Inference of target gene regulation via miRNAs during cell senescence by MiRaGE Server
Inference of target gene regulation viamiRNAs during cell senescence by MiRaGE Server Y-h. Taguchi Department of Physics Chuo University
1. What is cell senescence?2. What is miRNA?3. Previous works (wet)4. MiRaGE Method5. Correlation between target gene regulationby miRNA and miRNA expression changeduring cell senescence6. Summary & Conclusion
1. What is cell senescence?The cell division cannot continue foreverfor incubated cell lines. It must stop afterseveral rounds of proliferation. ⇓This is called “cell senescence”, which isbelieved to be related to aging. agingThus, cell senescence is caused by theinterruption of cell divisions, typicallyby cell cycle arrests.
2. What is miRNA?miRNA is a kind of non-coding RNA.miRNAs are believed to suppress target geneexpression by degradation of mRNAs.Important features:・ Typically, there are hundreds kinds of miRNAs foundfor each species (c.a., ≧1000 for human).・ Each miRNA targets more than hundreds of genes.・ miRNA mainly contributes to cell type change (e.g., cancer, defferentiation, diseases)・Infulence to target gene expression by miRNA is subtle (〜10%) and contexts dependent.・In spite of that, miRNA critically contributes to the related processes (e.g., induction of cell cycle arrest)
3. Previous works (wet)Several researches suggest the contribution of miRNAsto cell senescence.Upregulation of miRNAs during cell senescencemiR-34a,miR-486-5p,miR-494,miR-210...Downregulation of miRNAs during cell senescencemiR-15a/b,miR-20a,miR-92,miR-16b....Induction of cell senescence by suppression ofmiRNA downregulated during cell senescence.
It is likely true that miRNAs contribute to cell senescence.However, which one?Dhahbi et al (2011) recently reported the upregulaton of141(!) miRNAs and the downregulation of 131(!!)miRNAs during cell senescence by deep sequencing.The reason why limited number of miRNAs revealedsignificant expression change during cell senescenceseems to be due to less sensitivity of microarray analysis.Is it truly critical the down/upregulation of such largenumber of miRNAs for cell senescence?
4. MiRaGE Method In order to select “Critical ” miRNAs during cell senescence, regulation of target genes by miRNAs is inferred by MiRaGE Server.For details, seeY-h. Taguchi and J. Yasuda (2010) Inference of GeneExpression Regulation via microRNA Transfection,ICIC2010Y-h. Taguchi and J. Yasuda (2012) MiRaGE: Inference ofGene Expression Regulation via MicroRNA Transfection IIICIC2011
MiRaGE :MiRNA Ranking by Gene Expression considered miRNA target miRNA gene VS target gene significantly up/downregulated? gene (t test, Wilcoxon test, KS test)
5. Correlation between target gene regulationby miRNA and miRNA expression changeduring cell senescence A) Confirmation of independence of cell line Two Cell Lines: IMR90 : young (PD 30) vs senescent (PD 48) vs MRC5 : young (PDL 28) vs senescent (PDL 63) mRNA expression change (during cell senescence) MiRaGE ⇓ Server P-values attributed to each miRNA
Intersection between N top-ranked significant miRNAs based upon P-values (t test) IMR90 vs MRC5 100% down 10 % of com m on m iRNAsregulation binomial: -log random P=0.05 30% 0 N 500 1500 N 500 1500 10 P10 100% 4 random up 20%regulation P=0.05 Thus, the results are cell line independent (possibly robust)
B) Confirm ation of reciprocal relationship between m iRNAexpression change and Pvalue (upregulation of target genes) IMR90,NGS Correlation Coefficient # of m iRNAs 700 0.35 100 0.05 quality score quality score -log 2 m iRDeep2: P=0.05 genom e alignm ent 10 1 01 10 2 10 4 program for m iRNA-seq P Threshold quality score Value (m iRDeep2)
Candidates of m iRNAs downregulaed (target genes are upreglated)NMRC : Norm alied m iRNA Read Counts P<0.05 RFC>1.0SCORE : m iRDeeps scoreP-value : upregulation of target genesRFC : Reciprocal Fold Change : young/senescent
Candidates of miRNAs upregulaed (target genes are downreglated)NMRC : Normalied miRNA Read Counts P<0.05 FC>1.0SCORE : miRDeeps scoreP-value : downregulation of target genesFC : Fold Change : senescent/young
Candidates of miRNAs upregulaed (target genes are downreglated) continuedNMRC : Normalied miRNA Read P<0.05 FC>1.0CountsSCORE : miRDeeps scoreP-value : downregulation of targe genes
6. Summary & Conclusion1. Selection of miRNAs commonly up/downregulated during cell senescence2. Reciprocal relationship between target gene regulation and miRNA expression change3. Reduction of number of critical candidate miRNAs during cell senescence (down: 131 ⇒１０, up: 141 ⇒ 32)
Full paper published inAging and Disease, (2012) No.4 in press(Epub 7 Jun 2012, online)MiRaGE Package will be included inBioconductior 2.11(planed to be released in this Autumn)MiRaGE Serverhttp://www.granular.com/MiRaGE/