Download Mutually exclusive splicing

Chapter 14 Opener
RNA Splicing (In Eukaryotes)
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Figure 14-1
Typical eukaryote gene
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Figure 14-2
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Figure 14-3
Sequences within the RNA
determine where splicing occurs
(Acceptor site)
(Donor site)
N: any base
R: purine
Y: pyrimidine
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Figure 14-4
Splicing reaction
by transesterification
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Figure 14-5
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Spliceosome machinery
-snRNA: U1, U2, U4, U5, U6
RNA splicing is performed by a large
complex called the spliceosome.
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Figure 14-6
Some RNA-RNA hybrids formed during splicing reaction:
In some cases, different snRNAs recognize the same
sequencers in pre-mRNA at different stage of splicing.
Branchpoint binding protein
(BBP)
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Figure 14-7
snRNAs: U1, U2, U4, U5,
U6
These RNAs (of 100-300
nucleotides) are
complexed with several
proteins and called
snRNPs (pronounced
“snurps”)
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(U2 auxiliary factor)
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tri-snRNP particle (U2-U5-U6)
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Table 14-1
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Spliceosome assembly (exon definition) is dynamic
and variable and
its disassembly ensures that the splicing reaction
goes only forward in the cell.
Assembly can be variable :
splicing pairing can occur before or after
tri-snRNAP(U4-U5-U5) recruitment
Dissembly is driven by one of the DEADbox helicase proteins (called Prp22)
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Figure 14-8
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Figure 14-8a
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Figure 14-8b
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Figure 14-8c
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Figure 14-9
Proposed folding
of the RNA
catalytic regions
for splicing of
pre-mRNA and
group II intron
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How does the spliceosome find the splice sites
correctly?
-One human gene of 363 exons (usually 7 to 8 exons)
-Drosophila gene of 38,000 alternative ways of splicing
-Average exon is only some 150 nucleotides long
-Whereas average intron is about 3000 nucleotides long
(Some introns can be as long as 800,000 nucleotides)
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Figure 14-10
How does the spliceosome find the
splice sites correctly?
Errors produced by mistakes in splice site selection
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Two ways in which the accuracy of splice-site
selection can be enhanced are as follows.
One way:
When 5’ splice site is encountered in the newly synthesized
RNA,
the factors that recognize that site(5’ splice site), transferred
from C-terminal tail of RNA polymerase II onto RNA.
Once in place, the 5’ splice site components are poised to
interact with those other factors that binds to the next 3’
splice site to be synthesize.
Thus correct 3’ splice site can be recognized before any
competing sites further downstream transcribed.
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Figure 14-11
Second way by SR to bind to exonic
splicing enhancers (ESEs):
SR(serine-arginine rich) proteins recruits
spliceosome components to the 5’ and 3’ splice sites
than to incorrect sites not close to exon.
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Figure 14-12
Alternative splicing by
SR (trans-splicing)
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Figure 14-13
AT-AC(minor) spliseosome
catalyzed splicing
A small group of introns is
spliced by an alternative (minor)
spliceosome composed of a
different set of snRNPs (perhaps
one in 1000 exons in human)
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Figure 14-14
Alternative splicing in the
troponin T gene
At least 40% of Drosophila and as many as 90% of
human genes undergoes alternative splicing.
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Figure 14-15
Five ways of splice an RNA
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Figure 14-16
Alternative splicing of SV40 T antigen
Splicing out intron for Large T
Splicing out intron for small t
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Figure 14-17
Mutually exclusive splicing:
Steric(입체적) hindrance(방햬)
U1-snRNP
U1-snRNP: now U2 snRNP can not bind
to branchpoint but bind to
U2 snRNP bind
first here
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Figure 14-18
Mutually exclusive splicing:
Combination
of major and
minor splice
sites
Non-sense
mediated decay
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Figure 14-19
Curious Drosophila Dscam (down syndrome cell
adhesion molecule) gene: Mutually exclusive splicing on
a grand scale (encode 38,016 protein sioforms)
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Figure 14-20
Mutually exclusive splicing of Dscam exon 6 can not be
accounted for by an standard mechanism and instead use
a novel strategy (docking site: select sequences)
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Figure 14-21
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Box 14-3-1
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Box 14-3-2
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Figure 14-22
Alternative
(대체)splicing(이
어맞추기) is
regulated by
activators and
repressors.
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Figure 14-22a
Exonic (or intronic) splicing silencers (ESS or
ISS): sites that repressor protein binds
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Figure 14-22b
Exonic (or intronic) splicing
enhancers (ESE or ISE): sites
that SR protein binds
SR protein has RMM
(RNA recognition
motif domain) and RS
domain (rich in
arginine and serine)
(SR protein)
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Figure 14-23
Two mechanisms of
silencer action
A: A1(repressor) competes
off SC35(activator) after
binding to ESS.
silencer
enhancer
Heterogeneous nuclear ribonucleoprotein (hnRNP)
B: PTB(hnRNP,
Pyrimidine binding
protein) interact with U1 at
the 5’ splicing site and then
block U1 to interact with 3’
splicing site.
So U1 pairs with exon at
downstream.
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(repressor)
Figure 14-24
Regulation of alternative splicing determines the
sex of flies (difference in ratio of activator and
repressor determine sex)
sis-a and sis-b = Sxl activators is in X chromosome
Dpn (Deadpan) = Sxl repressor is in autosome
X means X chromosome
A means autosome
Sxl: sex lethal
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Pe: promoter for establishment
Pm: promoter for maintenance
Figure 14-25
Dsx: double sex gene
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Figure 14-26
An alternative splicing switch(대체이어맞추기 전환) lies at
the heart(중추) of pluripotency(만능유도)
FOXP1 (Transcription
factor: Forkhead family
of DNA binding
protein)
FOXP1-ES
(protein encode by
17-18b-19 mRNA)
FOXP1-ES activates genes OCT4 and NANOG, etc.
Induced pluripotent stem cells (iPS cells)
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FOXP1
(protein encode by
17-18a-19 mRNA )
Shuffle: mix cards (poker game)
Exon shuffling
Exon are shuffled by recombination to produce
genes encoding new proteins.
Intron early model
Intron late model: Introns were added later in
evolution.
There is possibly another advantage afforded these
organisms: having coding sequence of genes divided
into several exons allows new genes to be created by
reshuffing exons.
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Figure 14-27
Exons encode protein
domains.
Borders between exons
and introns within a gene
coincide with boundaries
between domains.
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Figure 14-28
Genes made up of parts of other genes
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Figure 14-29
Accumulation, loss, and reshuffling of domains
during the evolution of a family of proteins
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Figure 14-30
RNA editing is another way of altering the
sequence of an mRNA
Stop codon
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Figure 14-31
RNA editing by deaminase
Inosine base pair with cytosine. So
can alter sequence of protein
ADAR (adenosine deaminase acting on RNA)
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Figure 14-32
RNA editing by guide RNA-mediated U
insertion (in trypanosome coxII gene RNA)
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Figure 14-32a
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Figure 14-32b
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Figure 14-32c
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Figure 14-33
Transport of mRNAs (active transport) out of
the nucleus through nuclear pore complex
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