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Tomoyo Ujisawa

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Graduate School of Natural Science, Konan University



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C. elegans 2Zq DNA 2Ðįīˍß3Ķɑɪȕȉ+3ȝà>/ 1998 ť/ǣŅE*CZq\Ma3jðȉ2+?Ȋ/Ŕɍ 100 Mb +D(C.

elegans Sequencing Consortium, 1998)C. elegans Zq DNA /3tkZq/?¿ĿE*

Dˇ°ľHĶ?(¹4ǣ;'$Ƴƹ/ǣ;'$ɑɪǚ0A/x

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and Horvitz, 1998)ˤ˙2űƋ/DxXɑɪǚ/ȞţH?(ĴȞµˇ°ŀȤ/ĶƧú˫E*C"EBĴȞµ(egl-1ced-9ced-4ced-3 .-)2þğˇ°ľ
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lx^? PDE ?ǭƁò*CɑɪÕ cGMP Ȃũ̀"EB cGMP »ĿƁgp(CNG) /ɕčD$>gp˛ƤEɮÝǐ*Dƪ+ƲD</ÜD,lx^k`j^HǭƁòB/ PDE ǭƁò*cGMP HÝʒD$>ʏɑɪ2ɑɪÕ cGMP ȂũǵŕD,+gp˚ˁÝǐʪD(Fain et al., 2001; Fesenko et al., 1985)ñ
+3ȊŅ2ñȉʨHąŉDƈʐnHǑƋDòŀȉʨ3 GPCR +ąŉE$ŵ/ Gα esWʨHǭƁò"EKjnˏ^Wc(AC)HǭƁòɑɪÕ2ȔȋKjq^
ˏ(cAMP)Hıí!DE CNG gp2˛ĆHŭʪɲ2ɮÝǐʪD(Kurahashi, 1990 ; Nakamura and Gold, 1987 ; Pace et al., 1985)
C. elegans 3ñ@ēȉʨ.-2ĵˌåȀH/˸ˌ@Ŗˌ/ĿĥDƈʐn+ąŉD"2&˸ˌ/3 12 ŏ2ƈʐn
Ȁ@"2ɑɪÕƅĬ°˂/˝GDÝľ?ˌÝȤ/G'*CąŉDå

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D ODR-3 HǭƁòB/XKn

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CASJ ƈʐnÕ/ĿĥDÝľƅĬ°˂ɔʫ3ɭǍðȉ2Ýľɔʫ,˺²*ÎƅĬHąĄ'$˦2ɲˬ³2ĴòɭǍðȉ,ɜɿ+3ʹ+D

D(Liu et al., 2010)

ÔµȤ/3ɭǍðȉ+3ÎHąŉD,k`j^, PDE ǭƁòcGMP HÝʒ* CNG gp˚DƪC. elegans +3ASJ ƈʐn+ÎąŉED,Gα esWʨ GC HǭƁòcGMP ȂũHư!D,+ CNG gpH˛Ć!D

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CTRPCTRPVTRPMTRPATRPN

TRPPTRPML 2 7 (2\wvJH?&tkBɜɿ;+Ũ¿ĿE*

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C. elegans 2ƈʐn/DǷũąŉ/˝*3Ƿũ/ŏDſɅʂð+/DǷũʩƁHƔǓ/ʒǃʂGE**Ƿũ3/ AFD ƈʐn@ AWC ƈʐn+ąŉED,

ĬĒE*
/

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D(Takeishi et al., 2016)GC /3ĈǻĨ2?2,ɲɕčĨ2?2ĿĥDGCY-8GCY-18GCY-23 3ɲɕčĨ2+?ɑɪĵ2ƅĬHɑɪÕ/ȨƜ
°˂DbxeĨ2 GC 2ǑʽHƓ'*-8-

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D TAX-4

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2015)
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D$>ǷũʩƁ/DǷũąŉn+3 GC Ƿũąŉµ+D,ȳęE*

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GE*

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AWC Ƿũąŉn2ǷũƅĬ°˂/˝GD 3 ˔µ G esWʨ/ODR-3 2ĴȞ
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ɑɪ2ɖCʵ<Hʂ'$20;$3 25+˼ɩE$ tax-6 ĴȞµ3´ǷåȀŵ?
90%«ȕĿ$9:Ð*2n+2ˇ°ľȣȑHʛœD unc-14 xe2Ǯ+ tax-6 2ˇ°ľHȣȑ!$ tax-6 ĴȞµ2k`_OnhWɌɗ+3
20˼ɩŵ/´ǷåȀH$˦2ȕĿȐ˓ȕǈ,Ďǒ2 10%«,.'$25˼ɩÀµ+?´ǷåȀŵ2ȕĿȐɍ 50%,.C˓ȕǈ2ʄȑĨ/ʶ)$B/

/ƈʐn+ tax-6 ˇ°ľHȣȑ!$ tax-6 ĴȞµk`_OnhWɌɗ
(tax-6;Ex[tax-6p(1.1kb)::tax-6])/

*?tax-6 ĴȞµȳ´ǷɦƁȞţ2ˌÝȤ.

ĞźʍBE$(ġ 9)«2ɕǅB´ǷɦƁ3ƈʐn+äŹEDĈɫƁ  Ÿ B E $   ƪ +   2 tax-6;Ex[tax-6p(1.1kb)::tax-6] Ɍ ɗ  Ɠ ' *tax-6p(1.1kb)::tax-6 x`l3ŕƧ+D

Dƈʐn«ĵ2¦ĥn+?ȣȑD$>´ǷɦƁ,ƈʐn,2ȩ˝ƁHŸD/3B.DʒǃŽʋ+D,ɤ$"+ƈʐn2ƈʐƼɂ(ƈʐɞǞ)2ǑʽűƋ@"2ɚƓ/˝GDŧ(2ˇ°ľ2ĴȞµHȗ*´ǷɦƁHʒǃ$,F(ġ 10ʄ 1)

IFT complex B/ che-13 , IFT52/osm-6 2ĴȞµ+ 20˼ɩŵ2´ǷɦƁ2Ȟţʑō
E$(ġ 10A)2 2 (2ˇ°ľ38*2ƈʐn+ȣȑ*D,B

E2ƈʐn´ǷɦƁ/˝D,ȳęE$(Collet et al., 1998;
Deane et al., 2001; Haycraft et al., 2003; Perkins et al., 1986)

 Bj{Y[~17>:<JO´ǷɦƁ/˝GDȊŅ2ƈʐnHĎŅD$>/ACŕƧ2ƈʐ

-18-n2ȣȕ@ǕɫȞţ/.'*DĴȞµHȗ

*ʒǃHʂ'$"2ɕǅ

cGMP »ĿƁgpH[lD tax-4 , tax-2 2ĴȞµ/*20+˼ɩ$À

µ2´ǷåȀŵ2ȕĿȐɍ 90%,˹ɻ.Ȟţ<BE$(ġ 10B)TAX-4 3˸ˌ2 10
ŏ2ƈʐn+ȣȑ*

D(Coburn and Bargmann, 1996 ; Komatsu et al., 1996)(ʄ 1)"+ȊŅ2ƈʐnȊȞȤ/ȣȑDˇ°ľ2xe2Ǯ/ tax-4 cDNA H(.

%.

%x`lH·ʇ"EH tax-4 ĴȞµ+ȣȑ!$

tax-4 k`_OnhWɌɗH·ʇ$tax-4 ĴȞµȳ´ǷɦƁ2Ȟţ3trx-1
xeHȗ*tax-4 ĴȞµ2 ASJ ƈʐnȊȞȤ/ tax-4 ˇ°ľHȣȑ!$,/Ƹ?˹ɻ.ĞźʍBE$(ġ 11);$str-3 xeHȗ*

tax-4 ĴȞµ2 ASI ƈʐn+ tax-4 ˇ°ľHȣȑ!$ĭč/?ǐG+D tax-4 ĴȞµȳ´ǷɦƁ2ıů´$ƪceh-36 xeHȗ*Ƿũąŉn,*ȭBE*D AWC ƈʐn+ tax-4 ˇ°ľHȣȑ!$ĭč/3tax-4 ĴȞµ2´ǷɦƁ2Ȟţ3Ğź.'$(;CcGMP»ĿƁgp TAX-4 2 ASJ ƈʐn/DǕɫ´ǷɦƁ2ƋɁ/Ž˶+*D,ȳęE$
ASJ ƈʐn´ǷɦƁ/˝DĈɫƁŸBE$$>E;+

ASJ ƈʐn+ˇ°ľȣȑHʛœDxe,*¸ȗ*lU^/TRX-1 2ĴȞµ/($gR*?´ǷɦƁHʞ8$,F20+˼ɩ$Àµ+´ǷåȀŵ2ȕĿȐ̀.DȞţ<BE$(ġ 12)B/˓ȕǈ2 ASJ ƈʐn
HɸɎ]/ACȉȓȤ/ȱĲD,+tax-4 ĴȞµ,Ďǒ2´ǷɦƁ2ıůȞţʑōEDHǎʘ$;
ɌɗH·ʇ$(ġ 13A)2ɌɗHȗASJ ƈʐn+ GFP HȣȑD* ASJ ƈʐn2³ɡHȲʚ.B

-19-

ASJ ƈʐn2ǉHɸɎ]+ȱĲ$ASJ ƈʐn/D GFP
2ʀÎǱ$,HȲʚ$ŵ"EB2Àµ2´ǷɦƁ2ʄȑĨHčʔ 15 Àµ/(*Ȳʚ$,F20+˼ɩ$ŵ/ 2/ɡE*?ķˌÝȕĿDȞţʑōE$(ġ 13B)2ʄȑĨ3tax-4 ĴȞµĎǒ2ʄȑĨ+'$«2ɕǅ

BASJ ƈʐnÀµ2´ǷɦƁ/˝D,ȳęE$

  [~17>:<%9(#5"6>)<'´ǷɦƁ/˝D,ȳęE$ ASJ ƈʐn3E;+/ÎHąŉDƈʐn,*ȭBE*$ASJ ƈʐn´ǷɦƁ2ƋɁ/˝*xM~.ǷũåȀHąĄDƈʐn+D-HǸŅD$>/ASJ ƈʐn2ǷũſɅƁHS^NM_XǧHȗ
$.Ƿũąŉn,*ȭBE**ʒǃ

D AFD ƈʐn@ AWC ƈʐn3ƽȰȾɕǅB´ǷɦƁ/˝*.

,ȳęE*

D$>

ASJ ƈʐn%/ȅȃHŰ**S^NM_XʒǃHʂ'$C.
elegans /3Ę˺2A.ˬ³»ĿƁmkNgpH¦$ǭðˬ³.

$>ɑɪÕ72S^NMR2ǮÏȨƜ2ȵɔǭð2Ĵò,*ʑō+D"+ǷũåȀƳ2ɑɪÕ2S^NȂũH in vivo +˰¼ʉȤ/ǸŅD$>/
ˇ°ľ/A'*[lED cameleon ,Ĕ4EDS^NMjLYeˇ°ľHȗ$cameleon 3 N Ƽ/ɾɸʀÎesWʨ+sWʨ+

D CFP HC Ƽ/̂ɸʀÎe

D YFP H?&"2˜/S^Nɕč\MkHƓ(UesWʨ+-20-

D(Miyawaki et al., 1997)ʻţɑɪÕS^NȂũ´

ȋƉ+3440 nm 2ïʪÎHŰ*D, CFP 2ʀÎʑōEDɑɪÕ7S^NMRǮÏD,
cameleon 2S^Nɕč\Mk/"EBɕčCFP , YFP Ƨ nm /ƜʶD
"2ɕǅCFP 2ʀÎʀÎÓ́PpVȹð̄FRET̅,Ĕ4EDPpVȹð/AC YFP /ȹðYFP 2ʀÎ?ʍBEDA/.D2Ƴ2CFP , YFP 2ʀÎ2ǝ(YFP/CFP)HʔɆD,+ɑɪÕS^NȂũHŅ˔òD,+DE;+/ǷũʩƁ/DǷũąŉn+cameleon Hȗ

D AFD @ AWC 2ǷũſɅƁ

$S^NM_X/A'*ʒǃE*

D(Kimura et al.,

2004; Kuhara et al., 2008)AFD ƈʐn@ AWC ƈʐn3ǷũåȀH
D, YFP/CFP 2ÄķĴòDƪ+ǷũąŉHʂG.

ASH ƈʐn

@ ASE ƈʐn+3ǷũåȀH*? YFP/CFP 2Ä/ĴòʪB.

,ĬĒE*

D(Kuhara et al., 2008)

15˼ɩŵ2˓ȕǈ2 ASJ ƈʐn2ǷũſɅƁHS^NM
_X+ǸŅ$(ġ 14A);˓ȕǈ2 ASJ ƈʐn/ cameleon Hȣȑ!$ɌɗH·ʇ$cameleon Hȣȑ!$ÀµH!$xskHÂɁ˹Ż˗` i  _  2 Ƿ ũ ä Ź x   k / Ģ Ņ   400 Á 2 Á Ȑ + ʑ ō  .  B 
172317,

.%B.ǷũĴòH"2˦2 ASJ ƈʐn+ʍBED CFP , YFP 2ʀÎÄHʖ˖ǸŅ$"2ɕǅ˓ȕǈ2 ASJ ƈʐn
+3Ƿũư,ˡ/±ɑɪÕS^NȂũ?ư,ˡHȳŅţȋƉBƸķÄ2˜/3ɍ 10̃2 YFP/CFP 2ĴòʑōE$Ďǒ/20˼ɩŵ;$325˼ɩŵ2˓ȕǈÀµ/˝*?ǷũåȀƳ2 ASJ ƈʐn2S^NM
_XHʂ'$"2ɕǅ20˼ɩE$Àµ+3ɍ 15%25˼ɩE$Àµ-21-+3ɍ 20%2 YFP/CFP 2ĴòʍBE˼ɩǷũ̀

9- YFP/CFP 2ĴòÄ?ưǷũ/ŏDĂſƁ̀.'$(ġ 14B)Ǘ/´ǷɦƁ/ȞţHȳ$ tax-4*ASJ ƈʐn2ǷũſɅƁHS^NM_X+ʒǃĴȞµHȗ$"2ɕǅtax-4 ĴȞµ+3ǷũåȀH*? ASJ ɑɪÕ2S^NȂũ3;CĴò!

Ƿũ/ŏDĂſƁHȳ.'$(ġ 15A, B)tax-4 ĴȞµ2 ASJƈʐn+2< tax-4 ˇ°ľHȣȑ!$Ɍɗ/˝*?S^NM_
XHʂ'$,F2Ɍɗ2 ASJ 3ǷũåȀ/ŏ*ĂſʍBEǷũſɅƁ2ȞţĞź$(ġ 15A, B)
ASJ ƈʐn3Îąŉn,*ȭBE*BˮɸÎHƈȭD,ȭBE*cameleon 2ïʪÎ3 440 nm +ǭƁ/Ų˲H*Ăſ*D¥Ğ2S^NM_X/¸ȗ$D,B2ïʪÎɳµ ASJ ƈʐn2DĈɫƁɤBE$"+ASJ ƈʐnˮɸÎ/D2+3.Ƿũ/Ăſ*DĚ2Îąŉµ+DȊ/ɐĵ

D,HƲȲ/D$>/ ASJ +Ǖɫ

D LITE-1 HǖƠ$ lite-1 ĴȞµHȗ*ASJ 2S^NM

_XHʂ'$"2ɕǅlite-1 ĴȞµ+3˓ȕǈĎǒ/Ƿũ2ư,ˡ/ŏ*ɑɪÕS^NȂũ2ĴòʑōEÎąŉµ LITE-1 .

ȋƉ+?Ƿũ

/ŏDĂſƁȲʚ+$(ġ 15A, B)(;CASJ ƈʐn3 cameleon 2ïʪÎ+3.ǷũHƈȭ*ĂſD,ȳęE$."E#E2Ɍɗ/ŏ*ǷũåȀH.

ǀ­+ cameleon 2ïʪÎ2<HȆŒ$ȋƉ2 ASJ 2S^NȂũ2Ĵò?ǸŅ$,FɑɪÕS^NȂũ2˹ɻ.Ĵò3ʍBE.'$(ġ 15C, D)¥Ğ2ʒǃ+3cameleon 2ïʪÎ(440nm)3 1 ȸ˜/ 100-50 msec ,˰ţ/ȮƳ˜2s`ȆŒH*

D$>ˮɸÎȆŒ2Ų˲3ˣĵ+DɈĠÕ,ɤ

-22-

$
«2S^NM_X/ADʒǃɕǅBŷǁÎąŉn,*ȭBE$$ ASJ ƈʐnǷũ/?ĂſDǷũąŉn+D,ȳęE$

  EDIPjSZKAE;+2ɕǅB´ǷɦƁ2ƋɁ/ąŉ** ASJ ƈʐnǷũHD,G'$ASJ ƈʐn3Îąŉn,*ȭBE*CASJ Õ/*ÎƅĬ°˂/˝GDÝľɔʫƲB,.'*2010) (ġ 16A)Î3˸ˌ ASJ n2ǔȋȿʪÍɂ/ŗĥ*
LITE-1 +ąĄBE3 ˔µ G esWʨ α \wƁòXKnˏ^Wc+nhk+

D(Liu et al.,DÎąŉµ

D GOA-1 , GPA-3 Hǭ

D DAF-11 , ODR-1 H¦*ɑɪÕ cGMP ȂũHĴò!"E/A'* cGMP »ĿƁgp TAX-4 2˛ĆHäŹD,+ɑɪÕ
ƅĬ°˂Hŭʪ;$cGMP HíǠÝʒD{`{_P`ic PDE-1
PDE-2"* PDE-5 ? ASJ Õ/*Ǖɫ*DEB ASJ ƈʐn2ÎąŉƅĬ°˂+ǕɫDÝľ´ǷɦƁ2äŹ/Óʻ*>ASJ 2ÎąŉƅĬ°˂/˝GDÝľ2ĴȞµ/(ASJ /ĿĥDĚ2Îąŉµ+D-Hʞ8D$*´ǷɦƁHʞ8$(ġ 16B)

D LITE-1 HǖƠ$ lite-1 ĴȞµ3˓ȕǈ,9:Ďǒ2ʄȑĨHȳ$,BÎąŉµ3´ǷɦƁ/˝.

,ȳęE$

ASJ 2ÎƅĬ°˂/˝GD 3 (2 Gα esWʨ(goa-1gpa-1gpa-3) 2Ĵ

-23-Ȟµ@2 (2XKnˏ^Wc(odr-1 , daf-11) 2"E#E2ĴȞµ320˼ɩŵ/´ǷåȀH*?ȕĿ+DȞţʑōE$B/{`{_P`ic2ĴȞµ/*?ȞţʑōE$«2,BASJ ƈʐn2Îƅ

Ĭ°˂/˝GDÝľ´ǷɦƁ/?˝GD,ȳęEÎ,Ƿũ2ąŉ3Óʻ2
Ýľ/AD̉esWʨɔʫ+°˂EDĈɫƁɤBE$B/Gα esW
ʨ2ĴȞµ+

D goa-1 ;$3 gpa-1 2´ǷɦƁ2Ȟţ3˒ĴȞµ@˒ĴȞµ/D,+ACů´ǷɦƁ2ȞţʍBE$ (ġ 16Bgoa-1; gpa-1 ˒ĴȞµ

goa-1; gpa-3 gpa-1 ˒ĴȞµ)Ďǒ/XKnˏ^Wc2ĴȞµ+

D daf-11, odr-1 2´ǷɦƁ2Ȟţ? daf-11; odr-1 ˒ĴȞ/D,+9: 100%2ȕĿȐHȳȞţʑōE$(ġ 16B)(;CEB Gα esWʨ@XKnˏ^Wc3ʈƧ2Ýľ"E#EÓĎ*Ǖɫ*D,ɤBE$ƪ+EB2

ˇ°ľ3 ASJ ƈʐn«ĵ+?ȣȑD,ĬĒE*

DGOA-1 39:Ð*2n+ȣȑ*CGPA-1 , GPA-3 ? ASJ ƈʐnHđ=ʈƧ2n+ȣȑ*

D(Jansen et al., 1999)tax-4 ĴȞµ2S^NM_

X2ɕǅBASJ ƈʐn/DǷũƅĬ°˂3ASJ 2ƈʐƼɂ/Ŀĥ
D cGMP »ĿƁgp/TAX-4 H¦*ʂGED,ȳęE*ąŉ3 ASJ 2ƈʐƼɂ+ʂGE*

D$>Ƿũ

D,ɤBED"+GOA-1GPA-1GPA-3"E#E ASJ ƈʐn2ƈʐƼɂ/ŗĥ*DHʞ8D$>/ASJ ȊȞ

Ȥ/ˇ°ľȣȑHʛœD trx-1 xe2Ǯ/ goa-1gpa-1gpa-3 2Č cDNA
, YFP(venus)ˇ°ľH(.

% DNA [`kWkH"E#E˓ȕǈ/ˇ°ľœÏ$(ġ 17)VENUS 2ʀÎHƔǓ/ʒǃHʂ'$,FGPA-1 , GPA-3 3ƈʐƼɂ/ŗĥ*$GOA-1 3ƈʐƼɂ«ĵ/?ɑɪµ@ʯɏ/?Ŀĥ*

-24-

D,G

'$
Ǘ/EB2 Gα esWʨ2ĴȞµ2´ǷɦƁ2Ȟţ ASJ Ƿũąŉn
2Ȟţ/AD?2Hʞ8$ÔµȤ/3goa-1; gpa-3 gpa-1 ˒ĴȞµ2
ASJ ƈʐnȊȞȤ/3 (2 Gα esWʨ2 cDNA Hȣȑ!$k`_
OnhWɌɗH·ʇ´ǷɦƁHʒǃ$"2ɕǅgoa-1; gpa-3 gpa-1 ˒ĴȞµȳ 20˼ɩŵ2´ǷɦƁ2ȞţˌÝȤ/Ğź$(ġ 18)ƪASE ēʐn
@ǷũʩƁ2ȵɔĞʫ2 AWC Ƿũąŉn+ 3 (2 Gα esWʨ
2 cDNA Hȣȑ!$ goa-1; gpa-3 gpa-1 ˒ĴȞµ+3´ǷɦƁ2Ȟţ3Ğź.
'$(;CEB2 Gα esWʨ2 ASJ ƈʐn/DǕɫ´ǷɦƁ/˒ʋ+

D,ȳęE$goa-1; gpa-3 gpa-1 ˒ĴȞµ2 ASJ ƈʐn+ 3 (2 Gα esWʨ2 cDNA Hȣȑ!$%+3˒ĴȞµȳ´ǷɦƁȞţ3ńÐ/Ğź.'$"+"E#E2 Gα esWʨ2ˇ°ľ
HČ2ˇ°ľɳʭ2xeHȗ2ɕǅ"EB2Ɍɗ/*ȣȑ!"2ĭč2ʄȑĨHʞ8$"*?˒ĴȞµȳ´ǷɦƁȞţ3ńÐ/Ğź!



ASJ ƈʐnȊȞȤ/ Gα esWʨ2ˇ°ľHȣȑ!$ĭč,ĎȺũ2Ğź
Ȑ,.'$«2ʒǃBASJ ƈʐnȊȞȤxeHȗ* 3 (2Gα esWʨ2 cDNA HĎƳ/ȣȑ!$˒ĴȞµ+?"E#E2 Gα esW
ʨɳʭ2xeHȗ*"EB2 Gα esWʨ2 cDNA HĎƳ/ȣȑ!$˒ĴȞµ+?ˌÝȤ.ĞźʍBE.'$(;CASJ ƈʐnȊȞȤ/
3 (2 Gα esWʨHȣȑ!$˒ĴȞµɌɗ+3´ǷɦƁ2ʄȑĨöÝ/Ğź*DĈɫƁɤBE$ƪ+ńÐ/Ğź.'$ȓș,*3Gα e

sWʨ2 cDNA /3ˆƐȤ`xM^X/ADKMdvQĿĥD$>¥-25-

$ Gα esWʨ2 cDNA 2KMdvQ%+3öÝ+Ğȗ

BED;$3 Ȼ˺2 Gα esWʨ2ˇ°ľHĎƳ/ȣȑ!*ȣȑ˔2r`?˒ʋ.ʋɎ?E.

'$ĈɫƁɤ
D$>"2  [~17>:<  -<3&v!@jSXt´ǷɦƁ/*ASJ Ƿũąŉn/D 3 (2 Gα esWʨ2ÓĎȤ.äŹŽ˶+D,ȳęE$$>Ǘ/ASJ n2ȵɔǭð2

y+?ʈƧ2ÝľÓĎ*Ǖɫ*

DHS^NM_XǧHȗ*ʒǃ$ASJ n2ǷũſɅƁʒǃ/ȗ
,$ǷũåȀ3 172317ǷũåȀse/í231723,ǷũåȀse?ȗ

$(ġ 19)2,ǩȧ$Ä,*ɑɪÕS^NȂũƸ?̀.'$}Mk+

D Maximum(Max),Ăŏ/Ƹ?´.'$}Mk+*ƸķĴòÄ,ƸŔĴòÄ2Š+D Minimum(Min)"

D Max-Min ƕBEDMax 317B 23/ǷũHĴò!$˦2ȵɔǭð2ĴòHʄ*CMin 3 23B 177ǷũHƌ$,2ŅţȋƉ72ƌCÔčHʄ*D;$17B 2372ǷũĴòH 1 ǗǷũåȀ,23B 1772ǷũĴòH 2 ǗǷũåȀ,ʍ.2 2 ǗǷũåȀ/ŏDĂſƁH Max-Min +ʄ*D2317232ǷũåȀH

$ĭč?Ďǒ/ƸķĴòÄH MaxƸŔĴòÄH Min ,EB2ŠH Min-Max+ʄ$20+˼ɩ$˓ȕǈ/ 1723172ǷũåȀHD,Ƿũư/±

ɍ 10%2 YFP/CFP 2ư<BE$(ġ 19B)2317232ǷũåȀ

-26-

HD,Ƿũ´/±ɍ 10% 2 YFP/CFP 2 ´   < B E $ (ġ 19C)  2

172317, 2317232ǷũåȀH Gα esWʨ̆XKnˏ^Wc{`{_P`ic2ČĴȞµ/ASJ ƈʐn2ǷũſɅƁHʒǃ$(ġ 20-25)2,G esWʨɔʫ/*Ǚ2ǷũƅĬ°˂Hʂ GαesWʨ@XKnˏ^Wc2ǕɫǖƠ*

DĴȞµ+3cGMP »ĿƁgpB2S^NMR2ɑɪÕǮÏĽBED$>1723172Ƿ
ũåȀ/ŏ*3ǷũſɅƁŮ.C¹4 Max 2Ä˓ȕǈAC?Ŕ.
D,Ɔ$Ăŏ/{`{_P`ic+3cGMP »ĿƁgpB2S
^NMR2ɑɪÕǮÏˁê/.D$>ǷũſɅƁů.CMax 2Ä˓ȕ
ǈAC?ķ.D,ɤBE$;1723172ǷũåȀH$˦/Gα esWʨ2ĴȞµ+D goa-1gpa-1gpa-3 "E#E2 ASJ ƈʐn/DS^NȂũ2Ĵ
òHʞ8$,Fgpa-1 ĴȞµ2<+ Max /G.ȞţʍBE$(ġ 20B, F)2ÈĐ3goa-1; gpa-1 ˒ĴȞµ@ goa-1; gpa-1 gpa-3 ˒ĴȞµ/?ʍBE$(ġ 20D,
E, F)2317232ǷũåȀH$ĭč/?gpa-1 ĴȞµ2<+ Max /G.ȞţʍBE(ġ 21B, F)Ķ˒ĴȞµ+?Ďǒ/ Max /G.ȞţʍBE$(ġ

21D, E, F)(;CEB Gα esWʨ2Ķ˒ĴȞµ/D Max 2Ȟţ3 gpa-1
ĴȞ/A'*ŭʪE*


D,ɤBE$;$goa-1; gpa-1 gpa-3 ˒ĴȞµ/*?ASJ 2ǷũſɅƁńÐ/ǖƠ*.

,B ASJ +ǕɫD¨2ƻĎŅ2 Gα esWʨĿĥ"E goa-1; gpa-1 gpa-3 ˒ĴȞµ2 ASJ +Ǖɫ*DĈɫƁ?ɤBED17B 2372åȀ/ŏD Max 2Ȟţ3 gpa-1 ĴȞµ2<+ʑōE$ƪ+23B 1772åȀ/ŏ*3goa-1gpa-1goa-3-27-ĴȞµ8*/

* Min ;$3 Max-Min 2ȞţʍBE$(ġ 20A-C, G, H)(;CEB2 Gα esWʨ3ɑɪÕ2S^NȂũHŅţȋƉ7ƌ˦/Ǖɫ*ĈɫƁɤBE$XKnˏ^Wc2ĴȞµ/(D*?ʒǃHʂ'$,F

1723172ǷũåȀ/ŏ* daf-11 ĴȞµ3 Max /ȞţHȳodr-1 ĴȞµ
3 Min , Max-Min /ȞţHȳ$(ġ 22)ƪ2317232ǷũåȀ/ŏ*3daf-11 ĴȞµ/

*2< Min-Max /ȞţʍBE$(ġ 23) (Ujisawa et al., 2016)

 4*4)$*/8>,LrC  [~17>:<jSXt
{`{_P`ic(PDE)3XKnˏ^Wc2Ǖɫ,3ʹ/cGMPHíǠÝʒD,+ɑɪÕ72S^NǮÏHƏäD,+ȭBE*2ĴȞµ2´ǷɦƁȞţ3 Gα @ GC 2ĴȞµ,ĎeMx2Ȟţ+
ASJ ƈʐn/

DPDE'$,B* PDE ǷũƅĬ°˂/ƏäȤ/É,¬Ņ$ĭčPDE

2ĴȞµȳ´ǷɦƁȞţ2ʄȑĨHʝƲD/3ȬȪ'$"+PDE ǖƠD,+ƽŰ/ ASJ ƈʐnˁêǭƁò*

D2-HS

^NM_X/ACǸŅ$1723172åȀH$ǀ­/
pde-2 ĴȞµ, pde-3 ĴȞµ/** Max 2ȞţʑōE$(ġ 24B, C, G)2ÈĐ

3 pde-1 pde-5; pde-2 ˒ĴȞµ+?ʍBEB/ pde-1 pde-5; pde-3; pde-2 ĝ˒ĴȞµ+3ȞţıůE$(ġ 24E, F, G)B/pde-1 pde-5; pde-3; pde-2 ĝ˒ĴȞµ+3Min , Max-Min 2Ä?´.'$ (ġ 24F, H, I)ʻţPDE ǖƠD,+ƈʐnÕ2 cGMP ÝʒE

CNG gp˛Ć$ȋƉ/.D$>

-28-

pde-5 2ĴȞµ/ʍBEDA/ɑɪÕS^NȂũ̀ȋƉɚƓED,ƆŅED¥Ğ2S^NM_X2ɕǅBPDE ǖƠD,ƈʐnÕS^NȂũ´*D,ȳE$ɭǍðȉ2ʏɑɪ+3ɑɪÕ72S^NMR2Ȟţ.ǮÏS^NɕčesWʨ.-H¦*
GC HƏä"2ɕǅS^NMR2ǮÏHĽD,ĬĒE*
and Stryer, 1988; Nakatani and Yau, 1988)2,BC. elegans /D(Koch*?PDE 2

ǖƠ/ACɑɪÕ72S^NMR2Ȟţ.ǮÏʪCE GC HƏäD
,/ACASJ ƈʐn2ǭƁòʪ*DĈɫƁɤBE$2A

.ȑʣ3 2317232åȀH$ɕǅ/?ʍBEpde-3 ĴȞµ+3˓ȕǈ
AC? Max 2Ä̀.DȞţʍBE$(ġ 25C, G)pde-1 pde-5; pde-3; pde-2 ĝ˒ĴȞµ+3Ä´.'*$(ġ 25F, G)

ASJ ƈʐn2S^NȂũ3 17B 2372 warming Ƴ/ư23B 1772 cooling Ƴ/3ǵŕDpde-3 ĴȞµ+3231723
2 cooling åȀH$ĭč/˓ȕǈAC?ĂſƁŮ.'$(ġ 25C, G)ƪpde-5
ĴȞµ+31723172 warming åȀH$ĭč/˓ȕǈAC?ĂſƁů
.'$(ġ 24D, G)E3PDE-3  cooling åȀ/ŏ*2Əäğľ,*Ǖɫ
PDE-5  warming åȀ/ŏ*2Əäğľ,*Ǖɫ*
BE$EB2ɕǅ3Ķ2ðȉ/
,D,Hʄ*

D,ɤ*PDE S^NǮÏHƏäD

ǕɫH?(,,ȬȪ.'$Ǘ/PDE-3 @ PDE-5 ˇ°ŀȤ/ G esWʨ2Ǯ+Ǖɫ*

D2HȲ>D$>/gpa-1 ĴȞµ, pde-3 ;$3 pde-5

ĴȞµ2˒ĴȞµH·ʇASJ ƈʐn2ǷũſɅƁHʒǃ$(ġ 26, 27);pde-5; gpa-1 ˒ĴȞµ/ŏ* 1723172ǷũåȀH$ĭč-29-

2˒ĴȞµ3 pde-5 ĴȞµ,Ďǒ2ǷũſɅƁHȳ$(ġ 26)B/pde-3; gpa-1
˒ĴȞµ/ŏ* 2317232ǷũåȀH$ĭč/?2˒ĴȞµ3 pde-3 ĴȞµ,Ďǒ2ǷũſɅƁHȳ$(ġ 27)(;CÝľɔʫ/
3 G esWʨAC?Ǯ/³ɡ*ȞË³(epistatic)+

D$>ȵɔǭðy/

* PDE*? PDE 2Ĵ'$,ɤBED(Ujisawa et al., 2016)

;+2ɕǅHġ 28 /;,>$ASJ ƈʐn2 G esWʨɔʫ2Ýľ3ʈƧ2ÝľÓĎ*ǕɫǷũĴòƳ/ǕɫD?2@S^NȂũHŅţȋƉ7ƌ˦/ǕɫD?2.-ȕȓŀȤ/ǒ.Æ˱/
ųëHƒ'**"E#E2

D,S^NM_XBȳęE$ASJ ƈʐn2ÎąŉƅĬ°˂/˝GDÝľɔʫ,ǝʰ*<D,Î3Îąŉµ LITE-1 +ąĄBEGα esWʨ,* GOA-1 , GPA-3XKnˏ^Wc,* DAF-11 ,ODR-1{`{_P`ic,* PDE-1PDE-2PDE-5 ǕɫDǷũƅĬ°˂/3 GPA-1 , PDE-3 ?ǕɫD,G'$B/pde-3 ĴȞµ3 coolingåȀ/(ġ 25)pde-5 ĴȞµ3 warming åȀ/ŏ*ȞţHȳ$,B(ġ 24)ǷũåȀ2Ȼ˺/A'*ǕɫDÝľȞ.DĈɫƁ?ɤBE$¥ĞȗĴȞµ/* ASJ ƈʐn2Ƿũ/ŏDſɅƁńÐ/3ǱǼ*$.

,B¨2ƻĎŅ2 Gα @ GC  ASJ ƈʐnÕ/ĿĥDĈɫƁ?ɤB
EDB/ASJ ƈʐn/DǷũąŉȵɔǭð,´ǷɦƁ2ʄȑĨ3ń
Ð/ɵ.

ˌÝ?ĿĥDŵʷ2A/´ǷɦƁ3ʈƧ2n@ɓɠH¦*äŹE*

D$>ASJ ƈʐn2ȵɔǭð2ĴòÄHse,*´ǷɦƁ2ʄȑĨHńÐ/ʝƲD/3¨2n@ɓɠ2se
@´Ƿǚ2þğ+

DKNkxhk2seHƲȲ/DŽʋ

-30-

D,ɤ

BED

Bj{Yuy2.0;>&HV;+2ʒǃB´ǷɦƁ2ƋɁ/3 ASJ ƈʐn˝D,ȳE$ASJ ƈʐnȨƜȤ/ǷũHƈȭ* ASJ 2ȵɔǭðHĴò!*D,ɤBED¨2nǷũHąŉ*"2ǷũąŉƅĬ ASJ

ƈʐn/°˂EASJ 2ȵɔǭðHĴò!*DĈɫƁ?ǛE*$"+ASJ ƈʐn«ĵB2^mx`H¦$ƅĬ°˂ǖƠ$ȋƉ+
ASJ ƈʐn2ǷũſɅƁHʒǃ$ÔµȤ/3^mx`°˂/˝GDĴȞµ,*^mxkwu/SNB-1ȵɔzxglčƋˎɎ/EGL-21^mxkeX
/SNT-1^eU^/UNC-64 Hȗ*ʒǃHʂ'$(ġ 29)ȵɔɑɪ˜2ƅĬ°˂

3ȵɔƜəˌ+.GEȵɔɑɪ2ɶļ/ſ*ȵɔ°˂ȉʨ3^mx`ŔɪÕ/ȂɝEDŔɪɲ/3^mxkeX,^mxkwuĿĥ"Eɑɪɲ/D^eU^,ʈčµHűƋ*ɲʁčHʪȵɔ°˂ȉʨH^mx`˜˥/ƤÜD,+ŵɑɪ7,ƅĬ°˂ʂGE*`Ŕɪɲ2ǑƋÝľ+D"+;^mx

D^mxkwu/ȞţH?( snb-1 ĴȞµ/*ASJ

2S^NM_XH.

ASJ ƈʐn«ĵB2^mx`°˂H

ƨ'$ȋƉ+ASJ ǷũåȀ/Ăſ+DHʞ8$(ġ 29A, B)"2ɕǅsnb-1
ĴȞµ2 ASJ n3˓ȕǈ,Ďǒ/ǷũĴòåȀ/ŏ*ſɅHȳ$$>ASJ ƈʐnúȍ+ȨƜǷũHąŉ*-31-

D,ȳęE$B/snb-1

ĴȞµHđ>egl-21 @snt-1 , unc-64 2ČĴȞµ3´ǷɦƁ2ȞţHȳ$(ġ 29C)
(;CASJ ƈʐn3úȍ+ǷũHƈD,+ASJ ƈʐn2^mx`H¦*ÝǦƁ2ǷũƅĬǮ2ɑɪ7°˂E*DĈɫƁɤBE$
ASJ ƈʐnBÝǦEDÝľ@Ǯɓɠ2ʒǃʂGE´ǷɦƁ/3 ASJ ƈʐn2Ǯɓɠ,*ɰ@ȵɔɌ˝*D,G'$(Ohta et al., 2014) (ġ 30A)ASJ nBÝǦEDM`Ýľ INS-6 ,DAF-28 ɰ/DM`ąŉµ DAF-2 /A'*ąŉED"+3DAF-2

2Ǯ2M`^Xm°˂ɔʫHɔ*ƅĬ°˂EFOXO ĨʮÖğľ/ADˇ°ľ2ȣȑäŹʂGED,+´ǷɦƁäŹED,ȳęE*

D2´ǷɦƁ3ɍ 3 Ƴ˜+ȏŸǱĺED$> (ġ 4)ʈƧ2ɓɠH¦*´ǷɦƁHȏŸD$>/3ASJ ƈʐn2ȵɔǭðĴòƳȤ.?2+öÝ.2˘'$"+15˼ɩÀµ/ 17B 23Ƴ˜/G$CŽʋ.2Hʞ8DŽʋ2ǷũåȀH"2ŵ5 Ƴ˜ 232ǷũåȀHə$ĭč2 ASJ ƈʐn2ȵɔǭðHS^NM_X+ʞ8$(Ujisawa et al., 2014a) (ġ 30B)ǷũåȀHDè,ǷũåȀŵ 30 Ý1 Ƴ˜3 Ƴ˜5 Ƴ˜2Ƴȃ+2 ASJ 2ȵɔǭðȋƉHʒǃ$ɕǅ30 Ýŵ/ ASJ 2ǭðƸķ,.Cɍ 1 Ƴ˜ŵ/ŅţȋƉ/ƌD,G'$;$ģȘB2ʒǃB´ǷɦƁ/3ɰ%+.Ɋľ˝
GD,G'*$Ɋľ2ĴȞµ+lD GSP-4 2ĴȞµ/

DxiM{`vJec 1(PP1)H[* ASJ ƈʐn2S^NM_XʒǃH

ʂ'$,FǷũåȀ/ŏDſɅƁ´*$(Sonoda et al., 2016)2ſɅƁ2´3ɊľȊȞȤ/ˇ°ľȣȑHʛœDxe(spe-11p)Hȗ

-32-*gsp-42ɊľȊȞȤ/ gsp-4 ˇ°ľHȣȑ!D,+Ğź$2,BɊľ˸ˌ
2 ASJ Ƿũąŉn2ȵɔǭð/Ų˲H*DĈɫƁɤBE$

B/XKɑɪ ASJ ƈʐn2Ƿũąŉ/DŲ˲Hʞ8$
ĉ,ĹȘB/A'*XKɑɪ+ǕɫD{RlMĨʮÖğľ PROS-1 2´ǷɦƁ72˝ʍ('$pros-1 3XKɑɪ(AMsh)+ȣȑEǖƠD,
ɵǚ,.D$>qhWKNkĴȞµHʒǃ/ȗ

D,3+.

"+feeding

RNAi /AC eri-1; lin-15B Ɍɗ2 pros-1(ceh-26)ˇ°ľHqhWfN!2qhW
fNÀµ2 ASJ ƈʐn2ǷũſɅƁHS^NM_X+ǸŅ$(Kage-Nakadai et al., 2016) (ġ 31)"2ɕǅ15@ 20+˼ɩ$ pros-1 qhWfN
Àµ2 ASJ ƈʐn2ǷũſɅƁ3qhWfNH*
2Ɍɗ,Ďǒ2ĂſƁHȳǷũſɅƁ3Ǚţ+.

eri-1; lin-15B

D,G'$pros-1 qhW

fNÀµ3´ǷɦƁ/ȞţHȳ15˼ɩŵ/ 2+ȕĿȐ´DȞţ+'$2Ȟţ3ASJ n2ǷũƅĬ°˂2ĴȞµȳ 20˼ɩŵ/ 2+ȕĿȐưDȞţ,3ʨȤ/Ȟ.'$EB2,Bpros-1 ȣȑ*

D

XKɑɪ(AMsh)3 ASJ «ĵ2n/Ų˲HD,+´ǷɦƁ/˝*D,ɤBE$ ?hANc
ASJ ƈʐnB3M`ǒÝľ DAF-28 , INS-6 ȣȑ*D$>ĹȘB/ACM`Ýľ2ʒǃʂGE$B/M`ąŉµ+-33-

D DAF-2 2ĴȞµ+ 20@ 25˼ɩŵ/´ǷɦƁưDȞţʑōE$"+daf-2 ĴȞµ2ɰ@ȵɔɌ+ȊȞȤ/ daf-2 ˇ°ľHȣȑ!$ɌɗH·ʇ´ǷɦƁ2ʒǃʂGE$"2ɕǅ´ǷɦƁ2ƋɁ/3ASJ ƈʐnBÝǦE$M`ɰ@ȵɔɌ2M`ąŉµ(DAF-2)+ąŉEM`ƅĬ°˂ɌH¦*ƸɒȤ/ FOXO ĨʮÖğľ(DAF-16)/ADˇ°ľȣȑäŹŽʋ+D,ȳęE$´ǷɦƁ2ƋɁ/M`ƅĬ°˂ɌH¦$ˇ°ľȣȑŽ˶+

D,B´ǷɦƁ/

*DAF-16 2Ǯ/³ɡDˇ°ľHʍ(D$>2ʒǃHʂ'$ˁÿ/ʂGE$ƾB2 DNA ~MWKMʒǃ/AD,
23B 17/˼ɩǷũHĴò!$˦/ȣȑ˔ĴðDˇ°ľ 79 ÀĿĥ$(Sugi et al., 2011)EB2ˇ°ľ&2ŕ.,? 20 À2ˇ°ľ/(ĿĥÒʄE*$$>"EB2ĴȞµ/(*3ĴȞµ2*´ǷɦƁHʞ8$"2ɕǅX[^blčƋˎɎ CGT-1^`iMxiKc CPR-1tlU^`iMl 17β ɮǠɎˎɎ DHS-4Xb-3-ˏɮǠɎˎɎ GPDH-1 H"E#E[
lDˇ°ľ2ĴȞµ/*20+˼ɩ$Àµ2´ǷɦƁ/ȞţʍBE$ƪdoublesex ĨʮÖğľ2 DMD-7 @ Gα esWʨ2 GPA-7 H[lDˇ°ľ2
ĴȞµ+325+˼ɩ$Àµ2´ǷɦƁ/ȞţʍBEPl|oWKc+

D M60.2 H[lDˇ°ľ2ĴȞµ+320+˼ɩ$Àµ, 25+˼ɩ$Àµ2ƪ+´ǷɦƁ2ȞţʑōE$(ġ 32)Ƿũ/»Ŀ*ȣȑ˔ĴðD
ˇ°ľ2/3Ƿũ`k`Hƈ$˦/ǕɫD,+ȭBE*WesWʨ(HSP)H[lDˇ°ľ+
đ;E*

Dtk^hD hsp-16.2hsp-16.41"* hsp-16.48 $EB 3 (2ˇ°ľ2ĴȞµ3"E#E´ǷɦƁ/Ȟţ3ȳ.'$(ġ 33)hsp ˇ°ľ2ʮÖʞɇ/˝GDÝľ,*tk^hWʮÖğľ

-34-

D HSF-1 +CHSF-1 H[lDˇ°ľ2ĴȞµ(hsf-1)+?´ǷɦƁ/Ȟţ3

ʍBE.'$(ġ 33)ĴȞµ´ǷɦƁ/ȞţHȳ$ 7 (2ˇ°ľM`ɔʫ2Ǯ+ȣȑʞɇHE*

D2Hʞ8D$>/Ņ˔Ȥ PCR ǧHȗľ2 mRNA 2˔H˓ȕǈ,M`ąŉµ daf-2 ĴȞµ/*EB 7 (2ˇ°*ǸŅ$daf-2 ĴȞµ3̀Ƿ+˼ɩD, dauer /.'*;$>˓ȕǈ, daf-2 ĴȞµH"E#E
15˼ɩÀµ15+ L4 ;+˼ɩŵ 20+ 12 Ƴ˜˼ɩ$Àµ;$3 15+ L4;+˼ɩŵ 25+ 12 Ƴ˜˼ɩ$ÀµB"E#EµÕ mRNA HƑÜŅ˔Ȥ
PCR ʒǃHʂ'$(ġ 34);$ gpa-7 ˇ°25˼ɩŵ2´ǷɦƁ/ȞţHȳ*ľ2 mRNA ˔H˓ȕǈ, daf-2 ĴȞµ+ǝ8$,F25+˼ɩ$ daf-2 ĴȞµ/* gpa-7 2 mRNA 2˔˓ȕǈ/ǝ8* 10 Á«/ıí*+1520;$3 25+˼ɩE$˓ȕǈ/$,F˼ɩǷũ2˃/ADȣȑ˔2˃$ (ġ 34B)ƪ* gpa-7 ˇ°ľ2ȣȑ˔Hǝʰ3ʍBE.'$(ġ 34C)EB2

ɕǅBdaf-2 ˇ°ľǖƠD,/ACM`ƅĬ°˂ɌǕɫ..C
25˼ɩE$ÀµÕ+ gpa-7 ˇ°ľ2ȣȑ˔ıí"E´ǷɦƁ2Ȟţ/˝ʾ*D,ɤBE$Ďǒ/daf-2 ĴȞµ+3ĴȞµ´ǷɦƁȞţHȳ$ 7(2ˇ°ľ29,I-/˝*"2ȞţHȳ$˼ɩǷũ/Dȣȑ˔ıí*$ƪĴȞµ 20+˼ɩE$˦/´ǷɦƁ/ȞţHȳ*$ cpr-1 ˇ°ľ/˝*320˼ɩÀµAC? 25˼ɩÀµ2ƪdaf-2 ĴȞµ/Dȣȑ˔ıí*$2,Bcpr-1 ˇ°ľ/˝*3´ǷɦƁ/Ȟţ/3Ž?˓ȕǈ,2ȣȑ˔2ŠķDŽʋƁ.

,ȳęE$

DAF-2 2Ǯ/3 PI3 Umc2 AGE-1PDK-1AKT/PKB Umc2 AKT-1,-35-

AKT-2"* FOXO ĨʮÖğľ2 DAF-16 7,ɟDM`^XmɔʫĿĥD(Fielenbach and Antebi, 2008; Malone et al., 1996) (ġ 30A)2ɔʫ3̀ǷɦƁ(dauer)Ƴ/?Ǖɫ*CɑɪÕ/HäŹD,+ɦƁäŹE*

D DAF-16 ǉÕ/ȹʂǮ2ˇ°ľȣȑD,ĬĒE*

D$>¥Ğʞ8$Ƿũ»ĿȤ/ȣȑ˔ĴðDˇ°ľ3DAF-16 /ACäŹHą*DĈɫƁɤBED
ɑɪɲ2ǮðƁHǣ>D˽Ėɬɨˏ.-2ɬʨɲǑƋ2Ĵò3Ķ2ȕ
ȉ2´ǷɦƁ/,'*˒ʋ+Dɬɨˏ˽ĖòˎɎ+Ķ2ȕȉ2´ǷɦƁ/˒ʋ.ųëHǅ$*,D Δ9 j\gc3

DC. elegans /? fat-5fat-6fat-7

 Δ9 j\gcH[lDˇ°ľĿĥ"EB3DAF-16 /A'*ȣȑäŹHąDǓȤˇ°ľ,*ȭBE*ǷɦƁ?´D,

GE*

DEBǖƠD, C. elegans 2´D(Savory et al., 2011)2ĬĒB¥Ğ2´ǷɦƁƳ/?ɑɪɲ2ǮðƁHĴ*´ǷɦƁHȏŸ+DA/Ĵò*BE$"+Ņ˔Ȥ PCR Hȗ*EB2ˇ°ľ2ȣȑ˔H˓ȕǈ, daf-2 ĴȞµ+ǝʰ$,F20;$3 25+˼ɩ$ daf-2 ĴȞµ/2ȣȑ˔ư*DĈɫƁɤ

$(ġ 35)

-36-*fat-5 ˇ°ľzQ 













 [~17>:<HVBj{Y

C. elegans 2´Ƿ/ŏDɦƁǕǑ3E;+/ȭBE*ÔµȤ.Ƿũ.-2ǀ­HǎʘDŽʋ

.'$$>'$¥Ğ15+˼ɩE$Àµ3 2

2´ǷåȀH*?ȕĿ+20@ 25+˼ɩE$Àµ3 22´ǷåȀHD,ǚǼD,G'$(ġ 12)(;C2´ǷɦƁȑʣ3˼ɩE$Ƿũ/»Ŀ*

D,G'$B/2´ǷɦƁ3G

 3 Ƴ˜+ȏŸED,

G'$(ġ 4)E;+/Ƿũ`k`HD, HSF @ HSP Ǖɫ`k`/ADŲ˲BɑɪH¿ʡD,3¨2ȕȉ+?ȭBE*ǷɦƁ/˝*3EB2Ýľ3˝*.

D¥Ğ2´

,GC(ġ 33)2 3 Ƴ˜,ȮƳ˜+ˇ°ľ2ʮÖʂGE-2A.µÕSnaʪ'*
G'*

.

;

D23

-2ɓɠǕɫDHƛCȵɔɌ˒ʋ+D,G

'$Ƭ/Ƿũąŉ/˝½Dȑʣ2(,*ǷũʩƁ

C2ȵɔĞʫ/GD AFD Ƿũąŉn2ĴȞµHʞ8$´ǷɦƁ3Ǚţ+'$(ġ 8)2A/ǷũʩƁ/˝GD AFD Ƿũąŉn2ǕɫǖƠ**?´ǷɦƁƋɁD,BAFD Ƿũąŉn3´ǷɦƁ/˝.
,ɤBE$ǷũʩƁ2ȵɔĞʫ/G*í*¨2ƈʐn2
Ǖɫ/˝GD cGMP »ĿƁgp2 tax-4 ĴȞµ+320˼ɩŵ2´ǷɦƁ/Ȟţ
<BEtax-4 ĴȞµ2 ASJ ƈʐn+ tax-4 ˇ°ľHȣȑ!$k`_O
nhWɌɗ+3tax-4 ĴȞµȳ´ǷɦƁ2ȞţĞź$(ġ 11),F¨
2 AWC ƈʐn+ tax-4 ˇ°ľHȣȑ!AWC n2ǷũąŉHǙ

-37-

ţ/ƌ$Ɍɗ+3´ǷɦƁ3Ğź.'$(ġ 11)(;CǷũʩƁ/˝GD
AWC n?´ǷɦƁ/3˝.'$EB2ʒǃB?ǷũʩƁ/Ž
ʋ.Ƿũąŉn,3â2n´ǷɦƁHäŹD,G'$

 [~17>:<EZKFo
tax-4 ĴȞµ2`UɌɗ@S^NM_X2ɕǅ.-B´ǷɦƁ/Îąŉn+D ASJ ƈʐn´ǷɦƁHäŹ*D,ƲB,.'$B/3 ˔µ G esW.-2ƈʐƅĬ°˂/¸GEDÝľ/˝*?ASJ /DÎ,Ƿũ2ƅĬ°˂/

*9,I-Ď?2Ǖɫ*,ȳęE$(ġ 16)ąŉµ3Ȟ.CÎąŉµ+,´ǷɦƁ/3˝*.

D

D LITE-1 3Ƿũąŉ(ġ 1516)ɷȤ/ 3 ˔µ G esW2Ǯ/3GesWÓȦĨąŉµ GPCR 

DASJ 2Ƿũąŉµ3;%ʍ('*ũʩƁʂð2ʋ.Ƿũąŉn+.

Ƿ

D AFD ƈʐn+3bxeĨXKnˏ^Wc(rGC)Ƿũąŉµ2ųëH*

D,

GE*

D(Takeishi

et al., 2016)ASJ ƈʐn+3ɷȤ/ GC 2Ǯ/³ɡD Gα HǖƠ!D,+ǷũąŉƅĬ°˂´D,BGC , Gα 2Ǯ/3¶B2ǷũąŉµǕɫ*D,ȳęE$

ASJ ƈʐnǷũ,Î,Ȟ.DƅĬHąŉD,B(

2ȟě,*Î,Ƿũ2ƅĬ ASJ Õ+W`kW*E*

D2"E,?ôâ

D2ƕBE$ƍřȰȾŇ2rǷũ,ÎHĎƳ/Ű*Dņ˿-38-

,*ASJ ƈʐn+ąŉDˮɸÎ,ǷũåȀHĎƳ/´ǷɦƁ/Ų˲ÜDʞ8$Ų˲3Ü.'$2ɕǅ%+3ƨŅ3+.
Î3ôâE**D?ȭE.

Ƿũ,

ASJ ƈʐn2+Î,Ƿũ2ƅĬHôâ

Dƪǧ,*Ýľ2ÉÝɤBE$¹4ASJ ƈʐn3̀Ƿ`k`Hƈ$˦/ʪDɦƁŦɿ2Sna/?˝*ʣ/DɦƁŦɿȑ*  ASJ 2 X K n  ˏ ^ W   c DAF-11  cGMP » Ŀ Ɓ g  p TAX-4/TAX-2 H¾ʿDɔʫÉɦƁŦɿǕǑ/˝GDXKnˏ^Wc3
DAF-11 «ĵ/?ʈƧĿĥDAF-11 , ODR-1 3ȣȑɑɪ˒.'*ɪ2+ÉHôâ*D,ʓGE*

DĎɑ

Dņ˦/¥Ğ2´ǷɦƁ2ʒǃ/

*

DAF-11 2ĴȞµ/ǝ8* ODR-1 2ĴȞµ2ƪ´ǷɦƁ2Ȟţů'$̄ġ 16B̅
(;C̀Ƿ`k`/3 DAF-11 /É´ǷɦƁ/3 ODR-1 /É,'$¸ÝʂGE*DĈɫƁɤBE$"2A.ȓș+ASJ 2 G e

sWʨɔʫ2ĴȞµ/D ASJ ƈʐn2S^NM_XHʂÎƅĬ,ǷũƅĬ+ǕɫDÝľȵɔǭðy+ÝE*

D2ʒǃɕǅ,

*G esWʨɔʫ2ÝľǷũåȀ/DǷũĩ@ǷũĴòse.-;;.Æ˱/*"E#E¸ÝBE*Dĭč@ÓĎȤ/Ǖɫ*DĭčD,G'$(ġ 20-25)
ƽʒǃ+ȗ$ĴȞµ2´ǷɦƁ2Ȟţ,ASJ ƈʐn2ǷũſɅƁ2S^NM_X2ʒǃ+ŸBE$Ȟţ2ů.-ɵ.

ˌÝʍ

BE$¹4Gα esWʨ+3 goa-1 ĴȞµ, gpa-1 ĴȞµ+3´ǷɦƁ2Ȟţ
ʍBEgpa-3 ĴȞµ+3;C´ǷɦƁ2Ȟţ3ʍBE.'$(ġ 16)

EB2ĴȞµ2 ASJ ƈʐn2ǷũſɅƁHS^NM_X+ʞ8-39-

D,Ð*2ĴȞµ+ˌÝȤ+3DȞţʍBE$(ġ 20212425)goa-1; gpa-3

gpa-1 ˒ĴȞµ2´ǷɦƁ tax-4 ĴȞµ9-2ȞţHȳ/ǖƠ*.'$ɕǅBASJ ƈʐn/ASJ 2ȵɔǭð?ńÐ

*¨2 Gα esWʨĿĥDĈɫƁɤBE$XKnˏ^Wc/˝*3daf-11; odr-1 ˒ĴȞµ/*˹ɻ.´ǷɦƁ2ȞţʍBE$(ġ 16)daf-11; odr-1 ˒ĴȞµ2 ASJ 2ǷũſɅƁ/3˹ɻ.Ȟţ3ʍBE.'$,BXKnˏ^W
c?¨/Ŀĥ*D?ȭE.

(ġ 2223)Gα esWʨ@XKnˏ^W

c.-3ASJ «ĵ2¨2Ķ2n+?ȣȑ*

D$>´ǷɦƁ2Ȟţ,*ʄED/3ASJ nÕ2Ȟţ%+3.¨2n@ɓɠ2Ȟţ?Ų˲*ʄEDĈɫƁ?ɤBED

Bj{YuyĹȘB2ʒǃBǷũåȀ/AC ASJ ƈʐnBM`ǒÝ
ľ+

D DAF-28 , INS-6 ÝǦEɰ@ȵɔɌ+ąĄBE´ǷɦƁHäŹD,ȳęE$(ġ 30A)2M`ǒÝľ3tk2M`/²$ǑʽH?(M`ǒ{,Ĕ4E*Cnzxgl2A/ȵɔɌ/·ȗD,ĬĒE*ąŉµ+

D (Kodama et al., 2006)M`ƅĬ°˂ɔʫ+3M`

D DAF-2 2Ǯ+FOXO ĨʮÖğľ DAF-16 /ADˇ°ľ2ȣȑä

Ź.-ʂGEÐʭ2´ǷɦƁäŹE*

D,ɤBED´ǷɦƁ3ɍ 3 Ƴ

˜+ȏŸǱĺED5 Ƴ˜2S^NM_X2ɕǅBASJ ƈʐn-40-2ȵɔǭð3ǷũĴòHƈ*Bɍ 30 Ý+ǭðƸķ,.C,.C"2ŵ
3ŅţȋƉ/ƌD,G'$ʍD,ASJ ƈʐn2ȵɔǭð,´ǷɦƁ2ȏŸǱĺ/DƳ˜2˜/Šȕ*DE3ASJ ƈʐn

B2ǷũƅĬM`Ýľ/ACǮ2ɓɠ7°˂E"2Ǯ+3B/
DAF-16 /ADˇ°ľ2ȣȑH±äŹ.GE"B´ǷɦƁ/Žʋ.ȕµʞɇ.GED$>Ƴ˜2aȕ*D,ɤBED

M`ƅĬ°˂ɔʫ3̀ǷɦƁƳ/?ǕɫDɥ2˃̀ǷɦƁ2ĭč/32M`ƅĬ°˂ɔʫ3 L1 Ŧɿƹ/
ŦɿűƋ,

,*

*ǕɫdauerűƉ2ĴòH±´ǷɦƁ/3"2A.ʍ$ȧ2űƉȤĴ

ò3<BE.

DAF-16 /ʞɇHąDˇ°ľ3˰ţ/Ķ

j\gc2 fat-5fat-6fat-7 ˇ°ľ+D,´ǷɦƁ?´D,ʓGE*D´ǷɦƁˇ°ľ/(

"2&2( Δ9

D(Savory et al., 2011)EBǖƠD"2¨2 DAF-16 2Ǯ+ȣȑäŹE**3ʒǃʿʂ+DE;+/´ǷɦƁ/ȞţHȳ$ĴȞµ2þğˇ°ľ2+DAF-16 2Ǯ+Ǖɫ*Ƿũ/»Ŀ*ȣȑ˔ĴðDˇ°ľ+Dˇ°ľ2Ãʆ,*3

D gpdh-1(Xb 3 ˏjtlZmc)@ cpr-1(^`iMxiKc).- 7 (2ˇ°ľʍ('$(ġ 32)Ưɿ2´ǷɦƁǕǑ2(/ɑɪÕ/Ùòȉʨ,*ǒ.Ȼ˺2ɋHɽȼ!ɑ
ɪÕÙɕHĞˈ$Cµǲ2Xb,ɑɪÕ2ǠHɡƞ$CD,+ɑɪÕÙɕHˠ,G'*DC. elegans /*ǰʺĤ3excretory cell ,Ĕ4EDǰʺĤHʞɇDɑɪ+ʞɇE*Cdoublesex ĨʮÖğľ2 dmd-7 @b
lX[^k`vOc2 cgt-1 ˇ°ľ32 excretory cell +?ȣȑ*DC. elegans 2µ3ǰʺĤ+ʞɇE*C̀-41-Ǯµ˯ìŀȤ.ÕĤ+ɚƓE

*DEB2ˇ°ľ2ÕĤHɚƓD$>2 excretory cell +É´ǷɦƁ/˝*DĈɫƁ?ɤBEDcgt-1cpr-1dhs-4dmd-7gpa-7gpdh-1M60.2

ˇ°ľ3C. elegans ˓ȕǈH̀Ƿ(23)B´Ƿ(17)/ȹ$,/ȣȑ˔ıǵD(Sugi et al., 2011)EB2ˇ°ľ2ĴȞµ+Dcgt-1cpr-1dhs-4gpdh-1 ĴȞµ3 20˼ɩŵ2´ǷɦƁ2ȞţHȳdmd-7 ĴȞµ, gpa-7 ĴȞµ325˼ɩŵ
2´ǷɦƁ/ȞţHȳM60.2 ĴȞµ3 20;$3 25˼ɩŵ2´ǷɦƁ/ȞţHȳ$EB2ɕǅ,Ņ˔Ȥ PCR 2ɕǅHčG!D,dhs-4 @ dmd-7gpa-7 ˇ°
ľ3"EB2ĴȞµ2´ǷɦƁ/ȞţʍBE$Ƿũ/Dȣȑ˔̀.'**daf-2 ĴȞµ/D(ġ 34)ƪ+cpr-1 @ gpdh-1 ˇ°ľ.-´ǷɦƁ2Ȟ

ţʍBE$Ƿũ,3Ȟ.DǷũ+ȣȑ˔̀.'*Dˇ°ľ?

D¥Ğ2Ņ˔Ȥ PCR 3ÀµÐµBĞă$ mRNA H?,/ǸŅE$?2+

D$>´ǷɦƁ/˝GDɓɠ+3EB2ˇ°ľ2ȣȑ˔ÀµÐµ+ʍBEDȣȑĴðse,Ȟ.DĈɫƁ?ɤBE$ƽȰȾBƲB/.'$ C. elegans 2´ǷɦƁ2äŹǕǑH;,>D,
ASJ ƈʐnǷũHąŉM`ÝľÝǦE"Eɰ+ąŉED,+ˇ°ľȣȑH¦*Àµ2´ǷɦƁƋɁD,a+Ʃʎ2ǷũäŹSn

D(ġ 36)2 ASJ ƈʐn/»Ŀ$Ƿũ^Xm3ƸɒȤ/´ǷɦƁHƏäDɔʫ+

D(;C´ǷɦƁ/3¨2ɑɪH¦$Ǚ2ɔʫ

2ĿĥɤBEǙ,ʥ2ƪ2ɔʫH¦$ʈ˪.Sna´ǷɦƁ2ƋɁ/
3Ž˶+D,ɤBED-42-

w

ƽȰȾBC. elegans 3˼ɩE$Ƿũ/»Ŀ*´Ƿ/ɦƁH?(,G'$B/E;+¨2ðȉ+ƻȭ+'$ȵɔɌ/DǷũƅĬ2Mx

hkBɰ7(.DÝľɔʫ´ǷɦƁ/˝GD,G'$E;+/ÀµHǑƋDÀɑɪ+ɬɨˏčƋˎɎɑɪɲ2ɬɨˏ/·ȗɑɪɲ2ǮðƁHĴD,+´Ƿ+?ȕĿ+D§ɓ<D,ȳęE*$ (Savory et al.,2011)¥Ğ2ʒǃBČ2ɑɪ/DǷũſɅÀµ2´ǷɦƁHäŹ
D2+3.ÎąŉnǷũHƈȭ"2ƅĬɰ/°˂ED,+´ǷɦƁäŹED,

Ʃʎ2´ǷɦƁ2ʒǃjHȲɁ$ASJ ƈʐnÕ+3 G esWʨɔʫH¦*ǷũƅĬ°˂ʂGED3 ˔µ G esWʨ2Ǯ/ĿĥD,ɤBEDǷũąŉ/˝GDąŉµ3;%ʍ('*^N_NrP+3Îąŉµ+Dlx^Ƿũąŉ/˝GD,

D(Shen et al., 2011)C. elegans 2 ASJ n/
LITE-1 3Ƿũąŉ/3˝*

.

.



ĬĒ?

*3Ě2Îąŉµ+D,ȳęE$(;CÎąŉµ«ĵ2Ƿũąŉµ2ĿĥȳęE$B/ASJ ƈʐnHđ>*1 ɑɪÕ+ʈƧ
2ƅĬʠâED§ɓ</3ƻȭ2ȃǛE*C¥Ğ2 ASJ n/DǷũ,Î2W`kW2ņ˿3ƈʐn@¦ĥnʈƧ2ƅĬHąĄC"2ƅĬʠâ*Ǯ7ƅĬ°˂D§ɓ<HʒƲDņ˿Ɍ/ãȗ+D,ɤBED-43-b]=^g

C. elegans 2˼ɩǧĪĦ@ Buffer 2ɓƋɃ3 THE NEMATODE CAENORHABDITISELEGANS /ŷ'$(Wood and Researchers, 1988)

StrainN2 Bristol (˓ȕĨ)
RB1415 catp-3 (ok1612), VC693 cgt-1 (ok1045), RB2095 clec-67 (ok2770), PR674
che-1(p674), CB1033 che-2(e1033), CB3323 che-13(e1805), RB1262 cpr-1 (ok1344), IK038
daf-2(e1370), CB1377 daf-6(e1377), DR47 daf-11(m47), CX2454 daf-11(sa195); odr-1(n1936),
FX2418 dhs-4 (tm2418), RB1772 dmd-7 (ok2776), eat-4 (ky5), eat-16(nj8), KP2018
egl-21(n476), FX2468 F58E6.7 (tm2468), glc-3(ok321), MT2426 goa-1(n1134), NL332
gpa-1(pk15), gpa-1(pk15); goa-1(n1134), NL335 gpa-3(pk35), NL795 gpa-7(tm5828), RB1373
gpdh-1 (ok1558), PS3551 hsf-1(sy441), VC475 hsp-16.2 (gk249), hsp-16.41 (tm1093), RB791
hsp-16.48 (ok577), RB883 kqt-2 (ok732), TQ1101 lite-1(xu7), CZ1758 max-1 (ju142), FX1770
mtl-1 (tm1770), FX4977 M60.2 (tm4977), FX11134 pept-1(opt-2/lon-2) (tm302/e678), CX2336
odr-1(n1936), MT3644 odr-3 (n1605), CX4 odr-7(ky4), PR802 osm-3(p802), PR813
osm-5(p813),

PR811 osm-6(p811), CX0010 osm-9 (ky10), MT3641 osm-10(n1602), IK616

pde-1(nj57), IK618 pde-2(nj58), pde-3(nj59), IK510 pde-5(nj49), IK792 pde-1(nj57);
pde-2(nj58), IK794 pde-1(nj57) pde-5(nj49), IK796 pde-5(nj49); pde-2(nj58), IK798
pde-1(nj57) pde-5(nj49); pde-2(nj58), pde-1(nj57) pde-5(nj49); pde-3(nj59); pde-2(tm3098),
FX830 pgp-9 (tm830), RB908 pmp-1 (ok773), rol-6(e187), NM467 snb-1(md247), NM204-44-

snt-1(md290), FK100 tax-2(ks10), FK127 tax-4 (p678), tax-6 (db60), IK659 tax-6 (db60);
njEx343[tax-6p(1.1kb)::tax-6, ges-1 NLS GFP], IK1053 tax-6 (db60); njEx424[unc-14p::tax-6],
trx-1(ok1449), trx-1(tm1786), IK615 ttx-1(p767), ttx-3 (mg158), ttx-4 (nj1), IK589 ttx-7(nj50),
ttx-8(nj34), T28C12.4 (tm1013), CB1092 unc-54(e1092), MC339 unc-64(md130), CB1265
unc-104 (e1265), unc-112 (r367); gkDf2 (X)ķɰɺOP-50 ǈ: ɜɿ2˾,* NGM xk/Įš$˹Ż˗ʺˁŬņµ˹Ż˗ (Leica: MZ16)ʺˁŬņµʀÎ˹Ż˗ (OLYMPUS SZX12)
Óȅȃ]˹Ż˗(FV1000-IX81 with GaAsP PMT, Olympus)

Nematode Growth Medium (NGM) Plate (1000 ml)
NaCl 3 gAgar (®ˊŌĸ) 20 gBacto Peptone (BD Falcon) 2.5 gH2O 975 ml HǴč
RkWw$ŵ/ Cholesterol (5 mg/ml in EtOH) 1 ml 1M CaCl2 1 ml1M MgSO4
1 ml1M Potassium Phosphate (pH6.0) 25 ml Hí$EHȨŴ 6 cm 2^/ 14
ml ;$33.5cm 2^/ 6 ml(Ýǩ*Ģ>$

M9 buffer (1000 ml)
KH2PO4 3 gNa2HPO4 6 gNaCl 5 gH2O 1000 ml HǴčŵ/-45-

1M MgSO4 1 ml í$´ǷɦƁi`k20+˼ɩ$˓ȕĨ2 C. elegans H1 ;$3 2 Àµ

( OP50 ĮBE$Ʃ

2%(w/v)NGM xk/ǌɘ(P0)Č˼ɩǷũ(152025)+ 8-12 Ƴ˜˼ɩȖü!$P0 Hˣÿü(F1) adult /.D;+Č˼ɩǷũ+˼ɩə$(15+
144-150 h20+ 85-90 h25+ 60-65 h)xk2 F1 70-150 Àµ adult ;+Ƌ˘$ŵNGM xk2ŌĸÆH/Đ$ȋƉ+ǡ/ 20 Ý˜$"2ŵxk/ 2+ 48 Ƴ˜2´ǷåȀH$´ǷåȀŵŇǷ/ĄCÜȕĿÀµ,ǚÀµHƧȕĿȐHÜ$(Ujisawa et al., 2014b)

´ǷɦƁ2Ƿũ^vkKhbMŦɿƹ/*15°C B 25°C 72Ƿũ^vkKhbMHDĭčɜɿHČŦɿƹ;+ 15°C ˼ɩ$ŵ˼ɩǷũH 25°C /^vkƋɿ/.D;+˼ɩ$"2ŵʻţ2´ǷɦƁi`k,Ďǒ/´ǷåȀHȕĿȐHǢ>$Ƌɿƹ/*15°C B 25°C 72Ƿũ^vkKhbMHDĭčɜɿHƋɿ/.D;+ 15°C ˼ɩ$ŵ˼ɩǷũH 25°C /^vk1-5 Ƴ˜˼ɩ$"2ŵ´ǷåȀH
$7-12 Ƴ˜.-˘Ƴ˜ 25+˼ɩDĭč15+ L4 Ŧɿ;+˼ɩ$,F+
25/ȹ$(Ujisawa et al., 2014b)

Molecular biology
trx-1p::tax-4 cDNA(pTOM4)str-3p::tax-4 cDNA(pTOM3) ceh-36p::tax-4 cDNA(pTOM10)-46-

3pDEST-tax-4cDNA(a gift from Dr. Iino) (Hirotsu and Iino, 2005)B PCR +ı@$
tax-4 cDNA , unc-54 2 3’UTR Hđ=pTOM4 2 ASJ ȊȞȤxe,*¸ȗ$ trx-1 xe3pQZ37 trx-1p::ICE (a gift from Dr. Alcedo) (Miranda-Vizuete etal., 2006)B¸ȗ$ pTOM3 2 ASI ȊȞȤxe,*¸ȗ$ str-3 xe(3.1Kb)3pQZ36 str-3p::GFP (a gift from Dr. Alcedo)B¸ȗ$ pTOM10
2 AWC ȊȞȤxe,*¸ȗ$ ceh-36 xe(368bp)3pAK203
B¸ȗ$ŗĥʒǃ/¸ȗ$ trx-1p::gpa-1 cDNA::venus (pUDA14)3site-directed mutagenesis /A C gpa-1 cDNA 2 ɒ ǘ [ l  H ç C  gpa-1 cDNA H trx-1 x    e  ,trx-1p::daf-28::venus (pOTA10) 2 venus ˇ ° ľ 2 ˜ / Ɨ Ï  $  trx-1p::gpa-3
cDNA::venus (pUDA15) , trx-1p::goa-1 cDNA::venus (pUDA13) ?Ďǒ/·ʇ$·ʇ$x`l3ʄ 2 /ʖʱ$

ˇ°ľœÏ
ˇ°ľœÏ3E;+/ĬĒE*$ƪǧH?,/ʂ'$(Mello et al., 1991)ˇ°ľ
œÏ2˦co-injection mix /3ņ˿/¸ȗD DNA H"E#E 5–50 ng/µl 2Ȃũ+Ï
E$M_OW^~S/3 pKDK66 ges-1p::nls-gfp H 50 ng/µl, pNAS88
ges-1p::nls-tag rfp H 50 ng/µl ;$3 pRF4 rol-6gf H 30 ng/µl 2Ȃũ+ÏE$ ·ʇ$Ɍɗ3ʄ 4 /ʖʱ$ƙčG!
˓ȕǈ/ Ex[trx-1p::yc3.60],~Sˇ°ľ,* Ex[ges-1p::taqRFP]H~MWM

-47-

_OW^/A'*œÏ$ˇ°ľœÏ+$?2/ɰ/ RFP 2ʀÎ, ASJ ƈʐn/ cameleon 2ˮɸ2ʀÎʑōE$ŸBE$Àµ(F1)H 1 NGM ȹ15°C ˼ɩ$ɳňąɊ/ACǗªHȕ;!F2 /?ʀÎȲʚ+$,F+EHS^NM_X/¸ȗ$Ǘ/2ɌɗHȗ*¡ˍ/AC tax-4 ĴȞµ, lite-1 ĴȞµ/?ˇ°ľœÏE$ɌɗH·ʇ$;ˇ°ľœÏE*.

˓ȕǈ2˧,ʖ+·ʇ$ˇ°ľœÏE$˓ȕ

ǈ2˩˧ĎµH¡ˍ!$"2$>/NGM ż2Ȏ
˧ 5 ó,˩˧Ďµ 3 óHɡ$F1 ª/˷ĩ/˾HĮš

*RFP 2

,/ACǆɸµĵˇ°ľHƓ(ÀµŸBE$2˧, tax-4 ĴȞµ2˩˧ĎµHB/¡ˍ$F1 ª/*¥ũ3 RFP 

lite-1(xu7); Ex[trx-1p::yc3.60, ges-1p::taqRFP]?Ďǒ/·ʇ$
KHR80 goa-1(n1134); gpa-3(pk35) ,KHR81 gpa-3(pk35) gpa-1(pk15) 

KHR82

goa-1(n1134); gpa-3(pk35) gpa-1(pk15)KHR88 pde-3(nj59); gpa-1(pk15); Ex[trx-1p::yc3.60,
ges-1p::taqRFP,

rol-6gf]  KHR89

pde-5(nj49);

gpa-1(pk15);

Ex[trx-1p::yc3.60,

ges-1p::taqRFP, rol-6gf]3ˇ°ŀȤ.ƙčG!ǧ,PCR ,ˬǟǪðHȗ
ÜǧHȗ

$ĴȞǎ

*·ʇ$·ʇ$Ɍɗ3ʄ 4 /ʖʱ$

in vivo S^NM_X
E;+/ʂGE*$ƪǧ/ŷ.'$(Kuhara et al., 2011; Ohnishi et al., 2011)

24×24 SrT`2/2%(v/w)KT`shlH·ʇ$"2/S^N
M_Xȗ2Ɍɗ Ex[trx-1p::yc3.60, ges-1p::taqRFP]H1~3 ƧóɡõȡȗƜȫé(õȡȗKKvJ A Ó )+ĢŅDM9 buffer HħB$/@-48-

SrT`HƙD~nUK+őH*ɼȣHˠM_XD2Ƣ
·H 3 Ý«Õ/ʂ\xH Olympus IX81 microscope(Olympus Corporation, Tokyo,
Japan)/ʅÇE*

DzgOeMx2ǷũäŹʅɡ(Tokai Hit Co. Ltd., Fujinomiya,

Japan) 2/ɡ2 Ý9-ƤɡDual-View (Molecular Devices, USA) optical system.
Hȗ*ʀÎHʑō$ʀÎȜÊ3 EM-CCD S EVOLVE512 (Photometrics, USA)+  50-100 ms 2 ˭ Î Ƴ ˜ + 1 ȸ  , / ơ Ų  $  K T   ` s h l 2 Ƿ ũ 3
MATS-5500RA-KY (Tokai Hit)2Ƿũ^`i/A'*ʖ˖$"E#E2ȜÊ2ʀÎ
ůũ3 MetaMorph (Molecular Devices) image analysis software system /ACʒǃ$ɑɪÕ2S^NȂũ2ȩŏĴò3 cameleon 2 acceptor/donor (YFP/CFP)ʀÎ2ǝ+ʔɆ$Ð*2 band-pass filters 3 cameleon /¸ȗ$ǷũäŹʅɡHȗ*E;+/ĬĒE*

D?2H*1723172ǷũĴòåȀHɍ 5 Ý˜

ASJ ƈʐn2ɑɪµÕ2S^NȂũ2ĴòHǸŅ$ÀµHĞźDƳ
3SrT`H`Ml!

ËƓ&$ɜɿ/ŕ˔2 M9 Hƙ*Ğă$
˘Ƴ˜2S^NM_XHʂĭč310%(v/w)KT`shlH¸ȗɿ 2 Ģ Ņ / 3 Ȩ Ŵ 0.1 µm 2.5%(w/v) polystyrene microsphere H ȗ$  2.5%(w/v)

polystyrene microsphere 2þǲ;$3Ţː$ǻǲHKT`shl2/ 0.5 µl
³

(Ƨȃɡ

$ɿHČȃ/ 1~Ƨóɡ@SrT`HƙDp

RM+őH*ɼȣHˠM_X$ǷũåȀH* 17B 23/ư*DƳ˜H -1 sec.,23/ǷũŃŅ$,FH 0 sec.,$"2;;ǷũäŹʅɡH 232;; 5 Ƴ˜ɡ$"2˜30 Ýŵ1 Ƴ˜ŵHʔǸ

"2ŵB3 1 Ƴ˜/ 5 Ƴ˜ŵ;+ʔǸ$Ƹŵ/17/ƌ$,2ɑɪÕ

-49-

S^NȂũ?ʔǸ$ʔǸɒŵɜɿ/ŕ˔2 M9 buffer Hƙ*ȕĿ*DȲʚ$(Ujisawa et al., 2014a)

ɸɎ]/ADɑɪȱĲɸɎ]Hȗ$ɑɪȱĲņ˿3E;+/ʂGE*$ƪǧ/ŷʂ'$(Kuhara et al., 2008; Mori and Ohshima, 1995 )˓ȕǈ/ Ex[trx-1p::GFP, ges-1p::GFP]ˇ
°ľœÏE$ɌɗHȗʖ2Ɍɗ2 L3$5%(v/w)KT`shl2/25M NaN3 H 4 µl ɡ

L4 ŦɿH 2 óɡ$SrT`H*]ȱĲʅɡ

Micropoint system laser microbeam(Photonic Instruments, USA)+ ASJ ƈʐn2ǉ
HȌ'*]HȆŒ$2,ïʪT`/3ɀɎT`laser dye /3W~
(Methanol < 1% BPBD 365 (6 mM))Hȗ
$ƈʐn3ŞĊ/ŏ2ɑɪµD$>ƪ2ɑɪµ2ǉHȱĲ$ASJ ƈʐn2 GFP 2ʀÎǱ

$2HȲʚSrT`HĄCM9 buffer H*ÀµHĞăNGM /ȹ$20°C  1 ƭ˼ɩASJ ƈʐn/ʀÎ.

,HȲʚ$,F+EH´ǷɦƁi`k/ȗ$2,[k,* ASJ ƈʐn/ȨƜ]HȆŒ3.

ĎƢ·Hą$(Mock)Àµ?ĎƳ/´ǷɦƁi`k$[vQS˹Ż˗Hȗ$ʑō`MlX`2/ 2%(w/v)2KT`ZH·ʇ100mM NaN3 H 10 µl ǽ$"2/ 2-3 ó2ƋɿHɡSrX`HD2xskH[vQS˹Ż˗(FV1000-IX81 with GaAsP photomultiplier tubes, Olympus)+ʑō-50-

FV10-ASW software(Olympus)+ʒǃ$mRNA 2Ğă
daf-2(e1370)3 25+ dauer /.D$>L1-L3 ŦɿƹH 25+˼ɩ+.

˓ȕǈ, daf-2(e1370)ĴȞµH 15+ 5 ƭ˼ɩ* L3 ŦɿƹHˁ$˴/20;$3 25/ȹ* 1 ƭ"E#E˼ɩ$mRNA 


ȗĞă$Č˼ɩǷũ/$2 NGM M9 buffer Hȗ*ɜɿH*ĎƳƹ/§ʵI% 1-2 Ǆ2˼ɩxk3˄Þ.ʄ

ȑĨHȳÀµ2 mRNA ,E$ȲʚD$>´ǷɦƁi`kH$mRNA 2
ɜɿH˼ɩ$ NGM xkB M9 buffer +ĞăM9 buffer H 2 ĞÏEƷ*ķ
ɰɺ@Ŧɿ.-HǬǮ$ɊʇƎ˵3RNeasy kit (QIAGEN)2Ǝ˵ƶ/ŷ'$

ǬǯE$ÀµH 650 l 2 RLT buffer +ƊȁǲµɀɎ+ÙɕCǾ$ǻǲB RNA HĞă$Ņ˔Ȥ PCR˓ȕǈ, daf-2(e1370)ĴȞµ2 mRNA Hʖ2Ǝ˵/ŷĞăɊʇHʂ'$Ɋʇ

$ mRNA H 100 ng /ʞʇiScript Advanced cDNA Synthesis kit for RT-PCR (Bio–Rad)Hȗ

*ʹʮÖ/AC cDNA HčƋ$Ņ˔Ȥ PCR 3 cDNA 

Sso Fast EvaGreen Supermix (Bio–Rad) "* DNA Engine Peltier
Thermal Cycler–CFX96 (Bio–Rad)Hȗ
ʂ*ʂ'$Č Sample /(*kxYMk+

ŷǁ2 protocol / step5 , 6 Hí*KnXƳ˜H 5 sec.B 8 sec./$

step1. 95 30 sec. > step2. 95 5 sec. > step3. KnXǷũ 8 sec. > step4. 2 7-51-

ƌD(×39 Ğ) > step5. 95 10 sec. > step6. 65 3 min > step7. Melt Curve 65 to 95ˇ°ľ2 fold change  lmn-1(n);$3 tba-1(α-gw)2 fold change ÄHīǺ/ǙʎòHʂ'$¸ȗ$x
M~3ʄ 3 /ʖʱ$ɗʔʒǃ
Ð*2ġ/ʄʖE*DǓǺʜŠ3 standard error of the mean (SEM)Hʄ*

Ð*2ġ2ɗʔʒǃ3ÌˍɡÝƦÝǃ(ANOVA)HʂDDunnett’s post hoc tests for

multiple comparisons /ACʒǃ$ġ2(*),(**)3"E#E p < 0.05 , p < 0.01 H
ƇēD

-52-





ƽȰȾHʂ/Ű$CȈż/Ɣœ*$%;$ȚùķŀȓŝŀˌȕµʞɇŀȰȾŇ þɉ ØƥƘ,ĹȘɹ ûĳ/żACýȴț;ˇ°ŀȤʒǃA6ȕȓŀȤʒǃ/øì˳;$Țùķŀ ȓŝŀˌ ȕȉŀȷ ȕµʞɇŀȰȾŇ2÷Ǐȕ ȯëāûIǊþȨ£I˘Ř5F<IǠʢ¤ɢI
ĻȽIȰȾė2ÑƺɢˑI/3żACƈʟț;"*Ķ2
C. elegans ĴȞµHƝº*˳;$ Caenorhabditis Genetic Center (CGC)National
Bioresource Project (Japan) ʢƱŤ ÍȕĴȞµ/í*x`lHƝº*˳;$ďćŘķŀ ǋˋƃ Íȕǂ¢ķŀ ˻˓˧ ÍȕUniversity of Michigan Dr. X.
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þ ɉ, łǤǿ ȭª, ĹȘ ɹ (2015) "ɜɿ Caenorhabditis elegans 2Ƿũ˄ſHäŹDȵɔ,ɰH¦$ƅĬÛȓ" ǝʰȕȓȕòŀ, Vol.32 No.2, p67-75,-60-

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73.3

Mock

1

20

21

4.76

013.čASJVÁŚŬŵųŴĭ¶¢Ųŵŋŵ6īĶĹ x­TįĭMß(A) ASJVÁŚŬŵųŴĭGFP»[sÎ "=, Vol.32 No.2, p67-75, 2015Ķĸ\7L]ďScale bar
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-73-

B

ASJ

D

A P
V

xJ{a"

0 (%)""YFP/CFP (%)"

xJ{ "ASJ"

15"15℃飼育"

25"

20℃飼育"

25℃飼育"

20"
15"
10"
5"
0"

温度"

tax-4!

野生株"

A

23"℃
17"℃
0"

50" 100" 150" 200" 250" 300"
aÚ (sec)"0íðêčóûøVÁŚŬŵųŴĭxJSTçóèæ×hĨ7ī ţųŪŵőŵħĈāĂĂąćæĀþăāĂāąĄç   èļĚğĢļŃűŌĿŨľũŵōŴŇīěĢďíñĊħá¯ěĢ×hĨ 7īxJ{ļēĢÝĭĎ»äŁśűŅŵ!ç÷úöüèĭŌŇřűďĮóûøVÁŚŬŵųŴĭ¤°īxJ{īAĝĹ&STļ Ńůŵ
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-74-

野生株"
lite-1!
tax-4; [trx-1p::tax-4]!

AB

tax-4!

10"
YFP/CFP (%)"

YFP/CFP (%)"

15"
10"
5"

温度"

0"

**

**

5"
0"

23"℃
17"℃
0"

50" 100" 150" 200" 250" 300"
aÚ (sec)"DCYFP/CFP (%)"

10"
5"

10"
5"
0"

0"

温度"

YFP/CFP (%)"

tax-4!
野生株"
lite-1!
tax-4;Ex[trx-1p::tax-4]!

17"℃
0"

50" 100" 150" 200" 250" 300"
aÚ (sec)"015.č7ĭASJVÁŚŬŵųŴĭŃűŌĿŨľũŵōŴŇ(A) tax-47ĭASJVÁŚŬŵųŴĮĎxJ{īAěĦ&STļĚĪđďxJ{ŞőŵŴĮŇůšīÅÏďlite-17ĲĢĮĎtax-47ĭASJVÁŚŬŵųŴħtax-4cDNAļĚğĢtax-4;
Ex[trx-1p::tax-4] ¨ĮxJ{ī&SěĢďĝİĦ15Ċá¯ďĠĺġĺ nČ12ď(B) ŇůšAĭ120-140secĭ
20ÚĭYFP/CFPĭH2ďŝŵŇůšĭ¶ĮŇůšAĭ¶ĨASď(C) xJ{ļēĪđ5*ĭASJVÁ
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-75-

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cGMP<T!
čččœūśű!
tax-2!
tax-4!

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lite-1!

daf-11!
odr-1!

goa-1!
gpa-3!

5’GMP

PDE! pde-1!

Ca2+!

pde-2!
pde-5!

ASJVÁŚŬŵųŴ"<(%)

B

'@GőŴŞņÈalphaŊŢŭŚŔŗ (Gα)

80
60
40

**

**

**

**
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**

**
*
*

100

ŇĽŚűÕ
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**

**
**

**

20℃ 2℃
**

**

20
0

016.čxJU4ĮASJVÁŚŬŵųŴĭcGMP¦Ëļ ěĦ ÑĚĺĹ(A) ASJVÁŚŬŵųŴħo±ĝĹ3ØGőŴŞņÈĨcGMP<Tœūśűļ ěĢ'@U4 Ñĭ;¦Ë0ď(B) '@ĭU4 Ñħo±ĝĹÒ ;ĭ7ĭ x­Tď'@(lite-1)Ď3ØGőŴŞņÈαŊŢŭŚŔŗ(goa-1Ďgpa-1Ďgpa-3)ĎŇĽŚűÕŌņůŵŐ(daf-11Ďodr-1)ĎťŎťōŁŎŕůŵŐ(pde-1Ď
pde-2Ďpde-3Ďpde-5)ďÖ7ĵÖ7ĭĽŰű,Į#7ĭĴĭĨ+ĜďĠĺġĺ n≥9ď*P <0.05; **P < 0.01ďGα: GőŴŞņÈαŊŢŭŚŔŗĎGC: ŇĽŚűÕŌņůŵŐĎPDE: ťŎťōŁŎŕůŵŐď

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A P

ASJ
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Í£
trx-1p::gpa-1::venus

trx-1p::dsRedm

Merge

trx-1p::gpa-3::venus

trx-1p::goa-1::venus

017.čASJVÁŚŬŵųŴĭGőŴŞņÈalphaŊŢŭŚŔŗĭD1
×hīASJVÁŚŬŵųŴīDsRedmĨĠĺġĺĭGα(GPA-1ĎGPA-3ĎGOA-1)::VenusļĚğĢďASJVÁŚŬŵųŴĭmK0ļīĝďEĭĮASJVÁŚŬŵųŴīĔĘĹGα::VenusĭD1ļěĦđĹď:ĭĮDsRedmĭASJħĭļěĦđĹď(ĭĮEĨ:ĭļ`À×ħÖĬ
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80

**

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**

**

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20℃

2℃(24h)

60
40
20
0

goa-1(n1134); gpa-3(pk35) gpa-1(pk15); Ex!

018.čGőŴŞņÈalphaŊŢŭŚŔŗĭÒ ;ŲŎńŬŵ?â
GαőŴŞņÈĭÒ ;ĭÖ7Į20Ċħá¯ĚĺĢNī2Ċħ24aaÚĭ x{ļēĦĴ¡60%Ė<ħėĹďęĭÖ7īĔđĦASJVÁŚŬŵųŴīGαÒ ;ļĚğĹĨÓĪ½3ĭ.RĖ¿ķĺĢďgoa-1pīĮ goa-1’s own promoter(3kb), gpa-1pīĮ gpa-1’s own promoter(1.5kb),
gpa-3pīĮgpa-3’s own promoter(3kb)ļđĢď, gcy-5pĮASER -ÁŚŬŵųŴţųŪŵőŵĎæ
ceh-36p ĮxJÉTĭ¦.ËīÛĻĹAWCVÁŚŬŵųŴţųŪŵőŵĎætrx-1p ĮASJ VÁŚŬŵ
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-78-

(17℃)" xJ{a(23℃)"
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10"

10"

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Minimum0"-5"0 (%)"xJ{ (23℃)"xJ{a(17℃)" "
ASJ"

23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"0"-10 (%)""C
5"
YFP/CFP (%)"

D
A P
V

YFP/CFP (%)"

Warming"xJ{Cooling"

B

ASJ

温度"

A

×h

0"
-5"

Min – MaxMaximum

温度"

-10"

Minimum

23℃"

17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"019.čASJVÁŚŬŵųŴĭWarming{ĨCooling{īAĝĹST(A) ×hĭASJVÁŚŬŵųŴīWarming{()ĲĢĮCooling{()ļēĢaĭ¤°ĭl;ď
Warming{ļēĹĨĎASJVÁŚŬŵųŴĭŃűŌĿŨ|JĖ_ĝĹďCooling{ļēĹĨĎŃű
ŌĿŨ|JĖ ĝĹď(B) 20Ċħá¯ěĢ×hĭ17-23-17ĊxJ{(Warming)īAĝĹ&STďbĴ
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-79-

D

goa-1

10"

YFP/CFP (%)"

YFP/CFP (%)"

A
5"
0"
-5"

E

gpa-1

10"
5"
0"
-5"

0"

goa-1; gpa-3 gpa-1

10"
5"
0"
-5"

温度"

YFP/CFP (%)"

C

5"

-5"

YFP/CFP (%)"

YFP/CFP (%)"

B

gpa-3 gpa-1

10"

gpa-3

10"

23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

5"
0"

温度"

-5"
23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

F

G

5"
0"

*

**

15"
Max-Min"
YFP/CFP (%)"

10"

5"
Minimum"
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Maximum"
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15"

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-5"

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10"5"0"020.čGαőŴŞņÈĭ7ĭWarming{īAĝĹST(A-C) GαőŴŞņÈgoa-1Ďgpa-1Ďgpa-37ĭxJSTď¸đÞ¶ĭ©Į×hĭASJVÁŚŬŵųŴ
ĭxJSTļĝď(D,E) GαőŴŞņÈĭ8Ö7ĭxJSTďĠĺġĺn=18-20ď(F)A-Eĭb97"(Maximum)a}ĕķ N5Ğĥĭ10ÚĭYFP/CFP7"ď(G) A-EĭMinimum(280-290 sec)a}ĭ10ÚĭYFP/CFP7"ď(H) A-EĭMaximum-MinimumĭYFP/CFP7"ĭFďŝŵŇůšĭ¶ĮĠĺġĺĭŇůšA-Eĭ¶īASď*P < 0.05; **P < 0.01ď

-80-

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goa-1

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YFP/CFP (%)"

YFP/CFP (%)"

A
0"
-5"

YFP/CFP (%)"

YFP/CFP (%)"

E

gpa-1

5"
0"
-5"
-10"

-5"

goa-1; gpa-3 gpa-1

5"
0"
-5"
-10"

gpa-35"

温度"

YFP/CFP (%)"

C

0"

-10"

-10"

B

gpa-3 gpa-1

5"

0"

23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

-5"

温度"

-10"
23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

G

-5"

-10"
**

** **

Min-Max"
YFP/CFP (%)"

0"

H

"
Minimum"
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Maximum"
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F

0"

-5"

-10"

10"

5"
*

*

0"
**

**

**

021.čGαőŴŞņÈĭ7ĭCooling{īAĝĹST(A-C) GαőŴŞņÈgoa-1Ďgpa-1Ďgpa-37ĭxJSTď¸đÞ¶ĭ©Į×hĭASJVÁŚŬŵųŴ
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-81-

YFP/CFP (%)"

10"

Maximum"
YFP/CFP (%)"

DA
daf-11

5"
0"

10"
*
5"
0"

-5"
odr-1

E

5"

Minimum"
YFP/CFP (%)"

YFP/CFP (%)"

10"

0"
-5"

**

**
-5"
**

C

daf-11; odr-1

5"

F

0"

Max-Min"
YFP/CFP (%)"

YFP/CFP (%)"

10"

温度"

5"
0"

B

-5"

23℃"
17℃"

**
10"
5"
0"

30"50" 100" 150" 200" 250" 300"
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šA-Cĭ¶īASď*P < 0.05; **P < 0.01ď

-82-

Maximum"YFP/CFP (%)"

YFP/CFP (%)"

5"D

daf-11

0"
-5"

-10"

0"

A

-5"

-10"
odr-1

E

0"Minimum"
YFP/CFP (%)"

YFP/CFP (%)"

5"

-5"
-10"

5"

daf-11; odr-1

-10"

0"

F

-5"
-10"

温度"

*

-5"

Min-Max"
YFP/CFP (%)"

YFP/CFP (%)"

C

0"

B

23℃"
17℃"

10"
*

*

5"

0"

30"50" 100" 150" 200" 250" 300"
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šA-Cĭ¶īASď*P < 0.05; **P < 0.01ď

-83-

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0"

0"

YFP/CFP (%)"

YFP/CFP (%)"

5"

温度"
10"

pde-3

5"

pde-1 pde-5; pde-3; pde-2

10"
5"
0"

-5"

pde-5

23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

G

0"

10"

0"

-5"

-5"

D

H

15"
*
10"
**

*

**
**

5"

5"
* *
0"

-5"

**
**

**
**

0"

5"
0"

I

-5"

温度"

YFP/CFP (%)"

F

pde-2Maximum"
YFP/CFP (%)"

YFP/CFP (%)"

B

C

5"

-5"

-5"

10"

pde-1 pde-5; pde-2

10"

Minimum"
YFP/CFP (%)"

10"

E

pde-1YFP/CFP (%)"

YFP/CFP (%)"

A

23℃"
17℃"

Max-Min"
YFP/CFP (%)"

15"

30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

**
**

10"

*

5"
0"

024.čťŎťōŁŎŕůŵŐĭ7ĭWarming{īAĝĹST(A-D) ťŎťōŁŎŕůŵŐpde-1Ďpde-2Ďpde-3Ďpde-57ĭxJSTď¸đÞ¶ĭ©Į×hĭASJ
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n=15-19ď(G) A-Fĭb97"(Maximum)a}ĕķ N5Ğĥĭ10ÚĭYFP/CFP7"ď (H) A-Fĭ
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-84-

YFP/CFP (%)"

5"E

pde-1YFP/CFP (%)"

A

0"
-5"

0"
-5"

温度"

30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

0"
-5"

0"
-5"

-10"
23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

-5"

**
0"
**
-5"

**

-10"
*

-15"

I
Min-Max"
YFP/CFP (%)"

5"

pde-5

H
Minimum"
YFP/CFP (%)"

GMaximum"
YFP/CFP (%)"

DYFP/CFP (%)"

-5"

23℃"
17℃"

pde-3

-10"

温度"

0"

0"

YFP/CFP (%)"

5"pde-1 pde-5; pde-3; pde-2

5"

-10"

-10"

C

-5"

F

pde-2YFP/CFP (%)"

YFP/CFP (%)"

5"

0"

-10"

-10"

B

pde-1 pde-5; pde-2

5"

10"
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5"
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-85-

YFP/CFP (%)"

A

×h

gpa-1

pde-5

pde-5; gpa-1

10"
5"
0"

温度"

-5"
23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

Maximum"
YFP/CFP (%)"

NS

10"
5"
0"

D

C
5"

**

NS

0"

Max-Min"
YFP/CFP (%)"**

Minimum"
YFP/CFP (%)"

B

10"
5"
0"

-5"

026.čGαĨPDE7īĶĹWarming{īAĝĹ ÛĭjÆ(A) GαőŴŞņÈgpa-1ĨťŎťōŁŎŕůŵŐpde-5ĨĭÖ7ĭxJSTĕķĎGαĭ7ĨPDEĭ
7ĭ ÛļÃfď(B) Aĭb97"(Maximum)a}ĕķ N5Ğĥĭ10ÚĭYFP/CFP7"ď(C) AĭMinimum(280-290 sec)a}ĭ10ÚĭYFP/CFP7"ď(D) AĭMaximum-MinimumĭYFP/CFP7"ĭFďpde-5; gpa-1ĭxJSTĮĎpde-5ī<ěĦđĹďŝŵŇůšĭ¶ĮĠĺġĺĭŇůšAĭ¶īASď
n=9-14ď*P < 0.05; **P < 0.01ď

-86-

A

×h
pde-3

gpa-1
pde-3; gpa-1

YFP/CFP (%)"

5"
0"
-5"

温度"

-10"
23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

-5"

-10"

NS

D
Min-Max"
YFP/CFP (%)"

0"

Minimum"
YFP/CFP (%)"

Maximum"
YFP/CFP (%)"

C
0"
-5"
-10"

10"
5"
0"

NS

**

B

**
027.čGαĨPDE7īĶĹCooling{īAĝĹ ÛĭjÆ(A) GαőŴŞņÈgpa-1ĨťŎťōŁŎŕůŵŐpde-3ĨĭÖ7ĭxJSTĕķĎGαĭ7ĨPDEĭ
7ĭ ÛļÃfď(B) Aĭb97"(Maximum)a}ĕķ N5Ğĥĭ10ÚĭYFP/CFP7"ď(C) AĭMinimum(280-290 sec)a}ĭ10ÚĭYFP/CFP7"ď(D) AĭMaximum-MinimumĭYFP/CFP7"ĭFďpde-3; gpa-1ĭxJSTĮĎpde-3ī<ěĦđĹďŝŵŇůšĭ¶ĮĠĺġĺĭŇůšAĭ¶īASď
n=11-13ď*P < 0.05; **P < 0.01ď

-87-

Warming 17-23-17ĊťŎťōŁŎŕůŵŐ (PDE)ŇĽŚűÕGőŴŞņÈ
ŌņůŵŐ (GC) alphaŊŢŭŚŔŗ (Gα)

Ò

Max

;

goa-1
*

gpa-1
gpa-3

Max-Min

Min

**

**

**

**

**

**

odr-1

****

pde-1

****

pde-2

****

pde-3

**

pde-5

*

温度"

23ĊV

Max

**

**

*

daf-11

Cooling 23-17-23Ċ

**

Min-Max

*

Min

**

*

*
**

**
**

*

2qxJ7" >GWįĭYĸ

17ĊV 2qxJ7" >GW
įĭYĸ

23℃"

23℃"

17℃"

17℃"
30" 50" 100" 150" 200" 250" 300"
aÚ (sec)"50" 100" 150" 200" 250" 300"
aÚ (sec)"

028.čASJVÁŚŬŵųŴĭWarming{ĨCooling{īAĝĹSTĭĲĨĳ
020-25ĭ7ĭxJSTīAĝĹGĭĲĨĳďMaximum(Max)Į1qxJ7"īAĝĹ&STļěĎMinimum(Min)Į>GWįĭYĸ*ļĝďMax-MinĲĢĮMin-MaxĮ2qxJ7"īAĝĹ&STļěĦđĹď*P < 0.05; **P < 0.01ď

-88-

A

B

snb-1!

snb-1!
15"

YFP/CFP (%)"

YFP/CFP (%)"

15"
10"
5"

10"
5"
0"

0"

温度"

23"℃

17"℃
0"

50" 100" 150" 200" 250" 300"

17"℃
0"

40"

100" 140"aÚ (sec)"aÚ (sec)"< (%)

C
100

**

**
** **

190 "15℃ 2℃
20℃ 2℃

80
60
40
20
0

029.čASJVÁŚŬŵųŴĮ#ħZxJļ'@ĝĹ(A) ŌřţŎ ÑīÛĻĹ7ĨěĦĎsnb-1/ŌřţŗŢŲŠŴĭ7īxJ{ļēĢÝĭASJVÁ
ŚŬŵųŴĭ&STļÃfďsnb-17ĮxJSTļěĢď(B) xJ{ļēĪđ5*ĭxJSTďaÚ¦ÐīAĝĹxJ7"ļŇůšīÅÏď(C)¦ ÑīÛĻĹ;ħĐĹ¦ŤţœŘ*XÔ¢(egl-21)ĎŌřţŗőŇŧŴ(snt-1)ĎŌřţŗŢŲŠŴ(snb-1)ĎŌŴőńŌŴ(unc-64)ĭ7ĭ x­TďĠĺġĺ n≥6ď**P < 0.01ď

-89-

A

インスリン
DAF-28
INS-6
DAF-2"
AGE-1" PDK-1"
インスリン PI3-"受容体 キナーゼDAF-16"
FOXO"

AKT-1,2"
AKT/PKB
キナーゼ
DAF-16"Ò

B

120 60

60

;

60

60

60

~

YFP/CFP (%)"

17Ċ23Ċ

60 照射時間"(sec)"
23Ċ

30"

野生株"

20"
10"
0"
-10"

温度"

-20"
23"℃

17"℃
-1" 0

xJ{

30"

60"

120"

180"

"

240"

300" ÄyaÚ"(min)"

030.čASJVÁŚŬŵųŴĭvįĭU4 Ñ(A) ASJVÁŚŬŵųŴĕķĮľŴŎŰŴĖtĚĺĎ³ĵ¦ ĭľŴŎŰŴ'@DAF-2ħ'@ĚĺĹď(B)×hĭASJVÁŚŬŵųŴĭÙaÚŃűŌĿŨľũŵōŴŇďxJ{ļēĦĕķ30NĎ¤°ŃűŌĿŨ|JĖb9ĨĪĤĢďĠĭNĎŃűŌĿŨ|JĮwCěĎ>GWīYĤĢďYFP/CFP7"Į-1Ď0Ď30Ď
60Ď120Ď180Ď240Ď300 minĭa}ħ60ÚĠĺġĺy>ěĢďaÚ¦ÐīAĝĹxJ7"ļŇůšīÅ
Ïďn=13ď

-90-

A

B

10"

YFP/CFP (%)"

YFP/CFP (%)"

eri-1; lin-15B!

5"
0"

23℃"
17℃"

10"
5"
0"
-5"

温度"

温度"

-5"

pros-1 knock down!

23℃"
17℃"
30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

30"50" 100" 150" 200" 250" 300"
aÚ (sec)"

031.čpros-1ŜŔņŒĿŴ ¨ĭxJ{īAĝĹASJVÁŚŬŵųŴĭST
pros-1p[hĮµrTļĝĢĳĎeli-1; lin-15B ¨īAěĦpros-1(ceh-26)Ò ;ļRNAiuļđĦŜŔņ
ŒĿŴěĎŇŰĽ¤°(AMsh)ĭASJVÁŚŬŵųŴĭxJSTīAĝĹMßļŃűŌĿŨľũŵōŴŇħÃ
fěĢď(A) eli-1; lin-15B ¨Į×hĨ+lĭxJSTļěĢďn=15ď(B) RNAiuļđĦeli-1; lin-15B¨īAěĦpros-1Ò ;ļŜŔņŒĿŴěĢ ¨ĭxJSTďn=19ďaÚ¦ÐīAĝĹxJ7"ļŇůšīÅÏď

-91-

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< (%)

60

20℃ 2℃
25℃ 2℃

40

**

**

**

**
**

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20
0

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-92-;ĭ7ĭ

< (%)

100

15℃ 2℃

80

20℃ 2℃

60
40

25℃ 2℃

20
0

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ĭ7ĭ15ĊĎ20ĊĎ25Ċá¯Nĭ x­TďĠĺġĺn≥6ďæ*P < 0.05; **P < 0.01ď

-93-

A

Ò

;

cgt-1
cpr-1
dhs-4!
dmd-7
gpa-7
gpdh-1!

ŏůŧŘŇűŉŌűŗůŴŎšŀůŵŐŌŎŕľŴţųŕĽŵŐşŘųńŌŎŕųľŘ 17-β ŖşŘųňřŵŐdoublesex3Ì/;
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M60.2

B
Fold change"(×hĨĭsÎ)


30"
20"
10"

őŴŞņÈ

**** **

**
**
**

5"

**
**

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2
1"
0.5"

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C

6"
5"
4"
3"
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20℃á¯
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20"
15"
10"
5
0"
fat-5

fat-6

fat-7!

035.č²ÈkXīÛĻĹÒ ;ĭdaf-27īĔĘĹØĭÃfC. elegansĭ²È´kXīÛĻĹÒ ;ħĐĹΔ9 ŖŊœŬůŵŐ (fat-5Ďfat-6Ďfat-7)ĭdaf-27īĔĘĹ>ØPCRĭ§gďĠĺġĺĭŝŵŇůšĮ)xJħá¯ĚĺĢ×hĨĭAļĝďn=3ď

-95-

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インスリン
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cGMP<Tčččččœūśű

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?"GPA-3
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INS-1
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TAX-2/-4

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PDE-5
ťŎťō"ŁŎŕůŵŐ"インスリン
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ASH, ASI, PHA, PHB

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cGMP gated channel alpha subunit

AFD, ASE, ASG, ASI, ASJ, ASK, AWB, AWC, BAG, URX

cGMP gated channel beta subunit

AFD, ASE, ASG, ASI, ASJ, ASK, AWB, AWC, BAG, URX

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1) Ohta*, A., Ujisawa, T*., Sonoda, S. & Kuhara, A. (*Equally contributed)
Light and pheromone-sensing neurons regulates cold habituation through insulin
signalling in Caenorhabditis elegans.
Nature communications, 5 (4412), 1-12, 20142) Ujisawa, T., Ohta, A., Uda-Yagi, M., Kuhara, A.
Diverse Regulation of Temperature Sensation by Trimeric G-Protein Signaling in

Caenorhabditis elegans.
PLOS ONE, 11(10), e0165518, 1-20, 2016

1) Sonoda S., Ohta, A., Maruo, A., Ujisawa, T., Kuhara, A.
Sperm affects head sensory neuron in temperature tolerance of Caenorhabditis elegans.
Cell Reports, 16, 1, 56–65, 20162) Kage-Nakadai, E., Ohta, A., Ujisawa, T., Sun, S., Nishikawa, Y., Kuhara, A., Mitani, S.
A Caenorhabditis elegans homolog of Prox1/Prospero is expressed in the glia and isrequired for sensory behavior and cold tolerance.
Genes to Cells, 21, 9, 936-948, 20163) Ujisawa, T., Ohta, A., Okahata, M., Sonoda, S. & Kuhara, A.
Cold tolerance assay for studying cultivation-temperature-dependent cold habituation in C.

elegans.
Protocol Exchange, doi:10.1038/protex.2014.032, 1-7, 20144) Ujisawa, T., Ohta, A., & Kuhara, A.
Long-term calcium imaging of ASJ sensory neuron controlling cold tolerance inCaenorhabditis elegans.
Protocol Exchange, doi:10.1038/protex.2014.034, 1-6, 20145), 

, " [$]# Caenorhabditis elegans %

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