Lu Cai*, Yongqiang Xing, Xiujuan Zhao, Tao Yu, Dong Liang, Jun Li and Guanyun Wei
School of Mathematics, Physics and Biological Engineering, Inner Mongolia University of Science and Technology, Baotou, 014010, China
Received date: November 23, 2014; Accepted date: November 25, 2014; Published date: November 27, 2014
Citation: Cai L, Xing Y, Zhao X, Yu T, Liang D, et al. (2014) Elucidating the Regulation Mechanism of Alternative Splicing. Mol Biol 3:e122. doi: 10.4172/2168-9547.1000e122
Copyright: © 2014 Cai L, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
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Alternative splicing (AS) have increasingly attracted researchers’ attention since the discovery that gene number on a genome is not linearly correlated with the complexity and functional diversity of an organism. AS is widespread in human genome and has been investigating intensively in many human genes. By recent estimates, over 90% of human multi-exon genes are spliced alternatively [1,2]. AS of pre-mRNA is essential for regulating gene expression in higher eukaryotes. It alters the action of a gene in different tissues and developmental states by generating different mRNA isoforms which are composed of different selection of exons.It has been demonstrated that AS is implicated in numerous processes, including post-translational modification, development, differentiation, etc [3,4]. Moreover, the misregulation of AS has an impact on the function of the protein and leads to a number of human genetic diseases .
Tï»¿ï»¿he process of removing non-coding introns and concatenating remaining exons to form mature mRNAs occurs in the nucleus and is accompanied byinteraction of five small nuclear ribonucleoproteins (U1, U2, U4, U5 and U6 sRNPs) and over two hundred proteins on step-by-step by using ATP . AS patterns can be categorized into seven major types, such as (i)exon skipping; (ii) 3’-AS; (iii) 5’-AS; (iv)intron retention; (v)mutually exclusive exons; (vi)alternative first exons; (vii)alternative last exons .
A number of AS databases have been constructed so far and are available in free by internet. However, most of them were developed to identify AS events based on either automated large-scale comparisons of expressed sequence tags (ESTs) that are taken from publicly available databanks, such as GenBank, EMBL,DDBJ,or mining the experimental databases. Splice Aid-F was established by screening literatures and was the only database describing interactions of human splicing factors and their RNA-binding sites so far .
An integrated comprehension of SF, RNA elements, transcription factors (TF) and other molecules such as kinases and histone modifications so on, and of their interaction network implicated in pre-mRNA splicing, is pivotal for deciphering the splicing process on genome-wide scale.There is an increasing need to create value-added databaseson biomolecular interactionsimplicated with AS regulation. The investigation of biomolecular interactions is a difficult task due to a lot of important information isscattered in the literatures. At present, none of these databases were specially builtto efficiently store interaction information among SF, TF, and RNA elements etc involved in alternative splicing regulation although a large amount of interactions among biomoleculeshave been identified through experimental analysis, especially in human. By manually screenings of literatures, we retrieved experimentally validated interaction data regulatinghumanAS events and assembled them into an online database called “database of biomolecular interaction associated with alternative splicing (BiasDB) (unpublished)”.
Bias DB collected much interaction information of AS participators and have many potential applications. By querying the interactions of SF-SF, SF-RNA, SF-TF and SF-kinase etc., an integrating network model describing regulating pathway of AS can be constructed. Some regulated models of AS occurred in special gene such as Bcl-x, c-src, CD44, CD45, CFTR, Fas, FGFR2, FN1, INSR, NF1, SMN, Tau also be presented in the current version of BiasDB. User can observe the regulated models by searching given gene name. Here, the regulated model involved with Bcl-x was shown as example to elucidate the application of BiasDB. Others results can be queried by search gene name in BiasDB. Bcl-x is a member of the Bcl-2 family proteins that are key regulators of apoptosis. By mean of alternative 5’splice site utilization, Bcl-x can produce Bcl-xS and Bcl-xL isoforms with promotion and the prevention of apoptosis.SB1, a 361nt regulatory element, locate in the first half of exon2. Ceramide-responsive RNA cis-element 1 (CRCE 1) is a purine-rich cis-element located in exon2 of Bcl-x premRNA 277-295nt upstream from intron2.Exonic splicing enhancer CGGGCAlocated 64 to 69nt upstream of the Bcl-xS 5’ss within exon2. The E2d element locates about 30bp upstream 5’splice site of Bcl-xS. The 77nt region B2 is located immediately downstream of the 5’splice site of Bcl-xS, the 30-nucleotide G-rich element located between positions +10 and +39 (B2G). A 86nt region B3 locatedimmediately upstream of the Bcl-xL donor site is composed of AM1, AM2, ML1, ML2. AM2.
SB1 behaves as a splicing silencer, and its deletion stimulates the use of theBcl-xS.By stimulating thetranscriptional elongation of the Bcl-x gene, TCERG1would reduce the time for the assembly and activity of a functional repressor at SB1, thus favoring the production of the Bcl-xS isoform . Splicing factor SAP155 regulates the alternative 5’ splice site selection of Bcl-x by binding to the CRCE 1. Up-regulation of SAP155 induced a decrease in theBcl-xS with a concomitant increase in the Bcl-xL splice variant . Bcl-xS 5’ss selection was enhancedby RBM25 specifically bound ESE CGGGCA. The binding of RBM25 was shown to promote the recruitment of the U1snRNP to the weak 5’ssthrough interaction with hLuc7A .Sam68 did not interact with SAP155 andhnRNP F/H or with ASF/SF2.HnRNP A1and Sam68 cooperate to modulate the alternative splicing of Bcl-x. Depletion of Sam68 caused accumulation of Bcl-xL, whereas its up-regulation increased the levels of Bcl-xS. Tyrosine phosphorylation of Sam68 by Fyn inverted this effect and favored the Bcl-xL splice site selection . B2 favors Bcl-xS production, B2G might make the largest contribution toward enforcing the use of the Bcl-xS 5’ splice site by binding tohnRNP F and H proteins . ML2 enforceBcl-xL production by binding SRp30c factor.The B3 region also containsa silenceracross AM1 and AM2, U1 snRNP binding to this silencer represses the use of Bcl-xL . SRSF1 binds to element across ML1 and ML2, and do not interact with hnRNP F. The binding of hnRNP Ito E2d in exon 2 displaces SRSF1 from the proximal splice site and favors selection of the distal site and generate Bcl-xS . Bias DB provides an interactional dataset among participators for understanding the regulation mechanism of AS. This database is helpful for constructing regulatory network of AS andproviding a guide for experimental program of investigating the mechanism of AS (Figure 1).
Figure 1: The regulated model for Bcl-x. Boxes represent exons, separated by introns shown as lines. The cis-acting elements and trans-acting factors in exon 2 regulating 5â€™ splice site selection of Bcl-x were marked. The â€˜+â€™ denotes promoting the production of Bcl-xS and Bcl-xL. The â€˜-â€™ denotes suppressing the production of Bcl-xS and Bcl-xL. The â€˜Ã—â€™ denotes the protein factor doesnâ€™t bind to RNA sequence.
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