ABSTRACT
Small ubiquitin-related modifier(SUMOylation)is a dynamic post-translational modification that maintains cardiac function and can protect against a hypertrophic response to cardiac pressure overload.However,the function of SUMOylation after myocardial infarction(MI)and the molecular details of heart cell responses to SUMO1 deficiency have not been determined.In this study,we demonstrated that SUMO1 protein was inconsistently abundant in different cell types and heart regions after MI.However,SUMO1 knockout significantly exacerbated systolic dysfunction and infarct size after myocardial injury.Single-nucleus RNA sequencing revealed the differential role of SUMO1 in regulating heart cells.Among cardiomyocytes,SUMO1 deletion increased the Nppa+Nppb+Ankrd1+cardiomyocyte subcluster pro-portion after MI.In addition,the conversion of fibroblasts to myofibroblasts subclusters was inhibited in SUMO1 knockout mice.Importantly,SUMO1 loss promoted proliferation of endothelial cell subsets with the ability to reconstitute neovascularization and expressed angiogenesis-related genes.Computational analysis of ligand/receptor interactions suggested putative pathways that mediate cardiomyocytes to endothelial cell communication in the myocardium.Mice preinjected with cardiomyocyte-specific AAV-SUMO1,but not the endothelial cell-specific form,and exhibited ameliorated cardiac remodeling following MI.Collectively,our results identified the role of SUMO1 in cardiomyocytes,fibroblasts,and endothelial cells after Ml.These findings provide new insights into SUMO1 involvement in the patho-genesis of MI and reveal novel therapeutic targets.
ABSTRACT
Objective:To analyze the splicing mutation site of COL4A5 gene in a family with X-linked dominant Alport syndrome and explore the possibility of exon specific U1 small nuclear RNA (snRNA) gene therapy. Methods:The clinical data of the proband and family members of Alport syndrome were collected, and the gene mutations in the whole exon of a series of nephropathy genes in the proband were detected by high-throughput sequencing. The splice site changes and pathogenicity caused by COL4A5 c.546+5G>A mutation were analyzed by online software. Minigene experiment was used to verify and analyze the effect of COL4A5 gene mutation site c.546+5G>A in the proband of Alport syndrome family, and transient transfection and introduction of modified U1 snRNA to correct splicing mutation. Results:The results of gene sequencing showed that there was a hemizygous variation of COL4A5 gene in the proband and his half brother, and the variation site was c.546+5G>A. The results of online software for analyzing the pathogenicity of splice variation showed that the original donor splicing site could not be detected after mutation, suggesting that there was a great possibility of affecting splicing. The abnormal splicing mode of COL4A5 gene with c.546+5G>A mutation—deletion of exon 9 was verified by hybridized small gene detection. The abnormal splicing mutation could be partially corrected by the modified U1 snRNA. The correction ratios of ExSpeU1 (MT), ExSpeU1(E9+1), ExSpeU1(E9+9) and ExSpeU1(E9+11) to exon 9 deletion caused by c.546+5G>A were 0, 43.81%, 52.09% and 48.12%, respectively. Conclusions:The pathogenicity of the new splicing mutation of COL4A5 is verified, and the modified U1 snRNA can partially correct the abnormal splicing.
ABSTRACT
The discovery of integrator complex (INT) expanded our understanding of noncoding U-rich small nuclear RNA (U snRNA) maturation and transcriptional regulation, and has revived the research boom in related fields. This complex consists of at least 14 subunits, weighting over 1. 4 MD. On one hand, it functions by the cleavage of transcripts; on the other hand, it interacts with protein phosphatase 2A (PP2A) and dephosphorylates the C-terminal repeat sequence of RNA polymerase II (Pol II), thus regulating the transcriptional activity of RNA Pol II, and functions in the production of various RNAs (messenger RNA, small nuclear RNA, enhancer RNA, etc.). INT is recruited to the CTD during transcriptional initiation stage and moves along the U small nuclear RNA (snRNA) gene during transcription. Upon the recognition of 3' maturation sequence element, its cleavage activity is triggered and the matured transcripts are released. Besides, it is also involved in many other processes, including protein-coding gene transcription pause-release, transcriptional elongation, regulation of enhancer RNA transcription, and DNA and RNA metabolism etc. Meanwhile, the significance of INT complex or its components in tumorigenesis, pathogenesis, and ontogeny has being gradually highlighted. Nevertheless, the structural and compositional studies just took a new breakthrough recently. The 14 subunits that make up the complex are characterized by a large number of α-helices, which are further assembled into a huge transcription regulation machine based on the formation of functional modules. This article will mainly discuss about the composition, structural characteristics, functional studies, disease-association, and problem outlook of the integrator complex.
ABSTRACT
The integrator complex is multifunctional and contains at least 12 evolutionarily conserved subunits in hu-mans.It interacts with the C-terminal tail of the largest subunit of RNA-polymeraseⅡ ( RNAPⅡ) to promote 3′-end pro-cessing of small nuclear RNA (snRNA) U1/U2.It also interacts with RNAPⅡ, NELF and Spt5 to regulate NELF-mediated RNAPⅡpause/release and processivity at coding genes .Recently, the integrator complex is also reported to be involved in DNA damage response , dynein recruitment to the nuclear envelope , integrity of Cajal bodies , adipose differentiation , hem-atopoiesis , ciliogenesis , tumorigenesis and generation of viral microRNAs .This review discusses related research progress in the integrator complex .
ABSTRACT
Objective: To design the ribozymes to cleave human TIMP 1 mRNA, and embed them into U 6snRNA to make them stable. Methods: Ribozymes were designed according to the “hammerhead structure” described by Symons.Computer was used to analyze the possible cleavage sites. Results: Three ribozymes targeting the nt123, nt299 and nt353 on TIMP 1 mRNA were designed. Embedding ribozyme in U 6snRNA had little effect on its binding with the substrate. Conclusion: Computer assisted design is indispensable in studying ribozyme. Embedding ribozymes in U 6snRNA may be a good way to solve the problems existing in ribozyme study. [