Splicing inhibition of U2AF
            <sup>65</sup>
            leads to alternative exon skipping

Cho, Sunghee; Moon, Heegyum; Loh, Tiing Jen; Jang, Ha Na; Liu, Yongchao; Zhou, Jianhua; Ohn, Takbum; Zheng, Xuexiu; Shen, Haihong

doi:10.1073/pnas.1500639112

Cited by 46 publications

(58 citation statements)

References 54 publications

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“…20 Similarly, studies utilizing human cell lines show that knockdown of U2AF2 shifts the splicing toward introns harboring strong polypyrimidine tracts, 21 and that U2AF2 dependency can be eliminated by replacement of a stronger 5ss. 22 In addition, in fission yeast S. pombe, a large number of introns remain efficiently spliced upon the inactivation of U2AF, 59 the U2AF2 homolog. In this case stronger 5ss and A/U-rich sequence in the 5 0 intronic region are associated with U2AF 59 -insensitive introns.…”

Section: Introductionmentioning

confidence: 99%

RNA structure in splicing: An evolutionary perspective

2016

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Pre-mRNA splicing is a key post-transcriptional regulation process in which introns are excised and exons are ligated together. A novel class of structured intron was recently discovered in fish. Simple expansions of complementary AC and GT dimers at opposite boundaries of an intron were found to form a bridging structure, thereby enforcing correct splice site pairing across the intron. In some fish introns, the RNA structures are strong enough to bypass the need of regulatory protein factors for splicing. Here, we discuss the prevalence and potential functions of highly structured introns. In humans, structured introns usually arise through the co-occurrence of C and G-rich repeats at intron boundaries. We explore the potentially instructive example of the HLA receptor genes. In HLA pre-mRNA, structured introns flank the exons that encode the highly polymorphic b sheet cleft, making the processing of the transcript robust to variants that disrupt splicing factor binding. While selective forces that have shaped HLA receptor are fairly atypical, numerous other highly polymorphic genes that encode receptors contain structured introns. Finally, we discuss how the elevated mutation rate associated with the simple repeats that often compose structured intron can make structured introns themselves rapidly evolving elements.

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Section: Introductionmentioning

confidence: 99%

RNA structure in splicing: An evolutionary perspective

2016

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“…http://dx.doi.org/10.1101/199927 doi: bioRxiv preprint first posted online Oct. 8, 2017; is usually a rarer event in higher eukaryotes that contain longer introns 32 . We chose two longer intronic backbones (~300-600 bp) shown previously to not suffer from such intronic retention ( C. griseus DHFR and human SMN1 intron backbones), and found that the longer intron lengths improved both expression and assay reproducibility 33,34 . Exon inclusion metrics obtained from both of these intron contexts were highly reproducible between biological replicates (Fig.…”

Section: Main Textmentioning

confidence: 99%

Many rare genetic variants have unrecognized large-effect disruptions to exon recognition

Cheung

Yao

et al. 2017

Preprint

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Any individual’s genome contains ∼4-5 million genetic variants that differ from reference, and understanding how these variants give rise to trait diversity and disease susceptibility is a central goal of human genetics1. A vast majority (96-99%) of an individual’s variants are common, though at a population level the overwhelming majority of variants are rare2–5. Because of their scarcity in an individual’s genome, rare variants that play important roles in complex traits are likely to have large functional effects6,7. Mutations that cause an exon to be skipped can have severe functional consequences on gene function, and many known disease-causing mutations reduce or eliminate exon recognition8. Here we explore the extent to which rare genetic variation in humans results in near complete loss of exon recognition. We developed a Multiplexed Functional Assay of Splicing using Sort-seq (MFASS) that allows us to measure exon inclusion in thousands of human exons and surrounding intronic sequence simultaneously. We assayed 27,733 extant variants in the Exome Aggregation Consortium (ExAC)9 within or adjacent to 2,339 human exons, and found that 3.8% (1,050) of the variants, almost all of which were extremely rare, led to large-effect defects in exon recognition. Importantly, we find that 83% of these splice-disrupting variants (SDVs) are located outside of canonical splice sites, are distributed evenly across distinct exonic and intronic regions, and are difficult to predict a priori. Our results indicate that loss of exon recognition is an important and underappreciated means by which rare variants exert large functional effects, and that MFASS enables their empirical assessment for splicing defects at scale.

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“…In this study, we investigated the regulatory role of U2AF65 in milk synthesis and growth of bovine mammary epithelial cells (BMECs). 7 The U2AF homology motif kinase 1 (UHMK1), a serine/threonine protein kinase, regulates the cell cycle via a tumour suppressor cyclin-dependent kinase inhibitor. Treatment with amino acids (Met and Leu) and hormones (prolactin and β-estradiol) upregulated the expression of U2AF65 in these cells.…”

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U2AF65 enhances milk synthesis and growth of bovine mammary epithelial cells by positively regulating the mTOR‐SREBP‐1c signalling pathway

Zhen

et al. 2019

Cell Biochemistry & Function

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U2 snRNP auxiliary factor 65 kDa (U2AF65) is a splicing factor that promotes prespliceosome assembly. The function of U2AF65 in alternative splicing has been identified; however, the essential physiological role of U2AF65 remains poorly understood. In this study, we investigated the regulatory role of U2AF65 in milk synthesis and growth of bovine mammary epithelial cells (BMECs). Our results showed that U2AF65 localizes in the nucleus. Treatment with amino acids (Met and Leu) and hormones (prolactin and β-estradiol) upregulated the expression of U2AF65 in these cells. U2AF65 overexpression increased the synthesis of β-casein, triglycerides, and lactose; increased cell viability; and promoted proliferation of BMECs. Furthermore, our results showed that U2AF65 positively regulated mTOR phosphorylation and expression of mature mRNA of mTOR and SREBP-1c. Collectively, our findings demonstrate that U2AF65 regulates the mRNA expression of signalling molecules (mTOR and SREBP-1c) involved in milk synthesis and growth of BMECs, possibly via controlling the splicing and maturation of these mRNAs. U2 snRNP auxiliary factor 65 kDa (U2AF65) is a splicing factor that promotes prespliceosome assembly. The essential physiological role of U2AF65 remains poorly understood. In the present study, we confirmed that U2AF65 functions as a positive regulator of milk synthesis in and proliferation of bovine mammary epithelial cells via the mTOR-SREBP-1c signalling pathway. Therefore, our study uncovers the regulatory role of U2AF65 in milk synthesis and cell proliferation. KEYWORDS U2AF65, mTOR, SREBP-1c, β-casein, triglyceride, lactose 1 | INTRODUCTION U2 small nuclear ribonucleoprotein (U2snRNP) auxiliary factor (U2AF) is a conserved nuclear factor that plays a crucial role in mRNA splicing. U2AF is a stable heterodimer comprising U2AF65 subunit (65 kDa), which is encoded by U2AF2 gene, and the U2AF35 (35 kDa) subunit, which is encoded by the U2AF1 gene. U2AF65 interacts with the polypyrimidine tract (PPT), a conserved 3′ splice-site (ss) immediately downstream of the branch point sequence (BPS). On the other hand, U2AF35 interacts with invariant AG dinucleotides at the 3′ ss and stabilizes the binding of U2AF65 with RNA. 1,2A series of U2AF mutations have been characterized to investigate the functions of U2AF as a splicing factor, and previous studies have revealed that the stoichiometry of U2AF is important for accurate 3′ ss selection. 3,4 Consistent with the disease phenotype of myelodysplasia, one study indicated that mutations in U2AF cause widespread changes in alternative splicing and is linked to decreased cell proliferation. 5 U2AF has also been reported to be involved in other cellular processes, such as mRNA export, in which it binds to

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Splicing inhibition of U2AF ⁶⁵ leads to alternative exon skipping

Cited by 46 publications

References 54 publications

RNA structure in splicing: An evolutionary perspective

RNA structure in splicing: An evolutionary perspective

Many rare genetic variants have unrecognized large-effect disruptions to exon recognition

U2AF65 enhances milk synthesis and growth of bovine mammary epithelial cells by positively regulating the mTOR‐SREBP‐1c signalling pathway

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Splicing inhibition of U2AF 65 leads to alternative exon skipping

Cited by 46 publications

References 54 publications

RNA structure in splicing: An evolutionary perspective

RNA structure in splicing: An evolutionary perspective

Many rare genetic variants have unrecognized large-effect disruptions to exon recognition

U2AF65 enhances milk synthesis and growth of bovine mammary epithelial cells by positively regulating the mTOR‐SREBP‐1c signalling pathway

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Splicing inhibition of U2AF ⁶⁵ leads to alternative exon skipping