The causal link between long terminal repeat (LTR) retrotransposon-derived lncRNAs and hepatocellular carcinoma (HCC) remains elusive and whether these cancer-exclusive lncRNAs contribute to the effectiveness of current HCC therapies is yet to explore. Here, we investigated the activation of LTR retrotransposon-derived lncRNAs in a broad range of liver diseases. We found that LTR retrotransposon-derived lncRNAs are mainly activated in HCC and is correlated with the proliferation status of HCC. Furthermore, we discovered that an LTR retrotransposon-derived lncRNA, LINC01446, exhibits specific expression in HCC. HCC patients with higher LINC01446 expression had shorter overall survival times. In vitro and in vivo assays showed that LINC01446 promoted HCC growth and angiogenesis. Mechanistically, LINC01446 bound to serine/arginine protein kinase 2 (SRPK2) and activated its downstream target, serine/arginine splicing factor 1 (SRSF1). Furthermore, activation of the SRPK2-SRSF1 axis increased the splicing and expression of VEGF isoform A165 (VEGFA165). Notably, inhibiting LINC01446 expression dramatically impaired tumor growth in vivo and resulted in better therapeutic outcomes when combined with antiangiogenic agents. In addition, we found that the transcription factor MESI2 bound to the cryptic MLT2B3 LTR promoter and drove LINC01446 transcription in HCC cells. Taken together, our findings demonstrate that LTR retrotransposon-derived LINC01446 promotes the progression of HCC by activating the SRPK2/SRSF1/VEGFA165 axis and highlight targeting LINC01446 as a potential therapeutic strategy for HCC patients.

Background Poly (ADP-ribose) polymerase inhibitor (PARPi) resistance poses a significant challenge in ovarian carcinoma (OC). While the role of DOT1L in cancer and chemoresistance is acknowledged, its specific role in PARPi resistance remains unclear. This study aims to elucidate the molecular mechanism of DOT1L in PARPi resistance in OC patients.Methods This study analyzed the expression of DOT1L in PARPi-resistant cell lines compared to sensitive ones and correlated it with clinical outcomes in OC patients. Comprehensive in vitro and in vivo functional experiments were conducted using cellular and mouse models. Molecular investigations, including RNA sequencing, chromatin immunoprecipitation (ChIP) and Cleavage Under Targets and Tagmentation (CUT&Tag) assays, were employed to unravel the molecular mechanisms of DOT1L-mediated PARPi resistance.Results Our investigation revealed a robust correlation between DOT1L expression and clinical PARPi resistance in non-BRCA mutated OC cells. Upregulated DOT1L expression in PARPi-resistant tissues was associated with diminished survival in OC patients. Mechanistically, we identified that PARP1 directly binds to the DOT1L gene promoter, promoting transcription independently of its enzyme activity. PARP1 trapping induced by PARPi treatment amplified this binding, enhancing DOT1L transcription and contributing to drug resistance. Sequencing analysis revealed that DOT1L plays a crucial role in the transcriptional regulation of PLCG2 and ABCB1 via H3K79me2. This established the PARP1-DOT1L-PLCG2/ABCB1 axis as a key contributor to PARPi resistance. Furthermore, we discovered that combining a DOT1L inhibitor with PARPi demonstrated a synergistic effect in both cell line-derived xenograft mouse models (CDXs) and patient-derived organoids (PDOs).Conclusions Our results demonstrate that DOT1L is an independent prognostic marker for OC patients. The PARP1-DOT1L/H3K79me2-PLCG2/ABCB1 axis is identified as a pivotal contributor to PARPi resistance. Targeted inhibition of DOT1L emerges as a promising therapeutic strategy for enhancing PARPi treatment outcomes in OC patients.

Vascular smooth muscle cells (VSMCs) are the major contributor to vascular repair and remodeling, which showed high level of phenotypic plasticity. Abnormalities in VSMC plasticity can lead to multiple cardiovascular diseases, wherein alternative splicing plays important roles. However, alternative splicing variants in VSMC plasticity are not fully understood. Here we systematically characterized the long-read transcriptome and their dysregulation in human aortic smooth muscle cells (HASMCs) by employing the Oxford Nanopore Technologies long-read RNA sequencing in HASMCs that are separately treated with platelet-derived growth factor, transforming growth factor, and hsa-miR-221-3P transfection. Our analysis reveals frequent alternative splicing events and thousands of unannotated transcripts generated from alternative splicing. HASMCs treated with different factors exhibit distinct transcriptional reprogramming modulated by alternative splicing. We also found that unannotated transcripts produce different open reading frames compared to the annotated transcripts. Finally, we experimentally validated the unannotated transcript derived from gene CISD1 , namely CISD1-u , which plays a role in the phenotypic switch of HASMCs. Our study characterizes the phenotypic modulation of HASMCs from an insight of long-read transcriptome, which would promote the understanding and the manipulation of HASMC plasticity in cardiovascular diseases.

Differentiated expressed genes after lncMER52A knockdown.

The list of 464 HCC-specific lncRNAs.

<p>Supplementary data-The Supplementary information includes: Figure S1. The genomic location, subcellular distribution, full length and coding potential of lncMER52A. Figure S2. The knockdown efficiency and specificity of lncMER52A siRNAs and Crispr-Cas9 mediated knockdown. The effects of lncMER52A on the migration, invasion and metastasis of HCC cells. Figure S3. The correlation of lncMER52A and p120-catenin in HCC tissues and paired normal livers. Figure S4. lncMER52A overexpression significantly inhibited the interaction between β-TrCP1 and p120-catenin. Figure S5. The effects of p120-catenin on the migration and invasion of HCC cells in vitro. Figure S6. H3K4me3 and H3K27ac levels at the promoter of lncMER52A in Huh7 and HepG2-C3A cells. Table S1. Primers for RACE assays. Table S2. Sequence of siRNAs and gRNAs. Table S3. Primers used in molecular cloning of vectors. Table S4. Primers used in the RNA pulldown assays. Table S5. Primers used in the northern blot assays. Table S6. Primers used in the qPCR experiments. Table S7. Primers used in the ChIP assays. Table S8. The correlation of lncMER52A expression with HCC. clinicopathological features.</p>

Background Increasing evidences has indicated that primary and acquired resistance of ovarian cancer (OC) to platinum is mediated by multiple molecular and cellular factors. Understanding these mechanisms could promote the therapeutic efficiency for patients with OC. Methods Here, we screened the expression pattern of circRNAs in samples derived from platinum-resistant and platinum-sensitive OC patients using RNA-sequencing (RNA-seq). The expression of hsa_circ_0010467 was validated by Sanger sequencing, RT-qPCR, and fluorescence in situ hybridization (FISH) assays. Overexpression and knockdown experiments were performed to explore the function of hsa_circ_0010467. The effects of hsa_circ_0010467 on enhancing platinum treatment were validated in OC cells, mouse model and patient-derived organoid (PDO). RNA pull-down, RNA immunoprecipitation (RIP), and dual-luciferase reporter assays were performed to investigate the interaction between hsa_circ_0010467 and proteins. Results Increased expression of hsa_circ_0010467 is observed in platinum-resistant OC cells, tissues and serum exosomes, which is positively correlated with advanced tumor stage and poor prognosis of OC patients. Hsa_circ_0010467 is found to maintain the platinum resistance via inducing tumor cell stemness, and silencing hsa_circ_0010467 substantially increases the efficacy of platinum treatment on inhibiting OC cell proliferation. Further investigation reveals that hsa_circ_0010467 acts as a miR-637 sponge to mediate the repressive effect of miR-637 on leukemia inhibitory factor (LIF) and activates the LIF/STAT3 signaling pathway. We further discover that AUF1 could promote the biogenesis of hsa_circ_0010467 in OC. Conclusion Our study uncovers the mechanism that hsa_circ_0010467 mediates the platinum resistance of OC through AUF1/hsa_circ_0010467/miR-637/LIF/STAT3 axis, and provides potential targets for the treatment of platinum-resistant OC patients.

Full scan of all the western blots in the main figures.

<p>Supplementary data-The Supplementary information includes: Figure S1. The genomic location, subcellular distribution, full length and coding potential of lncMER52A. Figure S2. The knockdown efficiency and specificity of lncMER52A siRNAs and Crispr-Cas9 mediated knockdown. The effects of lncMER52A on the migration, invasion and metastasis of HCC cells. Figure S3. The correlation of lncMER52A and p120-catenin in HCC tissues and paired normal livers. Figure S4. lncMER52A overexpression significantly inhibited the interaction between β-TrCP1 and p120-catenin. Figure S5. The effects of p120-catenin on the migration and invasion of HCC cells in vitro. Figure S6. H3K4me3 and H3K27ac levels at the promoter of lncMER52A in Huh7 and HepG2-C3A cells. Table S1. Primers for RACE assays. Table S2. Sequence of siRNAs and gRNAs. Table S3. Primers used in molecular cloning of vectors. Table S4. Primers used in the RNA pulldown assays. Table S5. Primers used in the northern blot assays. Table S6. Primers used in the qPCR experiments. Table S7. Primers used in the ChIP assays. Table S8. The correlation of lncMER52A expression with HCC. clinicopathological features.</p>

Long noncoding RNAs (lncRNAs) are a type of transcript that is >200 nucleotides long with no protein-coding capacity. Accumulating studies have suggested that lncRNAs contain open reading frames (ORFs) that encode peptides. Although several noncoding RNA-encoded peptide-related databases have been developed, most of them display only a small number of experimentally validated peptides, and resources focused on lncRNA-encoded peptides are still lacking. We used six types of evidence, coding potential assessment tool (CPAT), coding potential calculator v2.0 (CPC2), N6-methyladenosine modification of RNA sites (m6A), Pfam, ribosome profiling (Ribo-seq), and translation initiation sites (TISs), to evaluate the coding potential of 883,804 lncRNAs across 39 species. We constructed a comprehensive database of lncRNA-encoded peptides, LncPep (http://www.shenglilabs.com/LncPep/). LncPep provides three major functional modules: 1) user-friendly searching/browsing interface, 2) prediction and BLAST modules for exploring novel lncRNAs and peptides, and 3) annotations for lncRNAs, peptides and supporting evidence. Taken together, LncPep is a user-friendly and convenient platform for discovering and investigating peptides encoded by lncRNAs.

Transcribed RNAs undergo various regulation and modification to become functional transcripts. Notably, cancer transcriptome has not been fully characterized at transcript resolution. Herein, we carry out a reference-based transcript assembly across >1000 cancer cell lines. We identify 498,255 transcripts, approximately half of which are unannotated. Unannotated transcripts are closely associated with cancer-related hallmarks and show clinical significance. We build a high-confidence RNA binding protein (RBP)-transcript regulatory network, wherein most RBPs tend to regulate transcripts involved in cell proliferation. We identify numerous transcripts that are highly associated with anti-cancer drug sensitivity. Furthermore, we establish RBP-transcript-drug axes, wherein PTBP1 is experimentally validated to affect the sensitivity to decitabine by regulating KIAA1522-a6 transcript. Finally, we establish a user-friendly data portal to serve as a valuable resource for understanding cancer transcriptome diversity and its potential clinical utility at transcript level. Our study substantially extends cancer RNA repository and will facilitate anti-cancer drug discovery.

Pancreatic cancer is a highly aggressive cancer with an exceedingly low rate of response to treatments, which calls for comprehensive molecular characterization of pancreatic cancer cell lines (PCCLs). We screened multi-layer molecular data of 36 PCCLs, including gene mutation, gene expression, microRNA (miRNA) expression, and protein profiles. Our comparative analysis of genomic mutations found that PCCLs recapitulated genomic alterations of the primary tumor and suggested potential therapeutic strategies for clinical interventions. The panel of 36 PCCLs was classified into 2 subgroups based on transcriptomic mRNA expression, wherein the C1 subgroup was characterized with differentiation, whereas C2 cell lines were featured with immunity, angiogenesis, epidermis, and proliferation. Transcriptomic classification was further recapitulated by miRNA and protein expression. Additionally, the differential proteins between C1 and C2 subgroups were prominently involved in epidermal growth factor receptor (EGFR) signaling, phosphatidylinositol 3-kinase (PI3K) signaling, and mitogen-activated protein kinase (MAPK) signaling pathways. Tumor samples from different subgroups exhibited distinct infiltration of CD4 naive cells and monocytes. Remarkably, patients in subgroups C1 showed longer survival, whereas those in C2 had worse clinical outcome. Further integrative analysis revealed that temozolomide and NVP-TAE684 showed higher sensitivity in the C1 subgroup, whereas the C2 cell lines were more sensitive to SR1001 and SRT-1720. Our results also showed that PCCLs with mutations in CDKN2A, TP53, and SMAD4 were more sensitive to certain anti-cancer drugs. Our integrative analysis identified molecular features of pancreatic cancer that were associated with clinical significance and drug sensitivity, providing potentially effective strategies for precision treatments of patients with pancreatic cancer.

Inflammatory skin diseases are induced by disorders of the host defense system of the skin, which is composed of a barrier, innate and acquired immunity, as well as the cutaneous microbiome. These disorders are characterized by recurrent cutaneous lesions and intense itch, which seriously affecting life quality of people across all ages and ethnicities. To elucidate molecular factors for typical inflammatory skin diseases (such as psoriasis and atopic dermatitis), transcriptomic profiling assays have been largely performed. Additionally, single-cell RNA sequencing (scRNA-seq) as well as spatial transcriptomic profiling have revealed multiple potential translational targets and offered guides to improve diagnosis and treatment strategies for inflammatory skin diseases. High-throughput transcriptomics data has shown unprecedented power to disclose the complex pathophysiology of inflammatory skin diseases. Here, we will summarize discoveries from transcriptomics data and discuss how to maximize the transcriptomics data to propel the development of diagnostic biomarkers and therapeutic targets in inflammatory skin diseases.

PTEN is a crucial tumor suppressor and loss of PTEN protein is involved in various cancers. However, the detailed molecular mechanisms of PTEN loss in cancers remain elusive, especially the involvement of lncRNAs. Here, lncRNA RP11-295G20.2 is found to be significantly upregulated in hepatocellular carcinoma (HCC) and promotes the growth of liver cancer cells both in vitro and in vivo. Furthermore, RP11-295G20.2 inhibits autophagy in liver cancer cells. Interestingly, RP11-295G20.2 directly binds to the PTEN protein and leads to its degradation. RP11-295G20.2 expression is inversely correlated with PTEN protein expression in 82 TCGA/TCPA-LIHC samples. Surprisingly, RP11-295G20.2-induced PTEN degradation occurs through the lysosomal pathway instead of the proteasome pathway. RP11-295G20.2 binds to the N terminus of PTEN and facilitates the interaction of p62 with PTEN. Thus, PTEN is translocated into lysosomes and degraded. RP11-295G20.2 also influences AKT phosphorylation and forkhead box O 3a (FOXO3a) translocation into the nucleus, in turn regulating the transcription of autophagy-related genes. Collectively, RP11-295G20.2 directly binds to PTEN and enables its lysosomal degradation. This newly identified RP11-295G20.2/PTEN axis reveals an unexplored molecular mechanism regarding PTEN loss in liver cancer and might provide new therapeutic benefits for liver cancer patients.

Retrotransposons have been known as major sources of genomic instability that trigger and promote cancer development. In a recent study, Gu et al. revealed that MPP8 deficiency impeded tumor development through reactivation long interspersed element-1 (LINE-1 or L1) retrotransposons in acute myeloid leukemia (AML), suggesting a tumor-suppressive role for retrotransposons.

Epigenetic alteration is one of the hallmarks of colorectal cancer (CRC). Many driver genes are regulated by DNA methylation in CRC. However, the role of DNA methylation regulating lncRNAs remain elusive. Here, we identify that SNHG11 (small nucleolar RNA host gene 11) is upregulated by promotor hypomethylation in CRC and is associated with poor prognosis in CRC patients. SNHG11 can promote CRC cell migration and metastasis under hypoxia. Interestingly, the DNA-binding motif of SNHG11 is similar to that of HIF-1α. In addition, SNHG11-associated genes are enriched with members of the HIF-1 signaling pathway in CRC. Mechanistically, SNHG11 binds to the pVHLrecognition sites on HIF-1α, thus blocking the interaction of pVHL with HIF-1α and preventing its ubiquitination and degradation. Moreover, SNHG11 upregulates the expression of HIF-1α target genes, i.e., AK4, ENO1, HK2, and Twist1. Notably, SNHG11 can bind to the HRE sites in the promoter of these genes and increase their transcription. In summary, these results identify a SNHG11/ HIF-1α axis that plays a pivotal role in tumor invasion and metastasis.

BACKGROUND AND AIMS:Alternative splicing (AS) is a key step that increases the diversity and complexity of the cancer transcriptome. Recent evidence has highlighted that AS has an increasingly crucial role in cancer. Nonetheless, the mechanisms underlying AS and its dysregulation in hepatocellular carcinoma (HCC) remain elusive. Here, we report that the expression of RNA-binding protein p54nrb /non-POU domain-containing octamer-binding protein (NONO) is frequently increased in patients with HCC and is associated with poor outcomes.APPROACH AND RESULTS:Knockdown of NONO significantly abolished liver cancer cell proliferation, migration, and tumor formation. RNA-sequencing revealed that NONO regulates MYC box-dependent interacting protein 1 (or bridging integrator 1 [BIN1]; also known as amphiphysin 2 3P9) exon 12a splicing. In the normal liver, BIN1 generates a short isoform (BIN1-S) that acts as a tumor suppressor by inhibiting the binding of c-Myc to target gene promoters. In HCC, NONO is highly up-regulated and produces a long isoform (BIN1-L, which contains exon 12a) instead of BIN1-S. High levels of BIN1-L promote carcinogenesis by binding with the protein polo-like kinase 1 to enhance its stability through the prevention of ubiquitin/proteasome-dependent cullin 3 degradation. Further analysis revealed that NONO promotes BIN1 exon 12a inclusion through interaction with DExH-box helicase 9 (DHX9) and splicing factor proline and glutamine-rich (SFPQ). Notably, frequent coexpression of DHX9-NONO-SFPQ is observed in patients with HCC.CONCLUSIONS:Taken together, our findings identify the DHX9-NONO-SFPQ complex as a key regulator manipulating the oncogenic splicing switch of BIN1 and as a candidate therapeutic target in liver cancer.

Background Hypoxic tumors are refractory to DNA damage drugs. However, the underlying mechanism has yet to be elucidated. We aimed to identify lncRNAs that upregulated under hypoxia and their effects on colorectal cancer (CRC). Methods CRC cells were treated with 1% O-2 to identify lncRNAs that upregulated under hypoxia. We integrated these lncRNAs with RNA-seq of 4 paired CRC tissues and TCGA data to get candidate lncRNAs. Multiple in vitro and in vivo assays were used to explore the role of LUCAT1 in CRC. Results We identified a hypoxia-induced lncRNA LUCAT1 that facilitated the growth of CRC cells and contributed to drug resistance of CRC cells both in vitro and in vivo. Mechanically, LUCAT1 interacts with polypyrimidine tract binding protein 1 (PTBP1) in CRC cells, facilitates the association of a set of DNA damage related genes with PTBP1, thus resulting in altered alternative splicing of these genes. Moreover, ectopic expression of PTBP1 in CRC cells with knockdown of LUCAT1 abrogated the effects induced by LUCAT1 knockdown. Chemotherapeutics drug combined with LUCAT1 knockdown via antisense oligonucleotides (ASO) would get a better outcome in vivo, compared with group treated with chemotherapeutic drug only. Notably, LUCAT1 is upregulated in CRC tissues, compared to adjacent normal tissues; and CRC patients with higher LUCAT1 have a worse prognosis and poorly responded to chemotherapy in the clinic. Conclusions Our data suggested CRC cells utilizes LUCAT1 to develop resistance to DNA damage drugs, and disrupting the LUCAT1/PTBP1 axis might be a promising therapeutic strategy for refractory hypoxic tumors.