1. Apical Dominance is controlled by Interaction between Cytokinin biosynthesis/degradation and Auxin in Stem.
Mori, H.
Nagoya University

5. 3days
after decapitation
intact

6. Apical Dominance auxin cytokinin axillary bud dormant outgrowth
decapitation

9. Northern Blot Analyses of Cell Cycle Related Genes
cdc2
CycD3 ;1
CycB1 ;2
histoneH4
PCNA 2 4 8 6 10 12 14 16 18 20 22 24
Time after decapitation (hours)
EtBr

10. 1cm
2cm
4cm
6cm

11. Expresson of PCNA mRNA

12. auxin cytokinin axillary bud outgrowth dormant decapitation
・Auxin derived from terminal bud is transported basipetally.
・When auxin applied to the cut stump after decapitation,
Auxin can repress axillary bud outgrowth.
・When auxin directly applied to axillary bud after decapitation,
Auxin cannot repress axillary bud outgrowth.

13. Auxin is basipetally transported in stem.
axillary bud
IAA
Scale leaf
at 2nd node

14. The basipetal transport of auxin is essential
for dormancy of axillary bud.
Auxin acts at node or in stem, not in axillary bud.

15. The basipetal transport of IAA is essential for axillary buds dormancy
intact
decapitated
IAA
3 hr
1cm
IAA
0 hr
1 hr
auxin transport rate: 1cm/hr

16. Subtracted cDNA clones
0hr-3hr
Ps-IAA4/5, IAA7, Aux28, Aux22, GH3,
F-box, cullin, auxin efflux carrier protein(PIN1 ),
RD29B, LEA4, zeaxanthin epoxidase
3hr-0hr
IAA-Ala hydrolase, GA2 oxidase,
HMG-CoA synthase, squalene synthase,
Isopentenyltransferase (IPT )

17. Current model of cytokinin metabolism in higher plants
LOG
LOG
LOG
LOG
(revised)

18. 1cm
2nd node

19. Expression of PsIPTs at the 2nd Node after Decapitation
0.5 4 3 2 1 5 6 9 15 18 24 12
PsIPT1
PsIPT2
rRNA
time after decapitation (hr)

20. Is IPT expression repressed ?
excise the segments
3hr after decapitation
IPT is expressed
incubate in MES buffer
with or without IAA
Is IPT expression repressed ?
1cm
excise the segments
from intact seedlings
IPT is NOT expressed
incubate in MES buffer
with or without IAA
Is IPT expressed?

21. PsIPTs Expression was Repressed by IAA1 2 3 4 5
IAA Free
10-5 M IAA
PsIPT1
PsIPT2
rRNA
incubation time (hr)

22. Is IPT expression repressed?
excise the segments
after 3hr decapitation
IPT is expressed
incubate in MES buffer
with or without IAA
Is IPT expression repressed?
1cm
excise the segments
from intact seedlings
IPT is NOT expressed
incubate in MES buffer
with or without IAA
Is IPT expressed?

23. PsIPTs Expression was Induced in IAA Free Buffer
10-5 M IAA 1 6 5 4 3 2
incubation time (hr) 9 8 7
PsIPT1
PsIPT2
rRNA

24. 10-5 M IAA IAA Free 10-5 M IAA excise the segments
after 3hr decapitation
IPT is expressed
incubate in MES buffer
with or without IAA
Repress IPT expression?
1cm
IAA Free 1 2 3 4 5
10-5 M IAA 1 2 3 4 5
IAA Free
10-5 M IAA 1 6 5 4 3 2 9 8 7 1 6 5 4 3 2
excise the segments
from intact seedlings
Not IPT expression
incubate in MES buffer
with or without IAA
IPT express?

25. PsIPTs Expression was Repressed by IAA Transported Basipetally in the Stem
PsPIN1 ＋
IAA
Lanoline paste w or w/o IAA after decapitation
3hr
Is IPT expression repressed?
1cm

28. pPsIPT2::GUS in Arabidopsis

29. pPsIPT2::GUS (2,4-D treated for 1 week)

30. pPsIPT2::GUS (2,4-D free for 3 days)

31. in Transformants harboring pPsIPT2::GUS
GUS Transcripts Deceased by Auxin
in Transformants harboring pPsIPT2::GUS
IAA Free
10-5M IAA 1 2 3 4 5
incubation time (hr)

32. Cytokinin Levels in the Stems and Axillary Buds
after Decapitation

33. Cytokinin Levels in the Excised Segments treated with/without IAA
100
200
300
iPMP
iPA iP
t-ZMP
t-ZR
t-zeatin 0h 3h 6h
-IAA
+IAA
100
200
300 0h 3h 6h
(p mol/ gFW)
(p mol/ gFW)
iPMP
iPA iP
t-ZMP
t-ZR
t-zeatin
excised the segments
put on the agar plate
± 1%(w/v) IAA treatment

34. Conclusion
One role of auxin in apical dominance is to repress the PsIPTs expression; local cytokinin biosynthesis in the nodal stem is negatively regulated by auxin through the control of IPT expression.
Cytokinin, which is thought to be derived from the roots so far, is locally biosynthesized in the stem rather than in the roots after decapitation.

35. PsIPTs Expression was Repressed by Basipetally Transported IAA in the Stem
PsPIN1 ＋
IAA
1% (w/v) IAA
after decapitation
3hr
Is IPT expression repressed?
1cm

36. Effect of IAA Applied to the Stump on PsPIN1 Expression
actin 1 2 3 4
(cm) 1 2 4
(hours) 1 2 3 4
IAA

37. Can IAA from Axillary Buds induce PsPIN1?
Biosynthesis
decapitated
intact
IAA
IAA
3hr
9hr
0hr

38. × IAA IAA Lower Stem Upper Stem PsPIN1 actin2 4 6 8 10 12
PsPIN1
actin
Time after decapitation ( hours )

39. Expression of PsIPTs at the 2nd Node after Decapitation
0.5 4 3 2 1 5 6 9 15 18 24 12
PsIPT1
PsIPT2
rRNA
time after decapitation (hr)

40. upper stem node lower stem IAA 81.7 CK 2.9 44.0 9.1 19.3 135.5 20.8
105.5
60.5
3.6
35.4
175.2
175.3
47.6
79.5
1.9
50.5
6.3
27.7
76.6
33.6
158.0
34.6
94.8
39.9
50.7
55.2
23.8
72.2
7.8 1 3 12 18 24 36 48
upper
stem
node
lower
stem
(p mol/gFW)

41. Cytokinin oxidase cDNAs were isolated from the stem 12 hr after decapitation.
PsCKX1：
was constantly expressed in stem before and after
decapitation,
was not induced by auxin.
PsCKX2：
was increased in stem after decapitation
was induced by auxin.

42. Expression of PsCKX2 in Pea Stem10 20 30 40 80
120
decapitation
TIBA treatment
Relative amounts of mRNA
Time after treatment (hr)

43. PsCKX2 Expression in Excised Pea stem by Auxin10 20 30 40
(x 103)
Relative amounts of mRNA
+NAA
-NAA
Time after treatment (hr)

44. PsCKX2 Expression in Pea stem
- NAA 40 80
120
Relative amounts of mRNA 10 20
Time after treatment (hr)

45. PsCKX2 Expression in Excised Pea stem by Auxin10 20 30 40
(x 103)
Relative amounts of mRNA
+NAA
-NAA
Time after treatment (hr)

46. upper stem node lower stem IAA 81.7 CK 2.9 44.0 9.1 19.3 135.5 20.8
105.5
60.5
3.6
35.4
175.2
175.3
47.6
79.5
1.9
50.5
6.3
27.7
76.6
33.6
158.0
34.6
94.8
39.9
50.7
55.2
23.8
72.2
7.8 1 3 12 18 24 36 48
upper
stem
node
lower
stem
(p mol/gFW)

48. Expression of PsCKX2, PsPIN1, PsIPT2 in stem
120
PsCKX2 80
PsPIN1
Relative amounts of mRNA
PsIPT2 40 g. 10 20 30
Time after decapitation (hr)

49. Model of Interaction Between Auxin and Cytokinin in Apical Dominance

52. Effect of Striolactone GR24 on outgrowth of axillary bud after decapitation
control

53. Apical dominance Nagoya University ・Mina TANAKA ・Sae SATO-SHIMIZU
・Aiko ISHIWATA
・Yuka MADOKA
RIKEN
・Mikiko KOJIMA
・Hitoshi SAKAKIBARA
This work was done by my laboratory in Nagoya University and Dr.Sakakibara group in RIKEN.
Thank you for your attention.