Huntington’s disease (HD) is a debilitating neurodegenerative disorder caused by an abnormal expansion of CAG repeats (Cytosine, Adenine, Guanine) in the huntingtin gene (HTT). This mutation leads to the production o...Huntington’s disease (HD) is a debilitating neurodegenerative disorder caused by an abnormal expansion of CAG repeats (Cytosine, Adenine, Guanine) in the huntingtin gene (HTT). This mutation leads to the production of a mutant huntingtin protein, resulting in neuronal dysfunction and cell death. Current treatments primarily focus on symptomatic relief and do not address the underlying genetic cause. This review explores spliceosome-mediated RNA trans-splicing (SMaRT) therapy as an innovative and potential approach for HD treatment. SMaRT leverages the cell’s natural splicing machinery to correct mutant mRNA, thereby reducing toxic protein levels while restoring functional protein production. We compare SMaRT with other gene therapy strategies, such as antisense oligonucleotides, RNA interference, and CRISPR-based systems, highlighting SMaRT’s dual-action mechanism and its potential advantages in clinical applications. Additionally, we discuss the challenges and future directions for SMaRT therapy, emphasizing the need for further research to optimize its efficacy and safety. This review aims to provide a comprehensive overview of current and emerging therapies for HD, with a focus on the innovative potential of SMaRT.展开更多
Dietary restriction usually suppresses biosynthesis but activates catabolic pathways in animals.However,the short-term starvation enhances biosynthetic activities and promotes ribosomal biogenesis in adult Caenorhabdi...Dietary restriction usually suppresses biosynthesis but activates catabolic pathways in animals.However,the short-term starvation enhances biosynthetic activities and promotes ribosomal biogenesis in adult Caenorhabditis elegans.The mechanism underlying the processes remains largely unknown.Here,we find that the short-term starvation enhances the SL1 trans-splicing of translation-related genes in adult C.elegans by transcriptome analysis.The small nuclear RNA-activating protein complex(SNAPc)promotes SL RNA production and mediates starvation-induced trans-splicing.TOFU-5,a core factor in the upstream sequence transcription complex(USTC)essential for piRNA production,is also involved in the starvationinduced trans-splicing processes.Knocking down components of the SNAPc complex and tofu-5 extends worm survival under starvation conditions.Taken together,our study highlights the importance of SL transsplicing in the nutrition response and reveals a mechanism of the survival regulation by food deprivation via SNAPc and TOFU-5.展开更多
A dual-vector system was explored for the delivery of the coagulation factor VIII gene,using intein-mediated protein trans-splicing as a means to produce intact functional factor VIII post-translationally.A pair of eu...A dual-vector system was explored for the delivery of the coagulation factor VIII gene,using intein-mediated protein trans-splicing as a means to produce intact functional factor VIII post-translationally.A pair of eukaryotic expression vectors,expressing Ssp DnaB intein-fused heavy and light chain genes of B-domain deleted factor VIII (BDD-FVIII),was constructed.With transient co-transfection of the two vectors into 293 and COS-7 cells,the culture supernatants contained (137±23) and (109±22) ng mL–1 spliced BDD-FVIII antigen with an activity of (1.05±0.16) and (0.79±0.23) IU mL–1 for 293 and COS-7 cells,respectively.The spliced BDD-FVIII was also detected in supernatants from a mixture of cells transfected with inteinfused heavy and light chain genes.The spliced BDD-FVIII protein bands from cell lysates were visualized by Western blotting.The data demonstrated that intein could be used to transfer the split factor VIII gene and provided valuable information on factor VIII gene delivery by dual-adeno-associated virus in hemophilia A gene therapy.展开更多
Protein trans-splicing based dual-vector factor VIII （FVIII） gene delivery is adversely affected by less efficiency of protein splicing. We sought to increase the amount of spliced FVIII protein and plasma coagulati...Protein trans-splicing based dual-vector factor VIII （FVIII） gene delivery is adversely affected by less efficiency of protein splicing. We sought to increase the amount of spliced FVIII protein and plasma coagulation activity in dual-vector FVIII transgene in mice by means of strengthening the interaction of inteins, protein splicing elements, thereby facilitating protein trans-splicing. Dual-vector delivery of the FVIII gene in cultured cells showed that replacement of Met226 in the heavy chain and Asp1828 in the light chain with Cys residues could facilitate inter-chain disulfide linking and improve protein trans-splicing, increasing the levels of spliced FVIII protein. In this study, C57BL/6 mice were coadministered dual vectors of intein-fused human FVIII heavy chain and light chain with Cys mutations via portal vein injection into the liver. Forty-eight hours post-injection, plasma was collected and analyzed for FVIII antigen concentration and coagulation activity. These mice showed increased circulating FVIII heavy chain polypeptide （442± 151 ng mL i vs. 305±103 ng mL-1） and coagulation activi- ty （1.46±0.37 IU mL i vs. 0.85±0.23 IU mL-1） compared with control mice co-administered dual vectors expressing the heavy and light chains of wild-type FVIII. Moreover, coagulation activity was similar to that of mice receiving a single vector ex- pressing FVIII （1.79_＋0.59 IU mL-l）. These findings indicate that improving protein trans-splicing by inter-chain disulfide bonding is a promising approach for increasing the efficacy of dual-vector based FVIII gene transfer.展开更多
Fusion transcripts are commonly found in eukaryotes, and many aberrant fusions are associated with severe diseases, including cancer. One class of fusion transcripts is generated by joining separate transcripts throug...Fusion transcripts are commonly found in eukaryotes, and many aberrant fusions are associated with severe diseases, including cancer. One class of fusion transcripts is generated by joining separate transcripts through trans-splicing. However, the mechanism of trans-splicing in mammals remains largely elusive. Here we showed evidence to support an intuitive hypothesis that attributes trans-splicing to the spatial proximity between premature transcripts. A novel trans-splicing detection tool(TSD) was developed to reliably identify intra-chromosomal trans-splicing events(i TSEs) from RNA-seq data. TSD can maintain a remarkable balance between sensitivity and accuracy, thus distinguishing it from most state-of-the-art tools. The accuracy of TSD was experimentally demonstrated by excluding potential false discovery from mosaic genome or template switching during PCR. We showed that i TSEs identified by TSD were frequently found between genomic regulatory elements, which are known to be more prone to interact with each other. Moreover, i TSE sites may be more physically adjacent to each other than random control in the tested human lymphoblastoid cell line according to Hi-C data. Our results suggest that trans-splicing and 3 D genome architecture may be coupled in mammals and that our pipeline, TSD, may facilitate investigations of trans-splicing on a systematic and accurate level previously thought impossible.展开更多
With the development of deep sequencing and bioinformatics technology, a large number of products produced by abnormal RNA splicing, such as chimeric RNA and chimeric/fusion proteins, have been discovered. Natural chi...With the development of deep sequencing and bioinformatics technology, a large number of products produced by abnormal RNA splicing, such as chimeric RNA and chimeric/fusion proteins, have been discovered. Natural chimeric/fusion genes are new genes formed by natural fusion of two or more independent genes. Chimeric RNAs can be transcribed by natural chimeric genes, and can also be formed by cis-splicing or trans-splicing of two or more precursor mRNAs. Unlike fusion genes, the production of chimeric RNAs does not involve changes in the DNA level of chromosomes. At first, chimeric RNAs were found as tumor markers. With the deepening of research, researchers also found a large number of chimeric RNAs in normal tissues. From the perspective of biological function, chimeric RNAs can play a biological role in regulating the expression of corresponding maternal genes, translating into chimeric proteins, and forming long non-coding RNAs. The objective of the present study focused on the frontiers of chimeric RNA and reviewed its role in health and tumor study to reveal research progress of chimeric RNA and health and provide a new sight of relative disease treatment. The main conclusion of this review is that chimeric RNA may serve as a biomarker for specific tumor diagnose and treatment while its role in normal physiology needs to be revealed.展开更多
Gene fusions are appreciated as ideal cancer biomarkers and therapeutic targets.Chimeric RNAs are traditionally thought to be products of gene fusions,and thus,also cancerspecific.Recent research has demonstrated that...Gene fusions are appreciated as ideal cancer biomarkers and therapeutic targets.Chimeric RNAs are traditionally thought to be products of gene fusions,and thus,also cancerspecific.Recent research has demonstrated that chimeric RNAs can be generated by intergenic splicing in the absence of gene fusion,and such chimeric RNAs are also found in normal physiology.These new findings challenge the traditional theory of chimeric RNAs exclusivity to cancer,and complicates use of chimeric RNAs in cancer detection.Here,we provide an overview of gene fusions and chimeric RNAs,and emphasize their differences.We note that gene fusions are able to generate chimeric RNAs in accordance with the central dogma of biology,and that chimeric RNAs may also be able to influence the generation of the gene fusions per the“horse before the cart”hypothesis.We further expand upon the“horse before the cart”hypothesis,summarizing current evidence in support of the theory and exploring its potential impact on the field.展开更多
Introduction.Specific gene fusions and their resultant fusion products,including chimeric ribonucleic acid(RNA)and protein,have long served as ideal tumor diagnostic markers and therapeutic targets.Despite this,only a...Introduction.Specific gene fusions and their resultant fusion products,including chimeric ribonucleic acid(RNA)and protein,have long served as ideal tumor diagnostic markers and therapeutic targets.Despite this,only a few systematic studies on chimeric RNAs have been conducted in prostate cancer(PCa).In this study,we summarize the discovery pipeline,formation mechanisms,method of action,and future perspectives of chimeric RNAs.We aim to provide a viewpoint for exploring novel targets for diagnosing or treating PCa.展开更多
基金National Natural Science Foundation of China under Grant 22376100.
文摘Huntington’s disease (HD) is a debilitating neurodegenerative disorder caused by an abnormal expansion of CAG repeats (Cytosine, Adenine, Guanine) in the huntingtin gene (HTT). This mutation leads to the production of a mutant huntingtin protein, resulting in neuronal dysfunction and cell death. Current treatments primarily focus on symptomatic relief and do not address the underlying genetic cause. This review explores spliceosome-mediated RNA trans-splicing (SMaRT) therapy as an innovative and potential approach for HD treatment. SMaRT leverages the cell’s natural splicing machinery to correct mutant mRNA, thereby reducing toxic protein levels while restoring functional protein production. We compare SMaRT with other gene therapy strategies, such as antisense oligonucleotides, RNA interference, and CRISPR-based systems, highlighting SMaRT’s dual-action mechanism and its potential advantages in clinical applications. Additionally, we discuss the challenges and future directions for SMaRT therapy, emphasizing the need for further research to optimize its efficacy and safety. This review aims to provide a comprehensive overview of current and emerging therapies for HD, with a focus on the innovative potential of SMaRT.
基金supported by grants from the National Key R&D Program of China (2019YFA0802600)the Strategic Priority Research Program of the Chinese Academy of Sciences (XDB39010600)+3 种基金the National Natural Science Foundation of China (91940303, 31870812, 32070619, 31871300 and 31900434)the China Postdoctoral Science Foundation (2018M632542)the Anhui Natural Science Foundation (1808085QC82 and 1908085QC96)supported in part by the Fundamental Research Funds for the Central Universities.
文摘Dietary restriction usually suppresses biosynthesis but activates catabolic pathways in animals.However,the short-term starvation enhances biosynthetic activities and promotes ribosomal biogenesis in adult Caenorhabditis elegans.The mechanism underlying the processes remains largely unknown.Here,we find that the short-term starvation enhances the SL1 trans-splicing of translation-related genes in adult C.elegans by transcriptome analysis.The small nuclear RNA-activating protein complex(SNAPc)promotes SL RNA production and mediates starvation-induced trans-splicing.TOFU-5,a core factor in the upstream sequence transcription complex(USTC)essential for piRNA production,is also involved in the starvationinduced trans-splicing processes.Knocking down components of the SNAPc complex and tofu-5 extends worm survival under starvation conditions.Taken together,our study highlights the importance of SL transsplicing in the nutrition response and reveals a mechanism of the survival regulation by food deprivation via SNAPc and TOFU-5.
基金supported by the Natural Science Foundation of Shandong Province, China (Grant No. Y2005D14)the Science and Technology Program of Yantai (Grant No. 2008152)+1 种基金the Scientific Research Foundation for the Returned Overseas Chinese Scholars, State Education Ministry (Grant No. 20071108)the Discipline Construction Funds of Ludong University
文摘A dual-vector system was explored for the delivery of the coagulation factor VIII gene,using intein-mediated protein trans-splicing as a means to produce intact functional factor VIII post-translationally.A pair of eukaryotic expression vectors,expressing Ssp DnaB intein-fused heavy and light chain genes of B-domain deleted factor VIII (BDD-FVIII),was constructed.With transient co-transfection of the two vectors into 293 and COS-7 cells,the culture supernatants contained (137±23) and (109±22) ng mL–1 spliced BDD-FVIII antigen with an activity of (1.05±0.16) and (0.79±0.23) IU mL–1 for 293 and COS-7 cells,respectively.The spliced BDD-FVIII was also detected in supernatants from a mixture of cells transfected with inteinfused heavy and light chain genes.The spliced BDD-FVIII protein bands from cell lysates were visualized by Western blotting.The data demonstrated that intein could be used to transfer the split factor VIII gene and provided valuable information on factor VIII gene delivery by dual-adeno-associated virus in hemophilia A gene therapy.
基金supported by the Natural Science Foundation of Shandong Province (ZR2010CM061)the Scientific Research Foundation from Ministry of Education for Returned Overseas Chinese Scholars (20071108)
文摘Protein trans-splicing based dual-vector factor VIII （FVIII） gene delivery is adversely affected by less efficiency of protein splicing. We sought to increase the amount of spliced FVIII protein and plasma coagulation activity in dual-vector FVIII transgene in mice by means of strengthening the interaction of inteins, protein splicing elements, thereby facilitating protein trans-splicing. Dual-vector delivery of the FVIII gene in cultured cells showed that replacement of Met226 in the heavy chain and Asp1828 in the light chain with Cys residues could facilitate inter-chain disulfide linking and improve protein trans-splicing, increasing the levels of spliced FVIII protein. In this study, C57BL/6 mice were coadministered dual vectors of intein-fused human FVIII heavy chain and light chain with Cys mutations via portal vein injection into the liver. Forty-eight hours post-injection, plasma was collected and analyzed for FVIII antigen concentration and coagulation activity. These mice showed increased circulating FVIII heavy chain polypeptide （442± 151 ng mL i vs. 305±103 ng mL-1） and coagulation activi- ty （1.46±0.37 IU mL i vs. 0.85±0.23 IU mL-1） compared with control mice co-administered dual vectors expressing the heavy and light chains of wild-type FVIII. Moreover, coagulation activity was similar to that of mice receiving a single vector ex- pressing FVIII （1.79_＋0.59 IU mL-l）. These findings indicate that improving protein trans-splicing by inter-chain disulfide bonding is a promising approach for increasing the efficacy of dual-vector based FVIII gene transfer.
基金supported by the National Natural Science Foundation of China(31671342,31871331,91540114,and 31401112)。
文摘Fusion transcripts are commonly found in eukaryotes, and many aberrant fusions are associated with severe diseases, including cancer. One class of fusion transcripts is generated by joining separate transcripts through trans-splicing. However, the mechanism of trans-splicing in mammals remains largely elusive. Here we showed evidence to support an intuitive hypothesis that attributes trans-splicing to the spatial proximity between premature transcripts. A novel trans-splicing detection tool(TSD) was developed to reliably identify intra-chromosomal trans-splicing events(i TSEs) from RNA-seq data. TSD can maintain a remarkable balance between sensitivity and accuracy, thus distinguishing it from most state-of-the-art tools. The accuracy of TSD was experimentally demonstrated by excluding potential false discovery from mosaic genome or template switching during PCR. We showed that i TSEs identified by TSD were frequently found between genomic regulatory elements, which are known to be more prone to interact with each other. Moreover, i TSE sites may be more physically adjacent to each other than random control in the tested human lymphoblastoid cell line according to Hi-C data. Our results suggest that trans-splicing and 3 D genome architecture may be coupled in mammals and that our pipeline, TSD, may facilitate investigations of trans-splicing on a systematic and accurate level previously thought impossible.
文摘With the development of deep sequencing and bioinformatics technology, a large number of products produced by abnormal RNA splicing, such as chimeric RNA and chimeric/fusion proteins, have been discovered. Natural chimeric/fusion genes are new genes formed by natural fusion of two or more independent genes. Chimeric RNAs can be transcribed by natural chimeric genes, and can also be formed by cis-splicing or trans-splicing of two or more precursor mRNAs. Unlike fusion genes, the production of chimeric RNAs does not involve changes in the DNA level of chromosomes. At first, chimeric RNAs were found as tumor markers. With the deepening of research, researchers also found a large number of chimeric RNAs in normal tissues. From the perspective of biological function, chimeric RNAs can play a biological role in regulating the expression of corresponding maternal genes, translating into chimeric proteins, and forming long non-coding RNAs. The objective of the present study focused on the frontiers of chimeric RNA and reviewed its role in health and tumor study to reveal research progress of chimeric RNA and health and provide a new sight of relative disease treatment. The main conclusion of this review is that chimeric RNA may serve as a biomarker for specific tumor diagnose and treatment while its role in normal physiology needs to be revealed.
基金Hao Wu was supported by China Scholarship Council(CSC,No.201706370109).We thank Emily Lin for her help in creating the figure.We thank Justin Elfman for his help with English editing.
文摘Gene fusions are appreciated as ideal cancer biomarkers and therapeutic targets.Chimeric RNAs are traditionally thought to be products of gene fusions,and thus,also cancerspecific.Recent research has demonstrated that chimeric RNAs can be generated by intergenic splicing in the absence of gene fusion,and such chimeric RNAs are also found in normal physiology.These new findings challenge the traditional theory of chimeric RNAs exclusivity to cancer,and complicates use of chimeric RNAs in cancer detection.Here,we provide an overview of gene fusions and chimeric RNAs,and emphasize their differences.We note that gene fusions are able to generate chimeric RNAs in accordance with the central dogma of biology,and that chimeric RNAs may also be able to influence the generation of the gene fusions per the“horse before the cart”hypothesis.We further expand upon the“horse before the cart”hypothesis,summarizing current evidence in support of the theory and exploring its potential impact on the field.
基金supported by the China Scholarship Council(No.201906380075)Guangdong Province Natural Science Foundation(No.2023A1515011905).
文摘Introduction.Specific gene fusions and their resultant fusion products,including chimeric ribonucleic acid(RNA)and protein,have long served as ideal tumor diagnostic markers and therapeutic targets.Despite this,only a few systematic studies on chimeric RNAs have been conducted in prostate cancer(PCa).In this study,we summarize the discovery pipeline,formation mechanisms,method of action,and future perspectives of chimeric RNAs.We aim to provide a viewpoint for exploring novel targets for diagnosing or treating PCa.
