Lecture 11
description
Transcript of Lecture 11
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Lecture 11
HW1 Feedback (ours)
(Upcoming Project – discuss Wed)
Non-Coding RNAs
Halfway Feedback (yours)
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“non coding” RNAs
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Central Dogma of Biology:
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RNA is an Active Player:
reverse transcriptionlong ncRNA
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What is ncRNA?
• Non-coding RNA (ncRNA) is an RNA that functions without being translated to a protein.
• Known roles for ncRNAs:– RNA catalyzes excision/ligation in introns.– RNA catalyzes the maturation of tRNA.– RNA catalyzes peptide bond formation.– RNA is a required subunit in telomerase.– RNA plays roles in immunity and development (RNAi).– RNA plays a role in dosage compensation.– RNA plays a role in carbon storage.– RNA is a major subunit in the SRP, which is important in protein trafficking.– RNA guides RNA modification.
– In the beginning it is thought there was an RNA World, where RNA was both the information carrier and active molecule.
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AAUUGCGGGAAAGGGGUCAACAGCCGUUCAGUACCAAGUCUCAGGGGAAACUUUGAGAUGGCCUUGCAAAGGGUAUGGUAAUAAGCUGACGGACAUGGUCCUAACCACGCAGCCAAGUCCUAAGUCAACAGAUCUUCUGUUGAUAUGGAUGCAGUUCA
RNA Folds into (Secondary and) 3D Structures
P 6b
P 6a
P 6
P 4
P 5P 5a
P 5b
P 5c
120
140
160
180
200
220
240
260
AAU
UGCGGG
A
A
A
GGGGUCA
ACAGCCG UUCAG
U
ACCA
AGUCUCAGGGG
AAACUUUGAGAU
GGCCUUGCA A A G G
G U A UGGUA
AU
A AG
CUGACGGACA
UGGUCC
U
A
A
CCA CGCA
GC
CAA
GUCC
UAA
GUCAACAG
AU C U
UCUGUUGAU
A
UGGAU
GC
AGU
UC A
Cate, et al. (Cech & Doudna).(1996) Science 273:1678.
Waring & Davies. (1984) Gene 28: 277.
We would like to predict them from sequence.
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RNA structure rules• Canonical basepairs:
– Watson-Crick basepairs:• G - C• A - U
– Wobble basepair:• G – U
• Stacks: continuous nested basepairs. (energetically favorable)
• Non-basepaired loops:
– Hairpin loop.
– Bulge.
– Internal loop.
– Multiloop.
• Pseudo-knots
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Bafna 1
RNA structure: Basics
• Key: RNA is single-stranded. Think of a string over 4 letters, AC,G, and U.
• The complementary bases form pairs.• Base-pairing defines a secondary structure.
The base-pairing is usually non-crossing.
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Ab initio structure prediction: lots of Dynamic Programming
• Maximizing the number of base pairs (Nussinov et al, 1978)
simple model:(i, j) = 1
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Pseudoknots drastically increase computational complexity
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Nearest Neighbor Model for RNA Secondary Structure Free Energy at 37 OC:
C G U U U G G GUU
CACAAACG
-2 .0
-2 .1
-0 .9
-0 .9
-1 .8
-1 .6
+ 5 .0
Ghelix = GCGGC + GGUCA + 2GUUAA + GUGAC =
-2.0 kcal/mol - 2.1 kcal/mol + 2x(-0.9) kcal/mol - 1.8 kcal/mol = -7.7 kcal/mol
Ghairpin loop = Ginitiation (6 nucleotides) + GmismatchGGCA =
5.0 kcal/mol - 1.6 kcal/mol = 3.4 kcal/mol
Gtotal = G
hairpin + Ghelix = 3.4 kcal/mol - 7.7 kcal/mol = -4.3 kcal/mol
Mathews, Disney, Childs, Schroeder, Zuker, & Turner. 2004. PNAS 101: 7287.
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Zuker’s algorithm MFOLD: computing loop dependent energies
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Energy Landscape of Real & Inferred Structures
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Unfortunately…
– Random DNA (with high GC content) often folds into low-energy structures.
– What other signals determine non-coding genes?
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Evolution to the Rescue
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a a cg u u c c c cu c ua g a cc
S
S
S
S
S aSu L aL
S uSa L cL
S gSc L a
S cSg L c
S L
• Each derivation tree corresponds to a structure.
Stochastic context-free grammar (SCFG)
L
L
L
L
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S aSu
S cSg
S gSc
S uSa
S a
S c
S g
S u
S SS
1. A CFG
S aSu
acSgu
accSggu
accuSaggu
accuSSaggu
accugScSaggu
accuggSccSaggu
accuggaccSaggu
accuggacccSgaggu
accuggacccuSagaggu
accuggacccuuagaggu
2. A derivation of “accuggacccuuagaggu” 3. Corresponding structure
Stochastic context-free grammar (cont’)
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MicroRNA
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Genomic context
known miRNAs in human
intergenic intronic
polycistronic
monocistronic
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tRNA
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tRNA Activity
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Human specific accelerated evolution
Chimp
Humanrapid change
conserved
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Human Accelerated RegionsHuman-specific substitutions in conserved sequences
28[Pollard, K. et al., Nature, 2006] [Beniaminov, A. et al., RNA, 2008]
HumanDerived
Chimp
Humanrapid change
HAR1:• Novel ncRNA•Co-expressed in Cajal-Retzius cells with reelin.•Similar expression inhuman, chimp, rhesus.•18 unique human substitutionsleading to novel conformation.•All weak (AT) to strong (GC).
conserved
ChimpAncestral
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Other Non Coding Transcripts
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mRNA
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EST
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lincRNAs (long intergenic non coding RNAs)
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X chromosome inactivation in mammals
X X X Y
X
Dosage compensation
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Xist – X inactive-specific transcript
Avner and Heard, Nat. Rev. Genetics 2001 2(1):59-67
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Microarrays, Next Gen(eration) Sequencing etc.
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End Results
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Transcripts, transcripts everywhere
Human Genome
Transcribed (Tx)
Tx from both strands
Leaky tx?
Functional?
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Or are they?
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Halfway Feedback