Radiation Effects on Regulation of Gene Expression

Mitsuru Nenoi, Bing Wang, Tetsuo Nakajima

Radiation Effect Mechanisms Research Group, Natl. Inst. Radiol. Sci., Chiba, Japan

Risk of Radiation-Induced Cancers

?

From Hall,E.J., Radiobiology for the Radiologist. J.B. Lippincott Company

Experiment of continuous low-dose-rate irradiation at IES

42-49days 400 days until spontaneous death

0 Gy/min

40 nGy/min

800 nGy/min

16,000 nGy/minsignificantly different

significantly differentin female

not significantlydifferent

Life Span

gene expression profile ?

SPF condition

Dose-rate range used in the study at IES

dose-rate(nGy/min)

Natural radiation in Guarapari, Brazil

Natural radiation level in J apan

Space station (ship outside)

Space station (ship inside)

Cosmic radiation at 12,000m elevation

1

10

100

1000

1000016 mGy/min

800 nGy/min

40 nGy/min

Large-scale study on the effect of continuous low-

dose-rate irradiation

atInstitute for

Environmental Sciences (IES),

J apan

Number of genes whose expression levels were changed more than 1.6-fold by irradiation

dose-rate(nGy/min)

32 1 5 6650 18 3 21

13,000 16 11 2732 & 650 1 0 1

650 & 13,000 3 1 413,000 & 32 0 0 0

up-regulated down-regulated total

Functional analysis of deregulated genes Four genes whose expression was enhanced after irradiat

ion at 650 nGy/min and 13,000 nGy/min were involved in mitochondrial oxidative phosphorylation pathway.

(mGy/ min) (mGy/ min)

Uqcrb

0

0.65

0.03

2

1.6

0.5

1

(mGy/ min)

13

Ndufb9

0 13

0.65

0.03

2

Atp5k

0

0.65

0.03

2

Ndufv2

0 13

0.65

0.03

213(mGy/ min)

Genes whose expression levels were significantly varied by irradiation

Welch’s ANOVAp<0.05MTC: None

621 genes

Clustering analysis of selected genes

①

③④⑤⑥⑦⑧

⑮

⑬⑭

⑫⑪⑩

⑯

⑨

②

0Gy/

min

13 m

Gy/

min

650

nGy/

min

32 n

Gy/

min

① ② ③

④

⑤ ⑥ ⑦ ⑧

⑨ ⑩ ⑪ ⑫

⑬ ⑭ ⑮ ⑯

0.5

1.6

1

0.5

1.6

1

0.5

1.6

1

0.5

1.6

1

0Gy/

min

13 m

Gy/

min

650

nGy/

min

32 n

Gy/

min

0Gy/

min

13 m

Gy/

min

650

nGy/

min

32 n

Gy/

min

0Gy/

min

13 m

Gy/

min

650

nGy/

min

32 n

Gy/

min

0Gy/

min

13 m

Gy/

min

650

nGy/

min

32 n

Gy/

min

GO category containing similar gene list to the clusters

①

78 2.8E-07cytoplasm

78 4.0E-10mitochondrion organization and biogenesis 78 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-10

nucleosome

78 4.0E-10

mitochondrion

78 4.0E-1078 4.0E-10

organelle organization and biogenesis

78 4.0E-10

cytoplasm organization and biogenesis

78 4.0E-10

cell organization and biogenesis

78 4.0E-10

transferase

78 4.0E-10

cytoplasm organization and biogenesis

78 4.0E-10

cell organization and biogenesis

78 4.0E-10

organelle organization and biogenesis

24 0.0013energy pathways

22 5.7E-04mitochondrion

24 0.0055

78 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1037 0.007137 0.0071

37 0.0071

78 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1017 1.4E-0548 0.031

17 1.4E-05

⑩

17 1.4E-05

94

Overlapped gene

numberp valueGO category

GO SLIMS Cellular Componentmitochondrion organization and biogenesis 78 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-1078 4.0E-109 0.032

39 0.040

mitochondrioncytoplasm 22 0.0016

9 0.00614 0.012

Conclusion1 Mitochondrial oxidative phosphorylation was suggested t

o be elevated after irradiation at 650 nGy/min and 13,000 nGy/min.

Mice irradiated with low dose-rate radiation in this range may suffer oxidative stresses caused by elevated mitochondrial respiratory activity. This oxidative stress may be one of the factors that cause life spans-shortening of irradiated mice.

Multiple genes were found whose expression was changed more than 1.6-fold after 32 nGy/min, but their biological significances are unclear.

Radioadaptive response

Chromatid deletions

1.5 Gy X-rays3H-thymidine

(0.01-0.1 µCi.mL-1)

Olivieri et al., 1984

When cells are exposed to a priming low-dose or low dose-rate of radiation, they show a reduced biological response to a challenging high dose of radiation.

Adaptive response

Humanlymphocytes

Radioadaptive response in mice during late embryogenesis

Digital defects

Prenatal death

E11 E12 E18 E20

Prim

ing

IRC

halle

ngin

g IR

Fertilization 1

2

3

4

5

6

7

0 10 20 30 40 50

Nu

mb

ers

per

Dam

of

Fet

use

s

Priming Dose (cGy)

*

*

Living FetusesLiving Malformed Fetuses

(A) (B)

Experimental model

E11 E12

Priming irradiation

cDNA Microarray analysis

Whole-fetus removed

RNA extraction

10.34F

Non-effective

dose

0.50.34E

Non-effective

DR0.32.5D

0.34.2C

Effective dose-rate

(DR)

0.30.34B

Control00A

Description

DOSE

(Gy)

Dose Rate

(Gy/min)

Priming irradiation parameters:

AR genes

Identification of global differences

AR-inducible conditions (B and C)

AR-uninducible conditions (D, E and F)

861 AR genes

Selection of genes with:

• similar expression ratios inside each group

• significantly different modulation ratios between both groups

Functional annotation

Gene enrichment

Functional annotation and gene enrichment analysis

Cyt

oki

nes

an

d In

flam

ma

tory

Re

spon

se

0.0%

Se

nso

ry p

erc

eptio

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ath

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ynth

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ell p

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Met

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and

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ucle

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Fa

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cid

Be

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xida

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3 B

iGC

aT

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DB

Cla

ss A

Rh

odo

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-like

Me

tallo

pep

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itoch

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rial

mem

bra

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ran

slat

ion

HS

P7

0 a

nd

Ap

opto

sis

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eta

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nalin

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ath

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row

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acto

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han

nel a

ctiv

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Tis

sue

s-B

loo

d a

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ymp

hp

38 M

AP

K s

igna

ling

pat

hway

1-T

issu

e-E

ndoc

rine

and

CN

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yto

sol

Hor

mo

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ix M

eta

llopr

otei

nas

esE

lect

ron

Tra

nsp

ort

Ch

ain

Cel

l pro

life

ratio

nT

ota

l

10.0%

20.0%

30.0%

40.0%

% of AR genes in each molecular pathway

Significantly enriched

Non significantly enriched

Conclusion2

Intracellular and intercellular signaling activities are suggested to be important bases for radioadaptive response in mouse fetuses during late organogenesis.

The present cDNA microarray analysis failed to show evidences for involvement of DNA damage repair genes in the radioadaptive response.

Acknowledgements

Tetsuya Ono,T (Tohoku Univ.)

Guillaum Vares (NIRS) Keiko Taki,K (NIRS) Jianyu Wu (ADSTEC)

Tsuneya Matsumoto (IES)

Yoichi Oghiso,Y (IES) Kimio Tanaka (IES) Kazuaki Ichinohe (IES) Shingo Nakamura,S (IES) Satoshi Tanaka (IES)

This work was supported by the Budget for New Nuclear Crossover Research from the Ministry of Education, Culture, Sports, Sciences and Technology, Japan