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Inference of gene expression regulation by miRNA using MiRaGE method
Yh. Taguchi/Dept.Phys.,Chuo Univ.Jun Yasuda /School Med.,Tohoku Uinv.
Inference of transfection of miRNA to human lung cancer cell
Three Topics:Three Topics:
Inference of gene regulation via miRNA in murine medulloblastoma
Identification of critical miRNAs for ES stemness during differentiation to neuronal cells
3
Target genes
Computer oriented prediction (uncertain)
genome
microRNA
microRNA
mRNA
mRNA
Regulation of gene expression via miRNARegulation of gene expression via miRNA
4
miRNA1 ○×○○○○××miRNA2 ○×○○××○○miRNA3 ×○○×○○××miRNA4 ○○○×○○××
Gene1
Gene2
Gene3
Gene4
Gene5
Gene6
Gene7
Gene8
miRNA target gene list simple seed match(Virtual)
VS
Human lung cancerHuman lung cancerMurine medulloblastomaMurine medulloblastoma
Murine ES cellMurine ES cell
RealRealgene1gene2gene3
Prediction
miRNA1
5
Control Treated
Gather the information of miRNA targets
Compare the expressions of targets for each miRNAs (see Next Slides)
Calculate False Discovery Rate
Generate ranking
6
miRNA Targets Down Pvalue FDRmiRa 54 3 0.5 0.4miRb 120 54 0.0001 0.005miRc 36 1 0.5 0.7... ... ... ... ...miRX 60 18 0.001 0.007
Reject miRa & c because the FDR > 0.05
Filtrate with miRNA expression profiles
Ranking
MiRaGMiRaGEE
Inference of transfection of miRNA to human lung cancer cell
Topics 1Topics 1
Gene expression byArray: AgilentatOne and three days after transfection ofMir107, mir185, and let7a.
log(xg[miRNA]) vs log(xg[Control]) xg: gene expression
Target gene list: simple seed match
ICIC2011, LNBI in press. (2011/8/1114)
Transfected miRNAmir107 mir185 let7a
time replicate 1 replicate 2 replicate 1 replicate 2 replicate 1 replicate 2
day1 1[1st] 1[1st] 7[1st] 9[1st] 2[1st] 2[1st]day3 1[1st] 0[] 0[1st] 0[] 1[1st] 1[1st]Numbers of significant miRNAs. The ranks of transfected micriRNAs are shown in square brackets.
Transfected miRNAs are correctly identified by MiRaGE method.
Topics 2Topics 2
Inference of gene regulation via miRNA in murine medulloblastoma
1 1
Materials(established at Tastuo Noda group)
P6P6=6 days after birth, normal but growingP30P30=30 days after birth, normal and not growingMBMB=a few month after birth, malignant neoplasm30% of the Ptc1 +/ mice suffers from MB.
12
mRNA/miRNA expression byArray: AgilentatP6, P30 and MB
log(xg[mRNA/miRNA:MB or P6]) vs
log(xg[mRNA/miRNA:P30])
xg: mRNA/miRNA expression
Target gene list: simple seed match
13
log(xg[miRNA:P6/MB]) vs log(miRNA:xg[P30]) of considered miRNA(*)
(*) each miRNA is measured by multiple probe
t test for miRNA expression
14
log(xg[mRNA:P6/MB]) – log(xg[mRNA:P30]) in target genes of considered miRNA
t test for miRNA target genes (MiRaGE method)
log(xg[mRNA:P6/MB]) – log(xg[mRNA:P30]) in target genes of
any of other miRNA
VS
15
1 mmu-miR-25mmu-miR-25 1 1
2 mmu-miR-466i-5p 1 1
3 mmu-miR-92ammu-miR-92a 0.75 1
4 mmu-miR-19ammu-miR-19a 1 0.69
5 mmu-miR-19bmmu-miR-19b 1 0.69
6 mmu-miR-3082-5p 1 0.56
7 mmu-miR-130a 1 0.5
8 mmu-miR-130b 1 0.5
9 mmu-miR-15b 1 0.5
10 mmu-miR-2861 1 0.5
11 mmu-miR-3096-5p 1 0.5
12 mmu-miR-32 0.5 1
13 mmu-miR-322 1 0.5
14 mmu-miR-721 1 0.5
15 mmu-miR-149* 0.5 0.88
16 mmu-miR-3081* 1 0.38
17 mmu-miR-574-5p 1 0.31
18 mmu-miR-669n 0.5 0.81
19 mmu-miR-1187 1 0.25
20 mmu-miR-182mmu-miR-182 0.5 0.75
miRNA P30<<MBtarget gene P30>>MB
miR17~92 cluster family miR17~92 cluster family membersmembers are ranked in top 5 by combination of MiRaGE methods and miRNA expression profiling.
selected bymiRNA miRNA expression / MiRaGE
suggested contribution to suggested contribution to cancer formation cancer formation
16
mmu-miR-100 1 1
mmu-miR-126-3p 1 1
mmu-miR-29cmmu-miR-29c 1 1
mmu-miR-376ammu-miR-376a 1 1
mmu-miR-451 1 1
mmu-miR-99b 1 1
mmu-miR-136* 1 0.9375
mmu-miR-299* 0.75 1
mmu-miR-26ammu-miR-26a 1 0.5
mmu-miR-26bmmu-miR-26b 1 0.5
mmu-miR-29ammu-miR-29a 0.5 1
mmu-miR-7a-1*mmu-miR-7a-1* 1 0.5
mmu-miR-3107 1 0.4375
mmu-miR-340-5p 1 0.3125
mmu-miR-369-5p 1 0.3125
mmu-let-7ammu-let-7a 1 0.25
mmu-let-7emmu-let-7e 1 0.25
mmu-let-7gmmu-let-7g 1 0.25
mmu-let-7immu-let-7i 1 0.25
mmu-miR-467b 0.25 1
selected bymiRNA miRNA expression / MiRaGE
tumorsuppressive miRNAs tumorsuppressive miRNAs
neuronspecific miRNAsneuronspecific miRNAs
miRNA P30>>MBtarget gene P30<<MB
Some of the neuronneuronspecific miRNAspecific miRNAs and tumorsuppressive tumorsuppressive miRNAsmiRNAs seem to contribute to the gene expression profiles of P30.
17
miRNA P30<<P6target gene P30>>P6
miR17~92, miR17~92, mir106bmir106b2525,,mir106a363mir106a363cluster family members are ranked in top 5 by combination of MiRaGE methods and miRNA expression profiling.
selected bymiRNA miRNA expression / MiRaGE
1 mmu-miR-106bmmu-miR-106b 1.00 1.00
2 mmu-miR-130a 1.00 1.00
3 mmu-miR-130b 1.00 1.00
4 mmu-miR-15b 1.00 1.00
5 mmu-miR-17mmu-miR-17 1.00 1.00
6 mmu-miR-20ammu-miR-20a 1.00 1.00
7 mmu-miR-20bmmu-miR-20b 1.00 1.00
8 mmu-miR-301b 1.00 1.00
9 mmu-miR-322 1.00 1.00
10 mmu-miR-721 1.00 1.00
11 mmu-miR-93mmu-miR-93 1.00 1.00
12 mmu-miR-542-3p 1.00 0.94
13 mmu-miR-3081* 1.00 0.88
14 mmu-miR-335-3p 1.00 0.88
15 mmu-miR-199a-5p 1.00 0.81
16 mmu-miR-199b* 1.00 0.81
17 mmu-miR-19ammu-miR-19a 1.00 0.81
18 mmmmu-u-mmiiRR--1199bb 1.00 0.81
19 mmu-miR-148a 0.75 0.94
20 mmu-miR-214 1.00 0.63
18
selected bymiRNA miRNA expression / MiRaGE
tumorsuppressive miRNAs tumorsuppressive miRNAs
neuronspecific miRNAsneuronspecific miRNAs
miRNA P30>>P6target gene P30<<P6
Some of the neuronneuronspecific miRNAspecific miRNAs and tumorsuppressive tumorsuppressive miRNAsmiRNAs seem to contribute to the gene expression profiles of P30.
mmu-miR-29c 1.00 1.00
mmu-miR-376ammu-miR-376a 1.00 1.00
mmu-miR-451 1.00 1.00
mmu-let-7bmmu-let-7b 1.00 0.94
mmu-let-7emmu-let-7e 1.00 0.94
mmu-let-7gmmu-let-7g 1.00 0.94
mmu-let-7immu-let-7i 1.00 0.94
mmu-miR-98 1.00 0.94
mmu-miR-126-3p 0.75 1.00
mmu-miR-299* 0.75 1.00
mmu-miR-29a 0.75 1.00
mmu-let-7ammu-let-7a 0.75 0.94
mmu-miR-3070b-3p 1.00 0.69
mmu-miR-138 1.00 0.63
mmu-miR-3107 1.00 0.56
mmu-miR-181a-1* 0.50 1.00
mmu-let-7dmmu-let-7d 0.50 0.94
mmu-miR-1937b 0.25 1.00
mmu-miR-1937c 0.25 1.00
mmu-miR-337-5p 1.00 0.25
19
MiRaGE method + miRNA expression successfully pick up biologically important miRNAs. Further (wet) experiments which supress miRNA expression with tiny LNA is now planed.
If it is successful, our method can find miRNAs which control tumor formation.
Published in IPSJ Technical report, 2011SIGBIO25, No5, pp.16
Topics 3Topics 3
Identification of critical miRNAs for ES stemness during differentiation to neuronal cells
Materials:
Gene expressionGene expression data is downloaded from GEO by the accession number GSE11523, “Defining Developmental Potency and Cell Lineage Trajectories by Expression Profiling of Differentiating Mouse ES Cells”. In this study, among those, data for the differentiation from ES cells to neuronal cells are used.
In parallel, we analyzed the difference of miRNA miRNA expressionexpression profiles between 2 datasets from ES cells and 6 datasets from mature neuronal tissues(*) and listed up the ES cell specific miRNAs.
*) http://www.mirz.unibas.ch/cloningprofiles/
miRNAs ratio
mmu-miR-200bmmu-miR-200b 1.00
mmu-miR-200c (+)mmu-miR-200c (+) 1.00
mmu-miR-23a 1.00
mmu-miR-23b 1.00
mmu-miR-291a-3pmmu-miR-291a-3p 1.00
mmu-miR-429mmu-miR-429 1.00
mmu-miR-294mmu-miR-294 0.98
mmu-miR-295mmu-miR-295 0.98
mmu-miR-302a (+)mmu-miR-302a (+) 0.98
mmu-miR-302b (+)mmu-miR-302b (+) 0.97
mmu-miR-302d (+)mmu-miR-302d (+) 0.97
mmu-miR-199a-5p 0.94
mmu-miR-141mmu-miR-141 0.69
mmu-miR-200ammu-miR-200a 0.69
mmu-miR-409-3p 0.67
mmu-miR-369-3p (+) 0.36
mmu-miR-96 0.11
mmu-miR-674 0.08
mmu-miR-467b 0.06Mallanna, S.K., and Rizzino., A.(2010)
miRNA ES> > Neuronal Celltarget gene ES< < Neuronal Cell
Our method identified critical miRNAs for ES stemness including iPS inducing miRNAs (marked with “+”in the previous slide) recently reported by Miyoshi, et al.(2011, Cell Stem Cell).
Published in IPSJ Technical report, 2011BIO25, No.38, pp.1 2
miRNA is short noncoding RNA which regulate many biological processes ranging from cancer formation to differentiation.We have proposed a new method, MiRaGE, MiRNA Ranking by Gene Expression, which assists to discriminate which miRNA really regulate target genes.Since there are some technical method which suppress miRNA machinery, it can also be a drug candidates related to several biological processes.
Acknowledgements:Acknowledgements: We thank Drs. Tetsuo Noda and We thank Drs. Tetsuo Noda and
Katsuyuki Yaginuma for Katsuyuki Yaginuma for providing reagents (Topics 2).providing reagents (Topics 2).
These works were supported by These works were supported by KAKENHI (23300357) .KAKENHI (23300357) .
Server will be soon available at:Server will be soon available at:
http://www.granular.com/MiRaGE.htmlhttp://www.granular.com/MiRaGE.html
e-mail: [email protected]: [email protected]