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Apparent microRNA-target-specific histone Apparent microRNA-target-specific histone modification inmodification in
mammalian spermatogenesismammalian spermatogenesis
Y-h. TaguchiY-h. Taguchi
Dept. Phys.Dept. Phys.
Chuo Univ.Chuo Univ.
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IntroductionIntroduction
Epigenetic regulation of gene expression is Epigenetic regulation of gene expression is known to be important nowadays.known to be important nowadays.
e.g.e.g.
- Development and Cellular Differentiation- Development and Cellular Differentiation- Diseases including Cancer - Diseases including Cancer - Aging- Aging
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Example of Epigenetic regulation of genesExample of Epigenetic regulation of genes
- Transcription Factor Bindings to - Transcription Factor Bindings to promoter regionpromoter region- Histon modification- Histon modification- Promoter Methylation- Promoter Methylation- miRNA regulation of target genes.- miRNA regulation of target genes.
etc. etc.etc. etc.InteractionInteractionRarely Rarely DiscussedDiscussed
Both gene regulationBoth gene regulation
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miRNA
cell nucleus
Gene
3'UTR
cytoplasm
histone modification ↔ cell nucleusmiRNA ↔ cytoplasm
histone modification
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Previous ResearchPrevious Research
Su et al (2011, BMC Genomics)Su et al (2011, BMC Genomics)““miRNAs tended tomiRNAs tended to targettarget the genesthe geneswith awith a lowlow DNA methylation level in their DNA methylation level in their promoter regions”promoter regions” Promoter methylation ↔ miRNA targeting Promoter methylation ↔ miRNA targeting
Purpose of Purpose of mymy study: study:Histone modification is miRNA-targeting-specific or not?Histone modification is miRNA-targeting-specific or not?
(In other words, histone modification is affected by being (In other words, histone modification is affected by being targeted by individual miRNA or not?)targeted by individual miRNA or not?)
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significantly up/downregulated?
(t test, Wilcoxon test, KS test)
MiRaGE :MiRNA Ranking by Gene Expression
miRNA
target gene
target gene
VS
gene
considered miRNA
MiRaGE Method
Histone modification
modified histone?
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My Previous Results
“Substantial number of miRNAs have target genes with significantly hyper/hypomethylated promoters”
References: - Y-h. Taguchi (2012) IPSJ SIG Tech. Rep. 2012-BIO-31(1) pp.1-6- Y-h. Taguchi, (2013) F1000Research [v1; ref status: approved 1, not approved 2, http://f1000r.es/wv]
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This Study
Comparison between miRNA and histone modification during spermatogenesis in miRNA-centric manner.
NGS:Spermatogonia(SG)→Spermatocytes(SC)→Spermatids(ST) vs Controls
H3K4me1, H3K27me3, H3K4me3, H3K27ac, H2AZ
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Calculation - Many miRNAs with significant (*) histone modification among multiple miRNAs for all SC, ST, SG vs Controls
(*) BH adjusted P <0.05
SC
SG
ST SCSG ST
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less modificated more modificated
Differenciation
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Cluste
Clustering using negative signed correlation coefficient of P-valuesP-values
Biologically feasible
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Clustering using negative signed correlation coefficient of logarithmic ratiologarithmic ratio
Less biologically feasible
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P-values computed by MiRaGE are more biologically
feasible than raw values (logarithmic ratio)
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less modificated
more modificated
Stemness Mouse ES cell specific
hsa-
↑during differentiation
hsa-
Critical roles in spermatogenesis
hsa-
Sperm abnormaly
hsa-
Expressed in abnormal semen
Selected miRNAs are biologically reasonableSelected miRNAs are biologically reasonable
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less modificated more modificated
Is it artifact ? → No, since essentially no miRNAs with Is it artifact ? → No, since essentially no miRNAs with significant histone modification after gene shufflingsignificant histone modification after gene shuffling
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Samples →
mR
NA
(g ene ) →
Test 2
Simultaneous shuffling with keeping correlation coefficients of logarithmic ratio
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Clustering using negative signed correlation coefficient of P-values P-values after simultaneous shuffling
More similar to cluster obtained by raw values (logarithmic ratio)
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Thus, converting logarithmic ratio to P-values by MiRaGE enphances
biological feasibility
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less modificated more modificated
Human sperm exhibits similar miRNA specific histone modification
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Too simple model? → comparison with lassoToo simple model? → comparison with lasso
differential expressionof i th mRNA and j th histone
modification
11 if k th miRNA targets ith mRNA 00 otherwise
Predicted activity of k th miRNA
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β
k
P-v
alu
es
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MiRaGE is well coincident with lasso. Thus, biological feasibilities obtained by miRNA specific histone modification will
not be likely fake.
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ConclusionsConclusions
・ P-values computed by MiRaGE are highly tissue/histone modification specific.・ Most significant miRNAs are stem cell or sperm specific. Thus, biologically feasible.・ Since gene order shuffling destroys miRNA specific significant histone modification, it is not an artifact.・ Since it is coincident with lasso, biological feasibility is not due to too simple algorithm・ Biological mechanism that causes miRNA specific histone modification should be cleared.