2. Cytokinins Biosynthesis, Signaling Pathways, Functions and Cross talk with Auxins
ARSHAD MAHMOOD KHAN
PhD Scholar (13-arid-3581)
BOTANY DEPARTMENT
UAAR RAWALPINDI

4. DISCOVERY CYTOKININS
Haberlandt (1913) – compound in phloem stimulates cell division
Van Overbeek (1941) – coconut milk (endosperm) also has the ability to stimulate cell division
Jablonski & Skoog (1954) – compounds in vascular tissues promote cell division
Miller (1955) – first cytokinin isolated from herring (Fish) sperm, named kinetin

5. DISCOVERY CYTOKININS
Miller (1955) – first cytokinin isolated from herring sperm, named kinetin
Miller (1961) – first naturally occurring cytokinin found in plants (Zea mays L.), later called Zeatin
Lathum & Guillfoyle (1974)
Zeatin

6. CHEMICAL STRUCTURE Adenine structure (amino purine ring) N6 Side chain
CYTOKININS
Adenine structure (amino purine ring)
N6 Side chain
Degree of unsaturation
Number of carbons in side chain

7. CHEMICAL STRUCTURE Free base forms are most hormonally
CYTOKININS
Free base forms are most hormonally
active (Yamada et al. 2001)
May have sugars attached to make
molecule more inactive and useful for
transport
Ribosides
Ribotides
Glycosides

8. BIOSYNTHESIS CYTOKININS
Biosynthesis started by the IPT (isopentenyl tranferase) enzyme
Major pathway
Figure on slide no. 9
Other pathway
tRNA biosynthesis
Figure on slide no. 10

9. Plant physiology, 4th Edi. Fig. 21.6, 2006 Sinauer Associate. Inc
BIOSYNTHESIS

10. BIOSYNTHESIS tRNA breakdown not the major pathway for
cytokinin synthesis
Plant physiology, 4th Edi. Fig. 21.7, 2006 Sinauer Associate. Inc

11. BIOSYNTHESIS Generated mostly in the root apical meristems
CYTOKININS
Generated mostly in the root apical meristems
but also found in:
Root cap cells
Ovules
Phloem cells
Leaf axils
Tips of young inflorescences
Fruit
Seeds

12. TRANSPORT Cytokinins move up the plant through the xylem
By contrast, auxin moves from top down.
Some signal in the shoot can also induce cytokinin transport from the root (Beveridge 2000).

13. SIGNALING PATHWAY
\figures\ch21\pp21270.jpg

14. Cytokinins perception and pathway
Cytokinin Signaling Pathway in Arabidopsis consists of four steps:
Initiation of a Phosphorelay Cascade by distinct Plasma membrane His (Histidine) Protein Kinases upon Cytokinin perception
Convergence of the signals initiated at His Protein Kinases, on AHP proteins, that serve as Phospho-relay carriers between the Cytokinin receptors and the downstream nuclear responses
Activation of B-type ARR proteins by nuclear AHP
Translocation and transcriptional activation of A-type ARRs
Mechanism:
First of all Cytokinins binds with the CHASE (Cyclases/Histidine kinases–Associated Sensory Extracellular) domain of receptors e.g. (CRE1, AHK2, AHK3) present in PM
The binding of Cytokinin activates the Transmembrane Domain, which Autophosphorylates on a His domain.
The phosphate then transferred to Receiver Domain and an AHP protein, which translocates to the nucleus, where it activates ARRs.

15. Cytokinins perception and pathway
ARR form a large gene family composed of 22 genes that includes two major classes (Type-A and Type-B)
Type-A ARRs (ARR3,4, 5, 6, 7,8,9,15,16,17,18,19)
Type-B ARRs (ARR1, ARR2, and ARR10,11,12,13,14)
AHP activates Type-B ARRs increases the transcription of the
Type-A ARRs, which feed back to inhibit their own transcription.
This transcription and translation of specific ARRs produces
specific cytokinins responses in plants

17. Cytokinin changes the structure of the His Kinase domain
SIGNALING SUMMARY
CYTOKININS
Cytokinin changes the structure of the His Kinase domain
Phosphate moves from the kinase domain to the receiver domain
Phosphate moves onto AHP
AHP moves from cytoplasm into nucleus
Phosphate on AHP moves onto receiver domain of the response regulator
Change in structure of the receiver domain affects the output domain
Output domain then signals transcription to cytokinin responses

18. FUNCTIONS Cell division Root and shoot meristems Cell differentiation
CYTOKININS
Cell division
Root and shoot meristems
Cell differentiation
Leaf senescence
Inflorescence growth
Nutrient mobilization
Cotyledon expansion
Apical dominance
Seedling morphology

19. CELL DIVISION CYTOKININ FUNCTION
Auxin and cytokinins influence the activity of:
Cyclin-dependant protein kinases (CDKs) and cyclins
Both are proteins that regulate transitions between G1 to S and G2 to mitosis stages in the cell cycle
Auxin stimulates the production of CDKs and cyclins
Cytokinins activate CDKs and cyclins through phosphrorylation and allow transition between stages

20. 26S Proteosome cyclin synthesized amino acids cyclin degraded cyclin
triggers mitosis
or DNA synthesis
dephosphorylation
CDK
CDK
CDK P
(inactive)
(active)
phosphorylation
Figure 15.7: Diagram showing how the cdc2 protein kinase (C-PK) and cyclin trigger cell cycle progression in cycling cells. Starting in the top left, cyclin has joined to C-PK in a phosphorylated form to make a new, but inactive, protein. An enzyme removes the phosphate (P) to activate this protein. This form of the protein acts as the trigger to switch on S phase and M phase. It is recycled by separating C-PK, which is phosphorylated again and recycled. The cyclin is broken down into amino acid subunits and must be resynthesized. ATP, Adenosine triphosphate; GTP, guanosine triphosphate.
C-PK
ATP?
GTP?
Fig. 15-7, p. 241

21. \figures\ch01\pp01262.jpg

22. ROOT AND SHOOT MERISTEMS CYTOKININ FUNCTION
Optimal levels of cytokinins are needed for normal cell division
Root: cytokinin overabundance inhibits cell division
Shoot: cytokinins promote cell division
Cytokinin oxidase dictates meristemic cytokinin concentrations
Mutants can either overproduce or underproduce this enzyme

24. ROOT AND SHOOT MERISTEMS
CYTOKININ FUNCTION

25. Figure 15. 10: The interplay of auxin and cytokinin
Figure 15.10: The interplay of auxin and cytokinin. Differences in the amounts of auxin and cytokinin can change the development of tobacco (Nicotiana tabacum) tissues in culture.
Fig , p. 244

26. CELL DIFFERENTIATION Bacteria: Agrobacterium tumefaciens
CYTOKININ FUNCTION
Bacteria: Agrobacterium tumefaciens

27. Auxin:cytokinin affects cell differentiation in callus tissue
CYTOKININ FUNCTION
Auxin:cytokinin affects cell differentiation in callus tissue
More auxin leads to roots development
More cytokinin leads to shoots development
Skoog and Miller (1965)

28. LEAF SENESCENCE
CYTOKININ FUNCTION
Cytokinin delays leaf senescence
Delay the degradation of chloroplasts
May increase the growing season for agricultural purposes.

29. LEAF SENESCENCE CYTOKININ FUNCTION
Control Sprayed
Delayed leaf senescence, help plants to recover from environmental stresses like
Drought (Rivero et al. 2007)
Flooding (Zhang et al. 2000)
Hyunh et al. 2005

30. Cytokinins induce division in inflorescence tips
INFLORESCENCE GROWTH
CYTOKININ FUNCTION
Cytokinins induce division in inflorescence tips
Leads to more flowering which yields more fruit
“Cytokinin Oxidase Regulates Rice Grain Production”
(Ashikari et al. 2005)
Plants in this study have lower levels of cytokinin oxidase. The plants then produce more fruit.

31. COTYLEDON EXPANSION CYTOKININ FUNCTION
Promotes expansion in cotyledons
Extends the cell wall
Differs from auxin expansion
Unlike auxin, no proton extrusion through the cell wall happens

32. APICAL DOMINANCE CYTOKININ FUNCTION
Antagonistic hormone interaction between cytokinin and auxin
Cytokinin stimulates growth in auxiliary buds
Inhibits shoot elongation
Auxin restrains growth in auxiliary buds
Causes shoot to lengthen.
Mutants that overproduce cytokinins in lateral meristems are bushy.

33. SEEDLING MORPHOLOGY CYTOKININ FUNCTION
Etiolated leaves treated with cytokinins produce more active/productive chloroplasts upon illumination
Dark-germinated seedlings treated with cytokinins
Shortened hypocotyls
Expanded cotyledons
Partial development of etioplasts into chloroplasts

34. SEEDLING MORPHOLOGY CYTOKININ FUNCTION
1 (control)
2 (most concentrated) 3
4 (most dilute)
Average Hypocotyle
(mm)
8.8
2.9
5.4
8.1
Standard
Deviation
1.398
0.567
0.699
0.875
Molarity
(mol/L)
none
4.651x10-4
4.651x10-5
4.651x 10-6

35. REFERENCES CYTOKININS
Ashikari, Motoyuki. “Cytokinin Oxidase Regulates Rice Grain Production.”
Science. 23 June May <
Davies, Peter J. ed. Plant Hormones. Boston: Kluwer Academic Publishers, 2004
Ma, Qing-Hu. “Genetic Engineering of Cytokinins and Their Application to Agriculture.” Critical Reviews in Biotechnology (2008)
InformaWorld. University of California Santa Cruz Lib., Santa Cruz, CA, 1 May < text= >
Mok, David W. S. , Machteld C. Mok, eds. Cytokinins Chemistry, Activity, and Function. Boca Raton:CRC Press Inc, 1994
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