Yi Xing's Alternative Splicing and Computational Genomics Blog

Human Genome Calendar

A very good collection of conferences related to genomics and bioinformatics.
Maintained by Oak ridge National Lab.

Click here.

U1/U2 splice sites weblogo

Trained from ~90000 constitutive splices in human. Logos are generated by WebLogo program.

You can use them in presentations if you want to show how splice sites look like. :)

Alternative splicing and mRNA nonsense mediated decay (I)

Nonsense mediated decay(NMD) is a mRNA surveillance mechanism to degrade aberrant transcripts that contain premature termination codons (PTCs). A premature termination codon is defined as a STOP codon located more than 50-55bp upstream from the last exon-exon junction. PTCs can be recognized by NMD, resulting in the degradation of the transcripts.

Steven Brenner and colleagues made the first genome-wide analysis on the relationship of pre-mRNA splicing and NMD. They found that alternatively spliced transcript isoforms frequently contained PTCs.

Evidence for the widespread coupling of alternative splicing and nonsense-mediated mRNA decay in humans. Lewis BP, Green RE, Brenner SE. Proc Natl Acad Sci U S A. 2003 Jan 7;100(1):189-92. Epub 2002 Dec 26.

This analysis showed that many alternatively spliced forms might be the result of aberrant splicing which will be cleared by NMD. Beyond that, Brenner and colleagues also advanced the interesting hypothesis that the coupling of alternative splicing and NMD might constitute a functional regulation of gene expression, i.e. using splicing and NMD to control the amount of productive transcripts being made. This hypothesis is supported by several lines of experimental evidence, such as the alternative splicing of PTB.

Comparative genomic studies of splicing regulatory elements

A new paper from Chris Burge lab, published at PNAS.

Variation in sequence and organization of splicing regulatory elements in vertebrate genes Gene Yeo, Shawn Hoon , Byrappa Venkatesh , and Christopher B. Burge

My GRC 2004 Poster on Splicing Graphs and Isoform Construction

Here is the poster I showed at Gordon Research Conference on Structural, Functional and Evolutionary Genomics (2004) at Ventura, CA.
It's about constructing multiple transcript isoforms from ESTs.
Download file

Out of town for NIH Sackler Colloquia at Irvine

Here is the program.

Will be back on Sunday. There will be a session for alternative splicing, featuring talks by Chris Burge (MIT) and Chris Lee (UCLA, my boss :) ).

I'll present a poster regarding alternative splicing and genome evolution.

Evolutionary convergence of alternative splicing in ion channels

This is an interesting paper about adaptive benefits of alternative splicing.
I'll write more about it later.

Evolutionary convergence of alternative splicing in ion channels. Copley RR. Trends Genet. 2004 Apr;20(4):171-6.

Alternative splicing and genome complexity: Do humans have more alternative splicing?

This is an interesting and controversial topic. The unexpectedly low number of genes in the human genome (~32000, which is roughly twice the number for Drosophila and worm) raised again the long-standing question in biology:

what is the source for organism complexity?

After it was discovered that alternative splicing, a mechanism for increasing transcript diversity, is actually widespread in the human genome, it appeared reasonable to think that alternative splicing might be one of the major contributors for human genome complexity.

Surprisingly, work from Peer Bork lab argued against this hypothesis. They found that after correcting for EST coverage, the frequency of alternative splicing seemed to be comparable among animals , and humans didn't appear to have a substantially higher rate of alternative splicing:

Alternative splicing and genome complexity. Brett D, Pospisil H, Valcarcel J, Reich J, Bork P. Nat Genet. 2002 Jan;30(1):29-30.

There is certain amount of doubt concerning this interesting and apparently counter-intuitive conclusion. In a recent correspondence published on Nature Genetics Kim H et.al. used an interesting but indirect method to re-evaluate this question. By estimating the number of total transcripts in different genomes, they suggested that human actually has more alternative splicing than mouse and fruitfly.

However, although Kim H et.al. claimed that their method is independent on EST coverage, in their reply Bork P and colleagues showed that the method used by Kim H et.al. was still dependent on the amount of EST coverage. For example, their method showed that there is substantially lower rate of alternative splicing in rat than mouse.

I believe there will still be more debates regarding this interesting question. No matter who is right ( although I am leaning towards the conclusion by Brett et.al., partly because I am amazed by DSCAM), I totally agree with the following argument put forward by Bork and colleagues:

"...no estimate should be considered absolute when extrapolating from data with many hidden biases...the art is to avoid hidden biases in derived data and their processing as much as possible."

Alternative splicing papers in Oct, 2004

This entry will be continuously updated till the end of Oct, 2004.
I would only list genome-scale studies of alternative splicing. Papers regarding AS of single genes won't be listed here.

Single nucleotide polymorphism-based validation of exonic splicing enhancers. Fairbrother WG, Holste D, Burge CB, Sharp PA. PLoS Biol. 2004 Sep;2(9):E268.

Genome-wide analysis of alternative pre-mRNA splicing in Arabidopsis thaliana based on full-length cDNA sequences. 1: Nucleic Acids Res. 2004 Sep 27;32(17):5096-103. Iida K, Seki M, Sakurai T, Satou M, Akiyama K, Toyoda T, Konagaya A, Shinozaki K.

How did alternative splicing evolve? Nature Reviews Genetics 5,773-782(2004). Ast G.

Alternative Splice Variants Encoding Unstable Protein Domains Exist in the Human Brain. J Mol Biol. 2004 Nov 5;343(5):1207-1220. Homma K, Kikuno RF, Nagase T, Ohara O, Nishikawa K.

Open thread for literature posting

Please post relevant articles in the comment.

Bioinformatics links

University of British, Columbia (CAN) maintains a good bioinformatics directory.
Click to enter.

Turning numbers into real biology: tissue specificity of alternative splicing

With large number of alternative splicing events detected from human and other genomes, one immediate question is: are these alternative splicing events regulated?

Here is a list of papers on tissue-specificity of alternative splicing:

Genome-wide detection of tissue-specific alternative splicing in the human transcriptome. Xu Q, Modrek B, Lee C. Nucleic Acids Res. 2002 Sep 1;30(17):3754-66.

Variation in alternative splicing across human tissues.Yeo G, Holste D, Kreiman G, Burge CB. Genome Biol. 2004;5(10):R74.

Strengths and weaknesses of EST-based prediction of tissue-specific alternative splicing. Gupta S, Zink D, Korn B, Vingron M, Haas SA.
BMC Genomics. 2004 Sep 28;5(1):72.

The basic idea for computational identification of tissue-specific alternative splicing, is to count number of EST evidences for each splice variants from different tissues, and look for evidence of tissue-specificity.

For example, Xu et.al. used the following strategy: for each pairs of alternative splices(S1,S2), and a given tissue(T), they counted the number of ESTs in tissue T which support either S1 or S2(N1,N2), as well as the number of ESTs which support either S1 or S2 in all other tissues(N1',N2'). Then they tested whether the frequency of splice S1 in tissue T was significantly greater than the frequency of splice S2 in tissue T, using bayesian inference.

Here is an Excel spreadsheet I made for a bioinformatics class at UCLA, which demonstrates the computation of Xu et.al. . Click to download.

Basic types of alternative splicing

Click to view PDF

The first wave: bioinformatic discovery of alternative splicing from expressed sequences

Alternative splicing is one of the fields in molecular biology that demonstrate bioinformatics can play a leading role in scientific discoveries. It's the bioinformatics analysis of ESTs data in late 90s which showed the striking abundance of alternative splicing in human and most eukaryotic genomes. This stimulated many experimental investigations. Those early bioinformatics work included:

Frequent alternative splicing of human genes. Genome Res. 1999 Dec;9(12):1288-93. Mironov AA, Fickett JW, Gelfand MS.
EST comparison indicates 38% of human mRNAs contain possible alternative splice forms. FEBS Lett. 2000 May 26;474(1):83-6. Brett D, Hanke J, Lehmann G, Haase S, Delbruck S, Krueger S, Reich J, Bork P.
ISIS, the intron information system, reveals the high frequency of alternative splicing in the human genome. Nat Genet. 2000 Apr;24(4):340-1. Croft L, Schandorff S, Clark F, Burrage K, Arctander P, Mattick JS.
Gene structure prediction and alternative splicing analysis using genomically aligned ESTs. Genome Res. 2001 May;11(5):889-900. Kan Z, Rouchka EC, Gish WR, States DJ.
Genome-wide detection of alternative splicing in expressed sequences of human genes. Nucleic Acids Res. 2001 Jul 1;29(13):2850-9. Modrek B, Resch A, Grasso C, Lee C.

These work used ESTs, or Expressed Sequence Tags, to discover alternative splicing. ESTs are fragments of the full length transcripts. The extensive EST sequenceing effort in the 1990s generated large amount of EST data. These EST sequences represent "snapshots" of the transcripts in different tissues, cell types and developmental stages, and are tremendously useful for detecting transcript variations such as Single Nucleotide Polymorphisms (SNPs) and alternative splicing.

This picture is taken from the review by Modrek B and Lee C, which illustrates how to detect alternative splicing using EST data.

A picture illustrating alternative splicing

I think it would be nice to have a picture showing what's alternative splicing.

Below is a picture taken from the homepage of Steven Brenner , who has done very interesting work on alternative splicing and mRNA nonsense mediated decay.

Recommended review articles on alternative splicing

Since some people were asking for it, I am providing a partial list of good review articles on alternative splicing. These articles are all from easily-accessible journals. I'll put a PubMed link for each article.

A genomic view of alternative splicing , by Christopher Lee and colleagues from UCLA. This is a very nice review on bioinformatic studies of alternative splicing.

The evolving roles of alternative splicing , by Steven Brenner and colleagues from UC Berkeley. This is a comprehensive review which summarizes the latest development in different fields of genomic studies of alternative splicing, including proteomic impact, genome evolution, NMD, etc.

Mechanisms of alternative pre-messenger RNA splicing , by Douglas Black from UCLA. This is a must-read for people who are interested in the mechanisms of alternative splicing.

Listening to silence and understanding nonsense: exonic mutations that affect splicing , by Adrian Krainer and colleagues from Cold Spring Harbor Laboratory. This is a nice review discussing the relationship of SNP and splicing.

The origins and implications of Aluternative splicing , by Brendon Graveley and colleagues from University of Connecticut. This article discusses alternative splicing events derived from Alu elements.

Update Pre-mRNA splicing and human disease. Faustino NA, Cooper TA. Genes Dev. 2003 Feb 15;17(4):419-37.

P.S. : please write to me or comment if you know of any other good reviews on this subject.

Intro: what is alternative splicing?

Alternative splicing refers to the production of multiple transcript isoforms from a single gene, due to variations in the splicing reaction of pre-mRNA. Most genes in eukaryotic genomes consist of exons and introns. After transcription, introns need to be removed from the pre-mRNA by a step called splicing. However, sometimes an exon can be either included or excluded from the final transcripts. Or there can be two splice sites at one end of an exon which can be recognized by the spliceosome ( the complex which carries splicing reaction). All these will lead to the production of multiple transcripts and are termed as "alternative splicing".

Alternative splicing has long been regarded as a rather rare event in eukaryotic genomes. For example, it was estimated to occur in less than 5% of human genes. However, recent genomic and bioinformatic analyses of vast amount of transcript data in human and other organisms suggest that alternative splicing is widespread in mammalian genomes. In human, it is estimated that alternative splicing occurs in more than 60% of genes. The "one gene, one protein" central dogma, a governing theory of modern molecular biology which has profound impact on biologists' thinking, therefore has to be revisited. As a result, our view towards many biological processes, such as protein interaction, gene expression, has to be adjusted.

Understanding the functional impact of alternative splicing, the regulatory mechanisms that govern RNA splicing, and the role of alternative splicing in genome evolution, has become one of the most challenging and exciting tasks for genomics and bioinformatics in the post-genomic era.

For a vivid illustration about alternative splicing, please check the animation from Exonhit.

Greetings

Hi, welcome to my alternative splicing blog!

This blog will be committed to discussing latest progress in genomics and bioinformatics of RNA alternative splicing.

I am a PhD student at University of California, Los Angeles. I am working with Dr. Christopher Lee at Molecular Biology Institute, UCLA.

For people who wants to know more about me, please go to my homepage .