Alternative splicing regulates vesicular trafficking genes in cardiomyocytes during postnatal heart development

Giudice, Jimena; Xia, Zheng; Wang, Eric T.; Scavuzzo, Marissa A.; Ward, Amanda J.; Kalsotra, Auinash; Wang, Wei; Wehrens, Xander H.T.; Burge, Christopher B.; Li, Wei; Cooper, Thomas A.

doi:10.1038/ncomms4603

Cited by 138 publications

(196 citation statements)

References 61 publications

(96 reference statements)

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“…However, postnatal heart development and function require an intact RNA splicing program, as both Hnrnpu and Srsf1 mutant mice die a few weeks after birth due to postnatal heart failure. A recent report showed that postnatal heart development is accompanied by major splicing changes (6). We have shown that hnRNP U plays a major role in the normal regulation of RNA splicing in the postnatal heart.…”

Section: Discussionmentioning

confidence: 66%

“…The developing heart is one of the best studied systems where splicing changes occur during normal development, and mutations affecting specific splicing outcomes contribute to cardiomyopathy (6,7). Although these mutations can either disrupt splicing elements or affect the expression of specific splicing factors, the latter mechanism is clearly responsible for the distinct splicing profiles at different developmental stages.…”

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confidence: 99%

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hnRNP U protein is required for normal pre-mRNA splicing and postnatal heart development and function

Beetz

O’Keeffe

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

106

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We report that mice lacking the heterogeneous nuclear ribonucleoprotein U (hnRNP U) in the heart develop lethal dilated cardiomyopathy and display numerous defects in cardiac pre-mRNA splicing. Mutant hearts have disorganized cardiomyocytes, impaired contractility, and abnormal excitation-contraction coupling activities. RNA-seq analyses of Hnrnpu mutant hearts revealed extensive defects in alternative splicing of pre-mRNAs encoding proteins known to be critical for normal heart development and function, including Titin and calcium/calmodulin-dependent protein kinase II delta (Camk2d). Loss of hnRNP U expression in cardiomyocytes also leads to aberrant splicing of the pre-mRNA encoding the excitation-contraction coupling component Junctin. We found that the protein product of an alternatively spliced Junctin isoform is N-glycosylated at a specific asparagine site that is required for interactions with specific protein partners. Our findings provide conclusive evidence for the essential role of hnRNP U in heart development and function and in the regulation of alternative splicing.T he expression of more than 95% of human genes is affected by alternative pre-mRNA splicing (AS) (1, 2). Differentially spliced isoforms play distinct roles in a temporally and spatially specific manner (3), and mutations that lead to aberrant splicing are the cause of many human genetic diseases (4). RNA-binding proteins (RBPs) play a central role in the regulation of alternative splicing during development and disease. They function primarily by positively or negatively regulating splice-site recognition by the spliceosome (1). Many RBPs are expressed in specific tissues, and AS is regulated by the combinatorial activities of these factors on specific pre-mRNAs through their interactions with distinct regulatory sequences in premRNA that function as splicing enhancers or silencers (5).The developing heart is one of the best studied systems where splicing changes occur during normal development, and mutations affecting specific splicing outcomes contribute to cardiomyopathy (6, 7). Although these mutations can either disrupt splicing elements or affect the expression of specific splicing factors, the latter mechanism is clearly responsible for the distinct splicing profiles at different developmental stages. For example, the dynamics of alternative splicing during postnatal heart development correlate with expression changes of many RBPs, including CUG-BP, Elavlike family member 1 (CELF1), Muscleblind-like 1 (MBNL1), and FOX proteins (8). Detailed biochemical studies have elucidated the mechanisms by which these splicing factors regulate splicing in a position-and context-dependent manner (9, 10). The function of other RBPs during heart development has also been studied. For example, two of the muscle-specific splicing factors, RBM20 and RBM24, play distinct roles in splicing regulation. RBM20 mainly acts as a splicing repressor, as its absence leads to multiple exon inclusion events in the heart. For example, the Titin gene is one of...

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Section: Discussionmentioning

confidence: 66%

mentioning

confidence: 99%

hnRNP U protein is required for normal pre-mRNA splicing and postnatal heart development and function

Beetz

O’Keeffe

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

106

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“…We identified CELF-regulated cassette exons using publicly available RNA-seq data from adult mouse hearts that ectopically expressed CELF1 or CELF2 (Giudice et al 2014;Wang et al 2015) and analyzed these cassette exons for enriched sequences and CLIP-seq features as done in the analysis of the human T cell data above. In line with the results from T cells, we found enriched occurrences of highly conserved [U]GCAUG motifs (see Methods) and RBFOX2 CLIP-seq peaks downstream of 12% -18% of CELF-repressed exons (Supplemental Fig.…”

Section: Celf2 Represses Rbfox2 Mrna and Protein Levels In T Cellsmentioning

confidence: 99%

Ancient antagonism between CELF and RBFOX families tunes mRNA splicing outcomes

Gazzara¹,

Mallory

Roytenberg

et al. 2017

Genome Res.

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Over 95% of human multi-exon genes undergo alternative splicing, a process important in normal development and often dysregulated in disease. We sought to analyze the global splicing regulatory network of CELF2 in human T cells, a well-studied splicing regulator critical to T cell development and function. By integrating high-throughput sequencing data for binding and splicing quantification with sequence features and probabilistic splicing code models, we find evidence of splicing antagonism between CELF2 and the RBFOX family of splicing factors. We validate this functional antagonism through knockdown and overexpression experiments in human cells and find CELF2 represses RBFOX2 mRNA and protein levels. Because both families of proteins have been implicated in the development and maintenance of neuronal, muscle, and heart tissues, we analyzed publicly available data in these systems. Our analysis suggests global, antagonistic coregulation of splicing by the CELF and RBFOX proteins in mouse muscle and heart in several physiologically relevant targets, including proteins involved in calcium signaling and members of the MEF2 family of transcription factors. Importantly, a number of these coregulated events are aberrantly spliced in mouse models and human patients with diseases that affect these tissues, including heart failure, diabetes, or myotonic dystrophy. Finally, analysis of exons regulated by ancient CELF family homologs in chicken, Drosophila, and Caenorhabditis elegans suggests this antagonism is conserved throughout evolution.

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“…Given the importance of Ca 2+ in GSIS and the fact that Ca 2+ signalling can be modulated by CUGBP1 in the heart [25], we hypothesised that the intracellular Ca 2+ concentration ([Ca 2+ ] i ) might modulate CUGBP1-induced GSIS impairment. To test this, [Ca 2+ ] i was recorded in isolated islets, as previously reported [40].…”

Section: Cugbp1 Is Overexpressed In the Islets Of Diabetic Micementioning

confidence: 99%

“…The association of CUGBP1 with its mRNA targets influences their alternative splicing, translation and turnover [17]. CUGBP1 is ubiquitously expressed and plays a critical role in various physiological and pathological cellular processes, including skeletal muscle differentiation and atrophy [18][19][20][21], cell proliferation [22], heart development and disease [23][24][25], and tumour growth [26]. Recently, several studies indicate that CUGBP1 may be involved in metabolic disorders.…”

Section: Introductionmentioning

confidence: 99%

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

Zhai

Yang

et al. 2016

Diabetologia

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Aims/hypothesis CUG-binding protein 1 (CUGBP1) is a multifunctional RNA-binding protein that regulates RNA processing at several stages including translation, deadenylation and alternative splicing, as well as RNA stability. Recent studies indicate that CUGBP1 may play a role in metabolic disorders. Our objective was to examine its role in endocrine pancreas function through gain-and loss-of-function experiments and to further decipher the underlying molecular mechanisms. Methods A mouse model in which type 2 diabetes was induced by a high-fat diet (HFD; 60% energy from fat) and mice on a standard chow diet (10% energy from fat) were compared. Pancreas-specific CUGBP1 overexpression and knockdown mice were generated. Different lengths of the phosphodiesterase subtype 3B (PDE3B) 3′ untranslated region (UTR) were cloned for luciferase reporter analysis. Purified CUGBP1 protein was used for gel shift experiments.Results CUGBP1 is present in rodent islets and in beta cell lines; it is overexpressed in the islets of diabetic mice. Compared with control mice, the plasma insulin level after a glucose load was significantly lower and glucose clearance was greatly delayed in mice with pancreas-specific CUGBP1 overexpression; the opposite results were obtained upon pancreas-specific CUGBP1 knockdown. Glucose-and glucagon-like peptide1 (GLP-1)-stimulated insulin secretion was significantly attenuated in mouse islets upon CUGBP1 overexpression. This was associated with a strong decrease in intracellular cAMP levels, pointing to a potential role for cAMP PDEs. CUGBP1 overexpression had no effect on the mRNA levels of PDE1A, 1C, 2A, 3A, 4A, 4B, 4D, 7A and 8B subtypes, but resulted in increased PDE3B expression. CUGBP1 was found to directly bind to a specific ATTTGTT sequence residing in the 3′ UTR of PDE3B and stabilised PDE3B mRNA. In the presence of the PDE3 inhibitor cilostamide, glucose-and GLP-1-stimulated insulin secretion was no longer reduced by CUGBP1 overexpression. Similar to CUGBP1, PDE3B was overexpressed in the islets of diabetic mice. Conclusions/interpretation We conclude that CUGBP1 is a critical regulator of insulin secretion via activating PDE3B. Repressing this protein might provide a potential strategy for treating type 2 diabetes.

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Alternative splicing regulates vesicular trafficking genes in cardiomyocytes during postnatal heart development

Cited by 138 publications

References 61 publications

hnRNP U protein is required for normal pre-mRNA splicing and postnatal heart development and function

hnRNP U protein is required for normal pre-mRNA splicing and postnatal heart development and function

Ancient antagonism between CELF and RBFOX families tunes mRNA splicing outcomes

RNA-binding protein CUGBP1 regulates insulin secretion via activation of phosphodiesterase 3B in mice

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