micro rna’s
DESCRIPTION
micro RNA’s. Computational biology seminar Ariel Jaimovich November 17 th 2005. Transcription. Translation. Protein. RNA. The central dogma of biology. This is not always the case: First ‘life forms’ viruses. Dioxy ribo @#$%##@?!. Rna performs many functions. Ribosomes tRNA - PowerPoint PPT PresentationTRANSCRIPT
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micro RNA’s
Computational biology seminar
Ariel Jaimovich
November 17th 2005
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The central dogma of biology
• This is not always the case:– First ‘life forms’– viruses
Transcription
RNA
Translation
Protein
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Dioxy ribo @#$%##@?!
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Rna performs many functions
• Ribosomes
• tRNA
• Nuclear detaining
• RNAi
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Micro RNA
• ~22nt rna
• Precursor stem& loop
• Post-transcription regulation
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miRNA History
• Lin-4 inhibits LIN14, but no LIN 4 protein was found (1993)
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miRNA appear in many organisms
• Highly conserved, many ‘copies’ in each organism4 paralogs of let7 4 in c elegans
15 in human
1 in drosophila
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miRNA Genes
• ~1/3 Reside inside introns
• ~ 2/3 independent transcription units
• Often in clusters.
• Many times near the genes they regulate or inside them.
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Expression• Stage\tissue specific
• Large number of copies (robust transcription \ slow decay)
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miRNA – biogenesis
Highly conserved in evolution
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Plants Vs Animals
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miRNA - function
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Sequence recognition
• Positions 2-8 are most important– How do we know– Why ?
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Base pairing Function
How do we know which process is active ?
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Protein coding rna
3’ utr
Function (cont)• Plants vs animals
• Number of target seq. on 3’ utr ?
• Some miRNA target the same mRNA in different sites
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siRNA vs miRNA
• Genomic origin –
– miRNA from genes
– siRNA from mRNAs, transposons, viruses...
• Synthesis
• One siRNA duplex many siRNA
• Conservation
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miRNA vs siRNA
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miRNA in plants
• Near-perfect complementarity
• mRNA cleavage, usually of TF’s related to developmental processes
• Conservation between Arabidopsis and rice
• Defend against viruses
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miRNA in animals
• mRNA cleavage or translational silencing
• Conservation is also high (?)
• Different numbers of paralogs
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Identification of hundreds of conserved and nonconserved human microRNAs
Isaac Bentwich, Amir Avniel, Yael Karov, Ranit Aharonov Shlomit Gilad, Omer Barad, Adi Barzilai, Paz Einat, Uri Einav, Eti Meiri,
Eilon Sharon, Yael Spector & Zvi Bentwich
Nature genetics - June 2005
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Goal
Find new human micro RNAs
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Motivation
Current gene search techniques:
• Hairpins
• Conservation
Try to search with a wider scope
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Search algorithm
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Fold the genome
~ 11 milion hairpins
Prediction (1)
Magic box
~ 430,000 hairpins
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Structure features•Hairpin length•Loop length•Stability score•Free energy per nucleotide•Matching pairs•Bulge size
Magic box
Sequence features•Sequence repetitiveness•Regular internal repeat•Inverted internal repeat
Build a classifier
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Prediction (2)
~ 430,000 hairpins
conservedNon- conserved
samplesample
800 clustered
3000 non-
clustered
1500 clustered
7500control
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Prediction (3)800
clustered 3000 non-
clustered
1500 clustered
7500control
Micro array in five tissue cultures
886 confirmed miRNA
Sample359 miRNA
Validate (clone and sequence)
69 ‘adjacent’ miRNA
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Prediction (3)
359 miRNA
Validate (clone and sequence)
69 ‘adjacent’ miRNA
89 (33 ‘adjacent’) cloned and sequenced NEW miRNA
Of these:
•1 from the control list
•36 conserved miRNA’s (32 validated in other experiments)
•43 in two new clusters
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New cluster
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The cluster conatin a few ‘seeds’
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Results summary
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Goal
ExpressionLocation on chromosome
?
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Creating miRNA micro array
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Design microRNA chip
• Normalization by synthetic samples
• Melting temperature
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Array Results
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Is Expression correlated with distance between microRNA’s?
55bp
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Is expression of micro RNA’s correlated with host genes?
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Caveats
• Numbers of pairs ?
• Quantitative comparison with host genes
Conclusions• Some miRNA are arranged in genes
• miRNA that are located inside introns are expressed similarly to their hosts
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Points for thought
• Is miRNA regulated ? On which levels ?
• Is there a regulation on the RISC ‘loading’
• Why is so many annotated miRNA related to differentiation ?– mRNA can be passed on during mitosis and
need to cleaved
• Control leaky transcription ?