Precursor messenger RNA (pre-mRNA) splicing is catalyzed by an intricate ribonucleoprotein complex called the spliceosome. Although the spliceosome is considered to be general cell "housekeeping" machinery, mutations in core components of the spliceosome frequently correlate with cell- or tissue-specific phenotypes and diseases. In this review, we expound the links between spliceosome mutations, aberrant splicing, and human cancers. Remarkably, spliceosome-targeted therapies (STTs) have become efficient anti-cancer strategies for cancer patients with splicing defects. We also highlight the links between spliceosome and immune signaling. Recent studies have shown that some spliceosome gene mutations can result in immune dysregulation and notable phenotypes due to mis-splicing of immune-related genes. Furthermore, several core spliceosome components harbor splicing-independent immune functions within the cell, expanding the functional repertoire of these diverse proteins.
Humans ; Neoplasms/metabolism* ; RNA Precursors/metabolism* ; RNA Splicing ; RNA Splicing Factors/metabolism* ; Spliceosomes/metabolism*

A complete proteomics study characterizing active androgen receptor (AR) complexes in prostate cancer (PCa) cells identified a diversity of protein interactors with tumorigenic annotations, including known RNA splicing factors. Thus, we chose to further investigate the functional role of AR-mediated alternative RNA splicing in PCa disease progression. We selected two AR-interacting RNA splicing factors, Src associated in mitosis of 68 kDa (SAM68) and DEAD (Asp-Glu-Ala-Asp) box helicase 5 (DDX5) to examine their associative roles in AR-dependent alternative RNA splicing. To assess the true physiological role of AR in alternative RNA splicing, we assessed splicing profiles of LNCaP PCa cells using exon microarrays and correlated the results to PCa clinical datasets. As a result, we were able to highlight alternative splicing events of clinical significance. Initial use of exon-mini gene cassettes illustrated hormone-dependent AR-mediated exon-inclusion splicing events with SAM68 or exon-exclusion splicing events with DDX5 overexpression. The physiological significance in PCa was investigated through the application of clinical exon array analysis, where we identified exon-gene sets that were able to delineate aggressive disease progression profiles and predict patient disease-free outcomes independently of pathological clinical criteria. Using a clinical dataset with patients categorized as prostate cancer-specific death (PCSD), these exon gene sets further identified a select group of patients with extremely poor disease-free outcomes. Overall, these results strongly suggest a nonclassical role of AR in mediating robust alternative RNA splicing in PCa. Moreover, AR-mediated alternative spicing contributes to aggressive PCa progression, where we identified a new subtype of lethal PCa defined by AR-dependent alternative splicing.
Humans ; Male ; Alternative Splicing ; Cell Line, Tumor ; DEAD-box RNA Helicases/metabolism* ; Disease Progression ; Gene Expression Regulation, Neoplastic ; Prostatic Neoplasms/pathology* ; Receptors, Androgen/metabolism* ; RNA Splicing Factors/metabolism*

OBJECTIVE: To construct an adenovirus vector expressing artificial splicing factor capable of regulating alternative splicing of Yap1 in cardiomyocytes. METHODS: The splicing factors with different sequences were constructed against Exon6 of YAP1 based on the sequence specificity of Pumilio1. The PCR fragment of the artificially synthesized PUF-SR or wild-type PUFSR was cloned into pAd-Track plasmid, and the recombinant plasmids were transformed into E. coli DH5α for plasmid amplification. The amplified plasmids were digested with Pac I and transfected into 293A cells for packaging to obtain the adenovirus vectors. Cultured neonatal rat cardiomyocytes were transfected with the adenoviral vectors, and alternative splicing of YAP1 was detected using quantitative and semi-quantitative PCR; Western blotting was performed to detect the signal of the fusion protein Flag. RESULTS: The transfection efficiency of the adenovirus vectors was close to 100% in rat cardiomyocytes, and no fluorescent protein was detected in the cells with plasmid transfection. The results of Western blotting showed that both the negative control and Flag-SR-NLS-PUF targeting the YAPExon6XULIE sequence were capable of detecting the expression of the protein fused to Flag. The results of reverse transcription-PCR and PCR demonstrated that the artificial splicing factor constructed based on the 4th target sequence of YAP1 effectively regulated the splicing of YAP1 Exon6 in the cardiomyocytes (P < 0.05). CONCLUSION We successfully constructed adenovirus vectors capable of regulating YAP1 alternative splicing rat cardiomyocytes.
Adenoviridae/metabolism* ; Alternative Splicing ; Animals ; Animals, Newborn ; Escherichia coli/metabolism* ; Genetic Vectors ; Myocytes, Cardiac/metabolism* ; Plasmids ; RNA Splicing Factors/metabolism* ; Rats ; Transfection

Long noncoding RNAs (lncRNAs) play a critical role in the regulation of atherosclerosis. Here, we investigated the role of the lncRNA growth arrest-specific 5 (lncR-GAS5) in atherogenesis. We found that the enforced expression of lncR-GAS5 contributed to the development of atherosclerosis, which presented as increased plaque size and reduced collagen content. Moreover, impaired autophagy was observed, as shown by a decreased LC3II/LC3I protein ratio and an elevated P62 level in lncR-GAS5-overexpressing human aortic endothelial cells. By contrast, lncR-GAS5 knockdown promoted autophagy. Moreover, serine/arginine-rich splicing factor 10 (SRSF10) knockdown increased the LC3II/LC3I ratio and decreased the P62 level, thus enhancing the formation of autophagic vacuoles, autolysosomes, and autophagosomes. Mechanistically, lncR-GAS5 regulated the downstream splicing factor SRSF10 to impair autophagy in the endothelium, which was reversed by the knockdown of SRSF10. Further results revealed that overexpression of the lncR-GAS5-targeted gene miR-193-5p promoted autophagy and autophagic vacuole accumulation by repressing its direct target gene, SRSF10. Notably, miR-193-5p overexpression decreased plaque size and increased collagen content. Altogether, these findings demonstrate that lncR-GAS5 partially contributes to atherogenesis and plaque instability by impairing endothelial autophagy. In conclusion, lncR-GAS5 overexpression arrested endothelial autophagy through the miR-193-5p/SRSF10 signaling pathway. Thus, miR-193-5p/SRSF10 may serve as a novel treatment target for atherosclerosis.
Humans ; Atherosclerosis/genetics* ; Autophagy/genetics* ; Cell Cycle Proteins/metabolism* ; Endothelial Cells/metabolism* ; Endothelium/metabolism* ; MicroRNAs/metabolism* ; Repressor Proteins/metabolism* ; RNA Splicing Factors ; Serine-Arginine Splicing Factors/genetics* ; RNA, Long Noncoding/metabolism*

OBJECTIVE: To observe the means of fibronectin(FN) alternative splicing and the expression of EIIIA-FN variant in rat skin after bruise, for the sake of providing some help for forensic estimation of wound interval. METHODS: Total RNA was isolated from wounded skin, and reverse transcription polymerase chain reaction was conducted to amplify target segments. RESULTS: Detectable EIIIA+(526 bp) segments, lacked in normal organize, was amplified at 1 h after experimental wound, and the levels were increased within 24 h. CONCLUSION The alternative splicing EIIIA-fibronectin variant would be a satisfied criterion for research of skin injury.
Alternative Splicing ; Animals ; Epithelium/metabolism* ; Fibronectins/genetics* ; Forensic Medicine ; Integrin alpha4beta1/biosynthesis* ; RNA, Messenger/metabolism* ; Rats ; Reverse Transcriptase Polymerase Chain Reaction ; Skin/metabolism* ; Time Factors

OBJECTIVETo investigate the expression of neural salient serine/arginine-rich protein 1 (NSSR1) in colorectal cancer.

METHODSRT-PCR, Western blot and immunohistochemical staining were used to detect the expression of NSSR1 mRNA and protein in different mouse tissues and human colorectal cancer, respectively.

RESULTSNSSR1 mRNA was expressed in mouse cerebrum, cerebellum, heart, liver, intestine, kidney and lung tissue, but NSSR1 protein was only expressed in neural tissues. In normal human intestinal mucosa, NSSR1 was expressed in the colorectal epithelial cells. In colorectal cancer, NSSR1 was highly expressed in the nucleus of tumor cells.

CONCLUSIONThe extensive expression of NSSR1 in colorectal cancer cells may hint it's roles in the biological function of colorectal cancer.

Animals ; Cell Cycle Proteins ; genetics ; metabolism ; Colon ; metabolism ; Colorectal Neoplasms ; metabolism ; Humans ; Mice ; Neoplasm Proteins ; genetics ; metabolism ; RNA, Messenger ; genetics ; RNA-Binding Proteins ; genetics ; metabolism ; Rectum ; metabolism ; Repressor Proteins ; genetics ; metabolism ; Serine-Arginine Splicing Factors

Enhancer of rudimentary homolog (ERH) is a small, highly conserved protein among eukaryotes. Since its discovery nearly 20 years ago, its molecular function has remained enigmatic. It has been implicated to play a role in transcriptional regulation and in cell cycle. We recently showed that ERH binds to the Sm complex and is required for the mRNA splicing of the mitotic motor protein CENP-E. Furthermore, cancer cells driven by mutations in the KRAS oncogene are particularly sensitive to RNAi-mediated suppression of ERH function, and ERH expression is inversely correlated with survival in colorectal cancer patients whose tumors harbor KRAS mutation. These recent findings indicate that ERH plays an important role in cell cycle through its mRNA splicing activity and is critically required for genomic stability and cancer cell survival.
Amino Acid Sequence ; Animals ; Cell Cycle ; Evolution, Molecular ; Humans ; Molecular Sequence Data ; Neoplasms ; metabolism ; RNA Splicing ; Transcription Factors ; chemistry ; metabolism ; Transcription, Genetic

OBJECTIVETo identify the interaction partners of a new splicing product of LMO2 gene (LMO2-C), and study its function in K562 cells.

METHODSMaltose binding protein (MBP) pull down and mammalian two-hybrid assay (MTHA) were used to identify the interaction partners of LMO2-C in K562 cells. Semiquantitative RT-PCR was used to detect the expression of hematopoietic specific gene glycoprotein (GPA) in K562 cells.

RESULTSMBP-LMO2-C fusion protein was expressed and purified in soluble form successfully. Endogenous GATA1 and LDB1 proteins were confirmed to bind to LMO2-C by MBP pull down analysis. The MTHA also showed that LMO2-C had comparable binding affinities to LDB1 with LMO2-L, and over expression of LMO2-C prevented LMO2-L from binding to LDB1, the inhibition rate being (81.13 +/- 0.68)%. RT-PCR results showed that the expression level of GPA was reduced [(51.00 +/- 1.58)%] in K562 cells while LMO2-C overexpressed.

CONCLUSIONLMO2-C can bind endogenous GATA1 and LDB1 protein in K562 cells and down regulates the expression of GPA.

Adaptor Proteins, Signal Transducing ; DNA-Binding Proteins ; genetics ; metabolism ; GATA1 Transcription Factor ; metabolism ; Humans ; K562 Cells ; LIM Domain Proteins ; Maltose-Binding Proteins ; Metalloproteins ; genetics ; metabolism ; Periplasmic Binding Proteins ; Proto-Oncogene Proteins ; RNA Splicing ; Transcription Factors ; metabolism ; Two-Hybrid System Techniques

Natural antisense transcripts (NAT) and alternative polyadenylation (APA) of messenger RNA (mRNA) are important contributors of transcriptome complexity, each playing a critical role in multiple biological processes. However, whether they have crosstalk and function collaboratively is unclear. We discovered that APA enriched in human sense-antisense (S-AS) gene pairs, and finally focused on RNASEH2C-KAT5 S-AS pair for further study. In cis but not in trans over-expression of the antisense KAT5 gene promoted the usage of distal polyA (pA) site in sense gene RNASEH2C, which generated longer 3' untranslated region (3'UTR) and produced less protein, accompanying with slowed cell growth. Mechanistically, elevated Pol II occupancy coupled with SRSF3 could explain the higher usage of distal pA site. Finally, NAT-mediated downregulation of sense gene's protein level in RNASEH2C-KAT5 pair was specific for human rather than mouse, which lacks the distal pA site of RNASEH2C. We provided the first evidence to support that certain gene affected phenotype may not by the protein of its own, but by affecting the expression of its overlapped gene through APA, implying an unexpected view for understanding the link between genotype and phenotype.
Cell Proliferation ; genetics ; Evolution, Molecular ; Gene Expression Regulation ; genetics ; HEK293 Cells ; Humans ; Polyadenylation ; genetics ; RNA, Antisense ; genetics ; RNA, Messenger ; genetics ; Ribonuclease H ; genetics ; Serine-Arginine Splicing Factors ; metabolism ; Transcription, Genetic ; Up-Regulation ; genetics

Animals ; Autophagy/genetics* ; Cholesterol/metabolism* ; Gene Expression Regulation ; Integrases/metabolism* ; Leydig Cells/metabolism* ; Male ; Mice, Knockout ; Multienzyme Complexes/metabolism* ; Phosphoproteins/metabolism* ; Primary Cell Culture ; Progesterone Reductase/metabolism* ; RNA Splicing Factors/metabolism* ; Scavenger Receptors, Class B/metabolism* ; Sequestosome-1 Protein/metabolism* ; Signal Transduction ; Sirtuin 1/genetics* ; Sodium-Hydrogen Exchangers/metabolism* ; Steroid 17-alpha-Hydroxylase/metabolism* ; Steroid Isomerases/metabolism* ; Testosterone/genetics*