Splice junction consensus sequences

invariant

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221 S16 Trends in Genetics 28:147-154 (2012)
 Why must splicing be so accurate?

Coding strand

5’ splice site
2
221 S16 Hartwell et al. (2008) Fig 8.27
 3’ splice site

RNA cleavage and poly(A) addition signal - acts at a distance 3

221 S16
Alternative splicing: Fibroblast growth factor receptor

Alternatively spliced
RNA isoforms

Protein isoforms
Different forms of a protein with slightly different amino acid
sequences, but with the same activity. May be produced from
different genes, or from the same gene by alternative splicing 4
 Types of alternative splicing

alternative splice-site donor

alternative splice-site acceptor

cassette-exon inclusion or skipping

intron retention

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221 S16 Nilsen & Graveley (2010) Nature 463, 457-46
 How are splicing decisions made?

U1 small nuclear U2 small nuclear

ribonucleoprotein (snRNP) ribonucleoprotein (snRNP)

cis-acting elements
There are additional splicing regulatory elements (SREs) in the both exons and introns
that can enhance or suppress splicing to (or from) nearby acceptor (or donor) sites.
These sites are termed exon-specific enhancers (ESE) or exon-specific suppressors
(ESS) – the same nomenclature for intronic sites (ISE and ISS)….
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221 S16 Nat Rev Genetics 15: 689-701 (2014) B. Keith
 How are splicing decisions made?

trans-acting factors
Splicing regulatory proteins (there are a LOT of them!) bind splicing regulatory
elements (SREs). Some (such as SR proteins) promote splicing to/from nearby splice
junctions, others (such as hnRNPs) repress splicing. Bottom line – these are highly
regulated decisions, and the cell invests a lot of effort in getting them right.

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Nat Rev Genetics 15: 689-701 (2014)
221 S16 B. Keith
 Alternative splicing generates protein diversity

Recent studies using high-throughput sequencing indicate that >95% of

human pre-mRNAs that contain more than one exon are processed to
yield multiple mRNAs…

8 457
Nilsen & Graveley (2010) Nature 463:
221 S16
 Alternative splicing of rat a-tropomysosin gene

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221 S16
 The Encyclopedia of DNA Elements (ENCODE)

Goal: To delineate all functional elements encoded in the human genome

ENCODE Project
32 institutions - 442 people
1,649 experiments – 15 TB data

ENCODE functional element:

A discrete genome segment that encodes
• A defined product (e.g., a protein or non-coding RNA) or
• Displays a reproducible biochemical signature (e.g.,
protein binding, or a specific chromatin structure).

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221 S16 Nature 489: 57-74; 75-82; 83-90; 91-100; 101-108; 109-113 + 25 more in other journals (2012)
 The Encyclopedia of DNA Elements (ENCODE)

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221 S16 Nature 489: 57-74; 75-82; 83-90; 91-100; 101-108; 109-113 + 25 more in other journals (2012)
 The Encyclopedia of DNA Elements (ENCODE)

The genome and its transcripts

Genome size ~3.1 x 109 bp

Gene number 20,687
Protein coding genes 2.94% of genome
Small RNAs (tRNA, miRNA, snRNA, snoRNA) 8,801
lncRNAs (long noncoding) 9,640
Promoters 70,292
Enhancers 399,124
ENCODE elements Cover 80% of genome

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221 S16 Nature 489: 57-74; 75-82; 83-90; 91-100; 101-108; 109-113 + 25 more in other journals
 The Encyclopedia of DNA Elements (ENCODE)

Alternative Splicing

Genome size ~3.1 x 109 bp

Gene number 20,687
Protein coding genes 2.94% of genome
Alternatively spliced transcripts 6.3/gene

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221 S16 Nature 489: 57-74; 75-82; 83-90; 91-100; 101-108; 109-113 + 25 more in other journals (2012)
 115 total exons in pre-mRNA – only 24 retained in mature Dscam I mRNAs

20 constitutive exons (grey)

4 alternatively spliced exons (exons 4, 6, 9, 17) - 95 choices…!

24 exons; ~8 Kb

~2000 aa

38,016 possible mRNA splicing combinations!

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221 S16