting local triiodothyronine (T3) production. Notably, CELF1 expression is significantly reduced in subcutaneous fat of individuals with obesity and negatively correlates with BMI. CELF1 enhances isoproterenol-induced beige adipocyte activation and mitochondrial respiration in vitro, and Celf1 overexpression ameliorates diet-induced obesity and metabolic dysfunction. Hence, our study identifies CELF1 as a physiological regulator of metabolic stress in activating thermogenesis and promoting energy expenditure at the post-transcriptional level, highlighting its potential as a therapeutic target for obesity and metabolic diseases. This study identifies CELF1 as a key regulator of fat metabolism. CELF1 promotes beiging and thermogenesis of white fat by stabilizing Dio2 mRNA. It is reduced in obesity, and its overexpression counters diet-induced metabolic dysfunction.

Rationale Esophageal squamous cell carcinoma (ESCC) is characterized by poor prognosis. tRNA-derived fragments (tRFs), a novel class of non-coding RNAs generated by tRNA cleavage, have emerged as key regulators of cancer development. However, the functional landscape of tRFs remains underexplored in ESCC. We here identified tRF-24-RPM8309M2S ( tRF-24 ), a 5’ tRF derived from mature tRNA LeuAAG/TAG , which promotes the malignant progression of ESCC and offers a promising therapeutic target. Methods The public GSE207635 dataset from Gene Expression Omnibus (GEO) database was analyzed to identify tsRNAs involved in ESCC progression. The clinical significance of tRF-24 was investigated in samples from 96 ESCC patients. CUGBP Elav-like family member 1 (CELF1) was validated as a tRF-24 interactor through RNA pull-down assays. CCK-8 and transwell assays were applied to measure malignant cell phenotypes. mCherry-GFP-LC3 reporter assay was performed to examine the autophagy. Colocalization between LC3 and mitochondria was employed to detect mitophagy. Immunofluorescent and colony formation assay were conducted to assess the impact of DNA damage repair and cisplatin resistance in ESCC. Extracellular acidification rate (ECAR), lactate production and glucose consumption were performed to analyze changes in glycolysis. Fluorescence recovery after photobleaching (FRAP) was adopted to evaluate CELF1 phase separation. Additionally, RNA sequencing and alternative splicing analyses were conducted to determine global transcriptome alterations following tRF-24 or CELF1 overexpression. Results Our findings demonstrate that tRF-24 is significantly upregulated in tumor samples and is associated with poorer survival of ESCC patients. Depletion of tRF-24 suppresses malignant cell phenotypes in ESCC cells both in vitro and in vivo. Mechanistically, tRF-24 binds to the Ser28 residue of CELF1, inhibiting AKT1-mediated phosphorylation at this site, which facilitates CELF1 nuclear translocation and subsequent liquid-liquid phase separation (LLPS) formation. These CELF1-enriched nuclear condensates potently regulate the alternative splicing of BIN1 and BECN1 pre-mRNAs, generating pro-oncogenic BIN1-L and pro-autophagic/mitophagic BECN1-α isoforms that collectively enhance tumor malignancy by promoting tumor cell EMT, DNA damage repair, cisplatin resistance and glycolysis. Targeting tRF-24 with an antagomir significantly suppresses tumor progression in ESCC xenograft models, highlighting its therapeutic potential. Conclusions Our findings establish tRF-24 as a promising therapeutic target in the comprehensive treatment of ESCC.

Understanding the mechanisms of early cardiac fate determination may lead to better approaches in promoting heart regeneration. We used a mesoderm posterior 1 (Mesp1)-Cre/Rosa26-EYFP reporter system to identify microRNAs (miRNAs) enriched in early cardiac progenitor cells. Most of these miRNA genes bear MESP1-binding sites and active histone signatures. In a calcium transient-based screening assay, we identified miRNAs that may promote the cardiomyocyte program. An X-chromosome miRNA cluster, miR-322/-503, is the most enriched in the Mesp1 lineage and is the most potent in the screening assay. It is specifically expressed in the looping heart. Ectopic miR-322/-503 mimicking the endogenous temporal patterns specifically drives a cardiomyocyte program while inhibiting neural lineages, likely by targeting the RNA-binding protein CUG-binding protein Elav-like family member 1 (Celf1). Thus, early miRNAs in lineage-committed cells may play powerful roles in cell-fate determination by cross-suppressing other lineages. miRNAs identified in this study, especially miR-322/-503, are potent regulators of early cardiac fate.

CUG-binding protein, ELAV-like Family Member 1 (CELF1) plays an important role during the development of different tissues, such as striated muscle and brain tissue. CELF1 is an RNA-binding protein that regulates RNA metabolism processes, e.g., alternative splicing, and antagonizes other RNA-binding proteins, such as Muscleblind -like proteins (MBNLs). Abnormal activity of both classes of proteins plays a crucial role in the pathogenesis of myotonic dystrophy type 1 (DM1), the most common form of muscular dystrophy in adults. In this work, we show that alternative splicing of exons forming both the 5′ and 3′ untranslated regions (UTRs) of CELF1 mRNA is efficiently regulated during development and tissue differentiation and is disrupted in skeletal muscles in the context of DM1. Alternative splicing of the CELF1 5′UTR leads to translation of two potential protein isoforms that differ in the lengths of their N-terminal domains. We also show that the MBNL and CELF proteins regulate the distribution of mRNA splicing isoforms with different 5′UTRs and 3′UTRs and affect the CELF1 expression by changing its sensitivity to specific microRNAs or RNA-binding proteins. Together, our findings show the existence of different mechanisms of regulation of CELF1 expression through the distribution of various 5′ and 3′ UTR isoforms within CELF1 mRNA.

Background/Objectives: The RNA-binding protein CELF1 is crucial for cardiac development, but its role in cardiomyocyte hypertrophy is unclear. This study investigates the effects of acute CELF1 knockdown on alternative splicing and hypertrophic growth in cardiomyocytes. Methods: Neonatal rat cardiomyocytes (NRCMs) were transfected with two siRNAs targeting CELF1. Hypertrophy was assessed by cell size and expression of hypertrophic markers via qPCR and Western blot. RNA sequencing was performed in NRCMs to identify alternative splicing events. Tead1 function was tested by knockdown in NRCMs. Selected mechanistic assays were performed primarily in HeLa cells. Results: CELF1 knockdown in NRCMs increased cardiomyocyte size and upregulated hypertrophic markers, while its overexpression restored the phenotype. RNA-seq revealed that CELF1 knockdown alters the alternative splicing pattern. Specifically, the splicing of the transcription factor Tead1 shifted from the full-length long Tead1 isoform (Tead1-L) to the exon 4-skipped short isoform (Tead1-S). In HeLa cells, CELF1 interacted with hnRNPC, an m6A reader and splicing factor, and CELF1 perturbation correlated with changes in global m6A abundance. Conclusions: These findings suggest that CELF1 regulates hypertrophic phenotypes in cardiomyocytes and is associated with alternative splicing of Tead1.

The importance of translational regulation in tumour biology is increasingly appreciated. Here, we leverage polyribosomal profiling to prospectively define translational regulatory programs underlying epithelial-to-mesenchymal transition (EMT) in breast epithelial cells. We identify a group of ten translationally regulated drivers of EMT sharing a common GU-rich cis- element within the 3′-untranslated region (3′-UTR) of their mRNA. These cis -elements, necessary for the regulatory activity imparted by these 3′-UTRs, are directly bound by the CELF1 protein, which itself is regulated post-translationally during the EMT program. CELF1 is necessary and sufficient for both mesenchymal transition and metastatic colonization, and CELF1 protein, but not mRNA, is significantly overexpressed in human breast cancer tissues. Our data present an 11-component genetic pathway, invisible to transcriptional profiling approaches, in which the CELF1 protein functions as a central node controlling translational activation of genes driving EMT and ultimately tumour progression. Epithelial-to-mesenchymal transition is a key process in tumorigenesis but little is known about the molecular mechanism regulating such process at the translational level. Here, the authors identify a subset of mRNAs important for this process that are specifically modulated by the RNA-binding protein CELF1.

Background RNA binding proteins (RBPs)-mediated regulation plays important roles in many eye diseases, including the canonical RBP CELF1 in cataract. While the definite molecular regulatory mechanisms of CELF1 on cataract still remain elusive. Methods In this study, we overexpressed CELF1 in human cultured lens epithelial SRA01/04 cells and applied whole transcriptome sequencing (RNA-seq) method to analyze the global differences mediated by CELF1. We then analyzed public RNA-seq and CELF1-RNA interactome data to decipher the underlying mechanisms. Results The results showed that transcriptome profile was globally changed by CELF1 overexpression (CELF1-OE). Functional analysis revealed CELF1 specifically increased the expression of genes in extracellular matrix disassembly, extracellular matrix organization, and proteolysis, which could be classified into matrix metalloproteinases (MMPs) family. This finding was also validated by RT-qPCR and public mouse early embryonic lens data. Integrating analysis with public CELF1-RNA interactome data revealed that no obvious CELF1-binding peak was found on the transcripts of these genes, indicating an indirectly regulatory role of CELF1 in lens epithelial cells. Conclusions Our study demonstrated that CELF1-OE promotes transcriptional level of MMP genes; and this regulation may be completed by other ways except for binding to RNA targets. These results suggest that CELF1-OE is implicated in the development of lens, which is associated with cataract and expands our understanding of CELF1 regulatory roles as an RNA binding protein.