Phototropin

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1. Phototropin Dr. Brijbhan Yadav

2. Phototropin It is a Flavoprotein which act as a blue light receptor. They are light activating serine/threonine protein kinases . Optimize the photosynthetic efficiency of plants . This include phototropism, light induced stomatal opening and chloroplast movements in response to changes in light intensity.

3. Phototropin is blue light (400-500nm) photoreceptor of plants . The chromophore for Phototropin is FMN (a Flavin), so Phototropin is a flavoprotein . The N-terminal half of Phototropin has 2 LOV domains each binds FMN and C-Terminal kinase domain that has a characteristic activity of serine/threonine kinase .   Phototropins membrane associated proteins & located at Plasma membrane & functions as Ser / Thr kinase . There are two different types of Phototropins in Arabidopsis they have been termed as PHOT1 and PHOT2 . Phototropin also absorbs UV-A light (320-400 nm) so Phototropin functions can also be mediated by UV-A light. Phototropin

4. PHOTOTROPIN GENES There are two different Phototropins in Arabidopsis that exhibits overlapping function in addition to having unique physiological role . The two Phototropins are Phot1 Phot2 Phot1 help in cotyledon/hypocotyl growth . The phot1 mutant lacks hypocotyl Phototropism in response to low-intensity blue light but retains a phototropic response at higher intensities.

5. STRUCTURAL DOMAIN OF PHOTOTROPIN LOV1 LOV2 FMN FMN Serine threonine kinase N C 1. Phototropin contain two light sensing Light-oxygen-voltage(LOV) Domains, LOV1 and LOV2 . 2. LOV1 and LOV2 bind a chromophore FMN . 3. Blue light irradiation/photoexcitation of protein bound FMN cause a conformational change of phototropin that triggers auto phosphorylation and start the sensory transduction cascade(phototropin signalling)

6. Photoexcitation of LOV domain result in activation Of C-terminal kinase domain, Which leads to auto phosphorylation of multiple Serine residues. In dark molecules binds non covalently to LOV domains In presence of blue light ,FMN molecule become covalently bound to LOV Domain. DARK Blue Light LOV1 LOV2 FMN FMN Serine threonine kinase N C P P

7. MM.DD.20XX LOV1 LOV2 FMN FMN Serine/threonine kinase N J α -Helix LOV1 LOV2 FMN FMN Serine/threonine kinase J α -Helix Substrate Phototropin response DARK Dephospho rhylation PP2A P P P Blue Light Auto- phosphorhylation Mechanism of Action

8. In the dark, FMN molecules are non-covalently bound to LOV domains. LOV2 domain "cages" and inhibits the activity of kinase domain. Upon blue light absorption FMN molecule becomes covalently bound to LOV domains which results in conformational change in protein & activation of LOV domains Which results in activation of Kinase Domain due to unfolding of Ja -helix Which leads to Autophosphorylation of kinase domain on multiple serine residues which activates Phototropin & starts signaling pathway A type 2A Protein Phosphatase mediates Dephosphorylation and inactivation of Phototropin in the dark

9. MUTATIONS CASE 1 PHOT1 mutant lacks phototropism response in low intensity blue light but retains a phototropic response at higher light intensities. PHOT2 mutant lacks phototropism response at higher light intensities but retains phototropic response at low light intensities. PHOT1 & PHOT2 double mutants lacks phototropism response. CASE 1 Stomata from PHOT1/ PHOT2 double mutants failed to exhibit specific blue light response where as in the single PHOT1 or PHOT2 mutant the blue light response is only slightly affected .

10. CASE 3 PHOT1 & PHOT2 double mutants does not exhibits chloroplast movements in response to blue light . PHOT2 mutant does not exhibits avoidance response in response to high light intensities.

11. Stomata from the phot1/phot2 double mutant fail to exhibit a specific blue-light response, whereas in the single phot1 or phot2 mutant the blue light response is only slightly affected . The phot1 mutant lacks hypocotyl phototropism in response to low-intensity blue light but retains a phototropic response at higher intensities MUTATION CASE

12. Function Chloroplast movement in leaves is an adaptive feature that occurs in order to control light absorption and prevent photodamage. Phototropin regulates chloroplast movement Case 1 Low Light Accumulation Response Under low light intensity When incident radiations are weak/ low chloroplasts move at the upper and lower surfaces of the mesophyll cells to maximize the light absorption called accumulation response. Both PHOT1 & PHOT2 mediates the Accumulation response. Vacuole

13. Chloroplast Movement Chloroplast movement in leaves is a adaptive features that occur in order to control light absorption and prevent photodamage .

14. High Light Case 2- In High Light Intensity Under high light intensity chloroplast moves to the cell surfaces that parallel to the incident light thus minimizing light absorption to prevent the photodamage called Avoidance Response only PHOT2 mediates the Avoidance Response. Case 3- During night time under dark conditions PHOT2 mediates chloroplast movement to the lower surface of the mesophyll cells called dark positioning . In Dark Dark Positioning Avoidance Response Vacuole Vacuole

15. Phototropism Auxin Will Accumulate On shaded side Phototropin mediate the auxin gradient change of plant in light. The plant will bend towards the light. The darker side will have high concentration of auxin as compare to light side. Rapid cell division and elongation at darker side due to high concentration of auxin.

16.   Both PHOT1 & PHOT2 mediates Phototropism (Bending of plants towards blue light).   Phototropin regulates chloroplast movements via. F-actin Filament with help of F-actin binding protein CHUP1(chloroplast Unusual Positioning 1). CHUP1 is localized in chloroplast envelop & function in chloroplast movement.

17. Both PHOT1 & PHOT2 Promotes stomatal opening under blue light.