The document discusses possible coregulation of target genes in brain regions by differential miRNA expression and miRNA-targeting-specific promoter methylation. It summarizes that numerous miRNAs have target genes with significantly hyper/hypomethylated promoters between brain regions. miRNA regulation of target genes is significantly correlated with miRNA-targeting-specific promoter methylation. Despite selecting diverse miRNAs that regulate target genes excluding promoter methylation, the KEGG pathways enriched by their union of target genes are largely common.

1. Possible miRNA coregulation of
target genes in brain regions
by both differential miRNA expression and
miRNA-targeting-specific promoter methylation
Y-h. Taguchi
Dept. Phys.
Chuo Univ.

2. Introduction
Epigenetic regulation of gene expression is
known to be important nowadays.
e.g.
- Development and Cellular Differentiation
- Diseases including Cancer
- Aging

3. Example of Epigenetic regulation of genes
- Transcription Factor Bindings to
promoter region
- Histon modification
- Promoter Methylation
- miRNA regulation of target genes.
Interaction
etc. etc. Rarely
Discussed
Both gene suppression

4. Methylation Gene
Promoter 3'UTR
cell nucleus
cytoplasm
miRNA
Methylation ↔ cell
nucleus
miRNA ↔ cytoplasm

5. Previous Research
Su et al (2011, BMC Genomics)
“miRNAs tended to target the genes
with a low DNA methylation level in their
promoter regions”
Promoter methylation ↔ miRNA targeting
Purpose of my study:
Promoter methylation is miRNA-targeting-specific or not?
(In other words, promoter methylation is affected by being
targeted by individual miRNA or not?)

6. MiRaGE Method
MiRaGE :
MiRNA Ranking by Gene Expression
Promoter Methylation
considered
miRNA target
miRNA gene
VS
target
gene
significantly
gene up/downregulated?
hypo/hypermethylated?
(t test, Wilcoxon test, KS test)

7. 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, approved with
reservations 1, http://f1000r.es/wv]

8. This Study
Comparison between miRNA, mRNA and
promoter methylation among distinct brain
regions (Frontal Cortex [FCTX], Temporal
Cortex [TCTX], Cerebellum [CRBLM], Pons
[PONS]) in miRNA-centric manner.
150 subject vs 4 regions = 600 samples
(We exclude samples without all four regions)

9. Calculation
- Number of miRNAs whose target genes are
significantly(*) up/downregulated among 1921 miRNAs
- Number of miRNAs with significantly (*)
hypo/hypermethylated promoters among 1921 miRNAs
- Correlation coefficients of mean ranks between miRNA
regulation of target genes and miRNA-targeting-specific
promoter methylation
for all six pairs of FCTX, TCTX, CRBLM, and PONS
(*) BH adjusted P <0.05

10. Corr. Coef. of Rank Number of miRNAs with
up/downregulated target genes
(or SD)
183 < +0.26 < 173 Number of miRNAs with
target genes with
(0.36) hyper/hypomethylated promoters
<
280 889
<
221 870 TCTX
FCTX
upregulated
>
>
>
+0 .49>
128
.4 3
+0 >
(0 . 3
.3 )
hypermethylated
(0
279
7)
608
1
267
>
> >
60 234 241
CRBLM
129
804
>
1038 168 450
>
>
>
>
- 0.
. 51 )
>
-0 5 +0.09 (0. 41
0. 3 42
( 490 )
>
(0.48)
439
>
>
1120 65 45
>
>
>
108
PONS

11. Present Results
- Substantial number of miRNAs have significantly
up/downregulated target genes.
- Substantial number of miRNAs have target genes with
significantly hyper/hypomethylated promoters.
- miRNA regulation of target genes and miRNA-
targeting-specific promoter methylation are significantly
correlated
- Correlation coefficients are not always negative
→ This indicates correlations between miRNA
regulation of target genes and miRNA-targeting-specific
promoter methylation are not bi-products of direct
correlation between gene expression and promoter
methylation

12. Selection of miRNAs that regulate target genes significantly
Liner regression analysis
x mjl
log P
l, l ' , mRNA l.l ' . Methyl
m , j,> =log P
m , j, > +log +a g e j +g e n d e r j
x mjl '
+ AIC based feature selection
Pl, l' j, ,mRNA : P-value of mth miRNA's target genes of jth
m, >
sample is upregulated between lth and l'th region
Pl, l', j, ,Methyl : P-value of mth miRNA's target genes' promoters
m >
of jth sample is hypermethylated between lth and
l'th region
x m j l : expression of mth miRNA of jth sample of lth region
Significant regulation by miRNAs of target genes

13. Bold : appear more than twice
underline : previously reported to be related to brain
non-
reciprocal
Reciprocal
Selected miRNAs are
diverse.
Biological meaning?

14. KEGG pathway analysis for the union of
target genes (DIANA-mirpath v2.0)
In contrast to
selected m iRNAs,
KEGG pathways
are com m on

15. Relation to Brain related facts
- TGFβ: relation to bipolar disorder
TGFβ
- MAPK: neuronal apoptosis
MAPK
- Wnt: Amyloid
Wnt
- ErbB: development of the nervous system
ErbB
…..
Many brain related KEGG pathways are
enriched by the union of miRNAs' target genes.

16. Q : Why can different set of miRNAs
target common KEGG pathways?
A : Target genes overlap
Example : TGFβ (CRBLM vs FCTX)
reciprocal vs non-reciprocal
no overlaped miRNAs (by definition)
share 36 mRNAs in TGFβ pathway....
Among 36 mRNAs, 25 mRNAs (RefSeq) are
differently expressed between CRBLM and
FTCX (see the next slide).

17. mRNAs expressed
differently
between FTCX and
CRBLM, targeted
by miRNAs
selected by present
study,
in TGFβ pathway

18. Conclusions
- numerous miRNAs have target genes with
hyper/hypomethylated promoter methylation
- miRNA-targeting-specific promoter
methylation is correlated with miRNA
regulation of target genes
- miRNAs that regulate target genes are
selected excluding contribution through
promoter methylation
- In spite of diverse selection of miRNAs,
KEGG pathway enrichments are largely
common