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 @#$%##@?!. Rna performs many functions. Ribosomes tRNA - PowerPoint PPT PresentationTRANSCRIPT
micro RNA’s
Computational biology seminar
Ariel Jaimovich
November 17th 2005
The central dogma of biology
• This is not always the case:– First ‘life forms’– viruses
Transcription
RNA
Translation
Protein
Dioxy ribo @#$%##@?!
Rna performs many functions
• Ribosomes
• tRNA
• Nuclear detaining
• RNAi
Micro RNA
• ~22nt rna
• Precursor stem& loop
• Post-transcription regulation
miRNA History
• Lin-4 inhibits LIN14, but no LIN 4 protein was found (1993)
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
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.
Expression• Stage\tissue specific
• Large number of copies (robust transcription \ slow decay)
miRNA – biogenesis
Highly conserved in evolution
Plants Vs Animals
miRNA - function
Sequence recognition
• Positions 2-8 are most important– How do we know– Why ?
Base pairing Function
How do we know which process is active ?
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
siRNA vs miRNA
• Genomic origin –
– miRNA from genes
– siRNA from mRNAs, transposons, viruses...
• Synthesis
• One siRNA duplex many siRNA
• Conservation
miRNA vs siRNA
miRNA in plants
• Near-perfect complementarity
• mRNA cleavage, usually of TF’s related to developmental processes
• Conservation between Arabidopsis and rice
• Defend against viruses
miRNA in animals
• mRNA cleavage or translational silencing
• Conservation is also high (?)
• Different numbers of paralogs
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
Goal
Find new human micro RNAs
Motivation
Current gene search techniques:
• Hairpins
• Conservation
Try to search with a wider scope
Search algorithm
Fold the genome
~ 11 milion hairpins
Prediction (1)
Magic box
~ 430,000 hairpins
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
Prediction (2)
~ 430,000 hairpins
conservedNon- conserved
samplesample
800 clustered
3000 non-
clustered
1500 clustered
7500control
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
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
New cluster
The cluster conatin a few ‘seeds’
Results summary
Goal
ExpressionLocation on chromosome
?
Creating miRNA micro array
Design microRNA chip
• Normalization by synthetic samples
• Melting temperature
Array Results
Is Expression correlated with distance between microRNA’s?
55bp
Is expression of micro RNA’s correlated with host genes?
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
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 ?