The Experts below are selected from a list of 48 Experts worldwide ranked by ideXlab platform
Andrea N Ladd - One of the best experts on this subject based on the ideXlab platform.
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Gene expression analyses implicate an alternative splicing program in regulating contractile gene expression and serum response factor activity in mice. PLoS One. 2013; 8:e56590. [PubMed: 23437181
2016Co-Authors: Twishasri Dasgupta, Samantha J. Stillwagon, Andrea N LaddAbstract:Members of the CUG-BP, Elav-like family (CELF) regulate alternative splicing in the heart. In MHC-CELFD transgenic mice, CELF splicing activity is inhibited postnatally in heart muscle via expression of a nuclear dominant negative CELF Protein under an a-myosin heavy chain promoter. MHC-CELFD mice develop dilated cardiomyopathy characterized by alternative splicing defects, enlarged hearts, and severe contractile dysfunction. In this study, gene expression profiles in the hearts of wild type, high- and low-expressing lines of MHC-CELFD mice were compared using microarrays. Gene ontology and pathway analyses identified contraction and calcium signaling as the most affected processes. Network analysis revealed that the serum response factor (SRF) network is highly affected. Downstream targets of SRF were up-regulated in MHC-CELFD mice compared to the wild type, suggesting an increase in SRF activity. Although SRF levels remained unchanged, known inhibitors of SRF activity were down-regulated. Conversely, we found that these inhibitors are up-regulated and downstream SRF targets are down-regulated in the hearts of MCKCUG-BP1 mice, which mildly over-express CELF1 in heart and skeletal muscle. This suggests that changes in SRF activity are a consequence of changes in CELF-mediated regulation rather than a secondary result of compensatory pathways in heart failure. In MHC-CELFD males, where the phenotype is only partially penetrant, both alternative splicing changes and down-regulation of inhibitors of SRF correlate with the development of cardiomyopathy. Together, these results strongly support a role for CELF-mediated alternative splicing i
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Gene Expression Analyses during Spontaneous Reversal of Cardiomyopathy in Mice with Repressed Nuclear CUG-BP, Elav-Like Family (CELF) Activity in Heart Muscle.
PloS one, 2015Co-Authors: Twishasri Dasgupta, Ryan J. Coram, Samantha J. Stillwagon, Andrea N LaddAbstract:CUG-BP, Elav-like family (CELF) Proteins regulate cell type- and developmental stage-specific alternative splicing in the heart. Repression of CELF-mediated splicing activity via expression of a nuclear dominant negative CELF Protein in heart muscle was previously shown to induce dysregulation of alternative splicing, cardiac dysfunction, cardiac hypertrophy, and dilated cardiomyopathy in MHC-CELFΔ transgenic mice. A “mild” line of MHC-CELFΔ mice that expresses a lower level of the dominant negative Protein exhibits cardiac dysfunction and myopathy at a young age, but spontaneously recovers normal cardiac function and heart size with age despite the persistence of splicing defects. To the best of our knowledge, this was the first example of a genetically induced cardiomyopathy that spontaneously recovers without intervention. In this study, we explored the basis for this recovery. We examined whether a transcriptional program regulated by serum response factor (SRF) that is dysregulated in juvenile MHC-CELFΔ mice is restored in the mild line with age, and evaluated global changes in gene expression by microarray analyses. We found that differences in gene expression between the mild line and wild type hearts are greatly reduced in older animals, including a partial recovery of SRF target gene expression. We did not find evidence of a new compensatory pathway being activated in the mild line with age, and propose that recovery may occur due to developmental stage-specific compatibility of CELF-dependent splice variants with the cellular environment of the cardiomyocyte.
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Gene Expression Analyses Implicate an Alternative Splicing Program in Regulating Contractile Gene Expression and Serum Response Factor Activity in Mice
PloS one, 2013Co-Authors: Twishasri Dasgupta, Samantha J. Stillwagon, Andrea N LaddAbstract:Members of the CUG-BP, Elav-like family (CELF) regulate alternative splicing in the heart. In MHC-CELFΔ transgenic mice, CELF splicing activity is inhibited postnatally in heart muscle via expression of a nuclear dominant negative CELF Protein under an α-myosin heavy chain promoter. MHC-CELFΔ mice develop dilated cardiomyopathy characterized by alternative splicing defects, enlarged hearts, and severe contractile dysfunction. In this study, gene expression profiles in the hearts of wild type, high- and low-expressing lines of MHC-CELFΔ mice were compared using microarrays. Gene ontology and pathway analyses identified contraction and calcium signaling as the most affected processes. Network analysis revealed that the serum response factor (SRF) network is highly affected. Downstream targets of SRF were up-regulated in MHC-CELFΔ mice compared to the wild type, suggesting an increase in SRF activity. Although SRF levels remained unchanged, known inhibitors of SRF activity were down-regulated. Conversely, we found that these inhibitors are up-regulated and downstream SRF targets are down-regulated in the hearts of MCKCUG-BP1 mice, which mildly over-express CELF1 in heart and skeletal muscle. This suggests that changes in SRF activity are a consequence of changes in CELF-mediated regulation rather than a secondary result of compensatory pathways in heart failure. In MHC-CELFΔ males, where the phenotype is only partially penetrant, both alternative splicing changes and down-regulation of inhibitors of SRF correlate with the development of cardiomyopathy. Together, these results strongly support a role for CELF-mediated alternative splicing in the regulation of contractile gene expression, achieved in part through modulating the activity of SRF, a key cardiac transcription factor.
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Expression of a dominant negative CELF Protein in vivo leads to altered muscle organization, fiber size, and subtype.
PloS one, 2011Co-Authors: Dara S. Berger, Michelle Moyer, Gregory M. Kliment, Erik Van Lunteren, Andrea N LaddAbstract:Background CUG-BP and ETR-3-like factor (CELF) Proteins regulate tissue- and developmental stage-specific alternative splicing in striated muscle. We previously demonstrated that heart muscle-specific expression of a nuclear dominant negative CELF Protein in transgenic mice (MHC-CELFΔ) effectively disrupts endogenous CELF activity in the heart in vivo, resulting in impaired cardiac function. In this study, transgenic mice that express the dominant negative Protein under a skeletal muscle-specific promoter (Myo-CELFΔ) were generated to investigate the role of CELF-mediated alternative splicing programs in normal skeletal muscle. Methodology/principal findings Myo-CELFΔ mice exhibit modest changes in CELF-mediated alternative splicing in skeletal muscle, accompanied by a reduction of endomysial and perimysial spaces, an increase in fiber size variability, and an increase in slow twitch muscle fibers. Weight gain and mean body weight, total number of muscle fibers, and overall muscle strength were not affected. Conclusions/significance Although these findings demonstrate that CELF activity contributes to the normal alternative splicing of a subset of muscle transcripts in vivo, the mildness of the effects in Myo-CELFΔ muscles compared to those in MHC-CELFΔ hearts suggests CELF activity may be less determinative for alternative splicing in skeletal muscle than in heart muscle. Nonetheless, even these small changes in CELF-mediated splicing regulation were sufficient to alter muscle organization and muscle fiber properties affected in myotonic dystrophy. This lends further evidence to the hypothesis that dysregulation of CELF-mediated alternative splicing programs may be responsible for the disruption of these properties during muscle pathogenesis.
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the neurofibromatosis type i pre mrna is a novel target of CELF Protein mediated splicing regulation
Nucleic Acids Research, 2010Co-Authors: Victoria A Barron, Melissa N Hinman, Andrea N LaddAbstract:The CUG-BP and ETR-3 like factors (CELF) are a family of six highly conserved RNA-binding Proteins that preferentially bind to UG-rich sequences. One of the key functions of these Proteins is to mediate alternative splicing in a number of tissues, including brain, heart and muscle. To fully understand the function of CELF Proteins, it is important to identify downstream targets of CELF Proteins. In this communication, we report that neurofibromatosis type I (NF1) exon 23a is a novel target of CELF Protein-mediated splicing regulation in neuron-like cells. NF1 regulates Ras signaling, and the isoform that excludes exon 23a shows 10 times greater ability to down-regulate Ras signaling than the isoform that includes exon 23a. Five of the six CELF Proteins strongly suppress the inclusion of NF1 exon 23a. Over-expression or siRNA knockdown of these Proteins in cell transfection experiments altered the levels of NF1 exon 23a inclusion. In vitro binding and splicing analyses demonstrate that CELF Proteins block splicing through interfering with binding of U2AF65. These studies, combined with our previous investigations demonstrating a role for Hu Proteins and TIA-1/TIAR in controlling NF1 exon 23a inclusion, highlight the complex nature of regulation of this important alternative splicing event.
Rodney F. Minchin - One of the best experts on this subject based on the ideXlab platform.
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The MBNL/CELF Splicing Factors Regulate Cytosolic Sulfotransferase 4A1 Protein Expression during Cell Differentiation
Drug metabolism and disposition: the biological fate of chemicals, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
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the mbnl CELF splicing factors regulate cytosolic sulfotransferase 4a1 Protein expression during cell differentiation
Drug Metabolism and Disposition, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
Misgana Idris - One of the best experts on this subject based on the ideXlab platform.
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The MBNL/CELF Splicing Factors Regulate Cytosolic Sulfotransferase 4A1 Protein Expression during Cell Differentiation
Drug metabolism and disposition: the biological fate of chemicals, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
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the mbnl CELF splicing factors regulate cytosolic sulfotransferase 4a1 Protein expression during cell differentiation
Drug Metabolism and Disposition, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
Neville J. Butcher - One of the best experts on this subject based on the ideXlab platform.
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The MBNL/CELF Splicing Factors Regulate Cytosolic Sulfotransferase 4A1 Protein Expression during Cell Differentiation
Drug metabolism and disposition: the biological fate of chemicals, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
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the mbnl CELF splicing factors regulate cytosolic sulfotransferase 4a1 Protein expression during cell differentiation
Drug Metabolism and Disposition, 2019Co-Authors: Misgana Idris, Neville J. Butcher, Rodney F. MinchinAbstract:Sulfotransferase 4A1 (SULT4A1) is a sulfotransferase-like Protein that is highly conserved between species. In human tissues, there are two transcripts, one that produces a full-length Protein and one that produces an unstable truncated Protein. The second transcript, which includes a pseudo-exon between exons 6 and 7 (6p), is widely expressed, whereas the first is more restricted. Differentiation of neuronal cells results in the removal of the pseudo-exon and subsequent SULT4A1 Protein expression. Recent studies with SULT4A1 knockout mice showed that the Protein is essential for normal development and that its absence leads to a severe neurologic phenotype. Here, the regulation of SULT4A1 6p splicing was investigated during neuronal differentiation using SH-SY5Y cells, human induced pluripotent stem cells, and mouse embryonic tissue. In all three models, pseudo-exon 6p was removed during differentiation, resulting in stable SULT4A1 Protein expression. Using a minigene splicing assay, a region upstream of pseudo-exon 6p was identified that is essential for correct splicing of SULT4A1 mRNA. Within this region, there were binding motifs for four RNA processing factors (MBNL-1, MBNL-2, CELF-1, and CELF-2). Time-dependent changes in SULT4A1 Protein and MBNL/CELF Protein during differentiation supported their role in correctly splicing the SULT4A1 mRNA. Furthermore, ectopic expression of each factor produced efficient splicing in the minigene assay as well as correct splicing of the endogenous SULT4A1 mRNA. These results show that SULT4A1 mRNA is a target for MBNL/CELF-dependent splicing, which may be essential in producing stable, functional SULT4A1.
Thomas A. Cooper - One of the best experts on this subject based on the ideXlab platform.
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Cardiac Tissue-Specific Repression of CELF Activity Disrupts Alternative Splicing and Causes Cardiomyopathy
Molecular and cellular biology, 2005Co-Authors: Andrea N Ladd, George E. Taffet, Craig J. Hartley, Debra L. Kearney, Thomas A. CooperAbstract:Members of the CELF family of RNA binding Proteins have been implicated in alternative splicing regulation in developing heart. Transgenic mice that express a nuclear dominant-negative CELF Protein specifically in the heart (MHC-CELFDelta) develop cardiac hypertrophy and dilated cardiomyopathy with defects in alternative splicing beginning as early as 3 weeks after birth. MHC-CELFDelta mice exhibit extensive cardiac fibrosis, severe cardiac dysfunction, and premature death. Interestingly, the penetrance of the phenotype is greater in females than in males despite similar levels of dominant-negative expression, suggesting that there is sex-specific modulation of splicing activity. The cardiac defects in MHC-CELFdelta mice are directly attributable to reduced levels of CELF activity, as crossing these mice with mice overexpressing CUG-BP1, a wild-type CELF Protein, rescues defects in alternative splicing, the severity and incidence of cardiac hypertrophy, and survival. We conclude that CELF Protein activity is required for normal alternative splicing in the heart in vivo and that normal CELF-mediated alternative splicing regulation is in turn required for normal cardiac function.
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ETR‐3 and CELF4 Protein domains required for RNA binding and splicing activity in vivo
Nucleic acids research, 2004Co-Authors: Gopal Singh, Nicolas Charlet-b., Jin Han, Thomas A. CooperAbstract:Members of the CUG-BP and ETR-3 like factor (CELF) Protein family bind within conserved intronic elements (called MSEs) flanking the cardiac troponin T (cTNT) alternative exon 5 and promote exon inclusion in vivo and in vitro. Here we use a comparative deletion analysis of two family members (ETR-3 and CELF4) to identify separate domains required for RNA binding and splicing activity in vivo. CELF Proteins contain two adjacent RNA binding domains (RRM1 and RRM2) near the N-terminus and one RRM (RRM3) near the C-terminus, which are separated by a 160-230 residue divergent domain of unknown function. Either RRM1 or RRM2 of CELF4 are necessary and sufficient for binding MSE RNA and RRM2 plus an additional 66 amino acids of the divergent domain are as effective as full-length Protein in activating MSE-dependent splicing in vivo. Non-overlapping N- and C-terminal regions of ETR-3 containing either RRM1 and RRM2 or RRM3 plus segments of the adjacent divergent domain activate MSE-dependent exon inclusion demonstrating an unusual functional redundancy of the N- and C-termini of the Protein. These results identify specific regions of ETR-3 and CELF4 that are likely targets of Protein-Protein interactions required for splicing activation.