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Domain Swapping to Assess the Mechanistic Basis of Arabidopsis Phototropin 1 Receptor Kinase Activation and Endocytosis by Blue Light

Kaiserli, E and Sullivan, S and Jones, MA and Feeney, KA and Christie, JM (2009) 'Domain Swapping to Assess the Mechanistic Basis of Arabidopsis Phototropin 1 Receptor Kinase Activation and Endocytosis by Blue Light.' The Plant Cell, 21 (10). pp. 3226-3244. ISSN 1040-4651

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Abstract

Phototropins (phot1 and phot2) are plasma membrane?associated receptor kinases that respond specifically to blue and UV wavelengths. In addition to a C-terminal Ser/Thr kinase domain, phototropins contain two N-terminal chromophore binding LOV domains that function as photoswitches to regulate a wide range of enzymatic activities in prokaryotes and eukaryotes. Through domain swapping, we show that the photochemical properties of Arabidopsis thaliana phot1 rely on interactions between LOV1 and LOV2, which are facilitated by their intervening linker sequence. Functional analysis of domain-swap proteins supports a mechanism whereby LOV2 acts as a dark-state repressor of phot1 activity both in vitro and in vivo. Moreover, we find a photoactive role for LOV1 in arresting chloroplast accumulation at high light intensities. Unlike LOV2, LOV1 cannot operate as a dark-state repressor, resulting in constitutive receptor autophosphorylation and accelerated internalization from the plasma membrane. Coexpression of active and inactive forms of phot1 demonstrates that autophosphorylation can occur intermolecularly, independent of LOV1, via light-dependent receptor dimerization in vivo. Indeed, transphosphorylation is sufficient to promote phot1 internalization through a clathrin-dependent endocytic pathway triggered primarily by phosphorylation of Ser-851 within the kinase activation loop. The mechanistic implications of these findings in regard to light-driven receptor activation and trafficking are discussed.

Item Type: Article
Uncontrolled Keywords: Chloroplasts; Plants, Genetically Modified; Arabidopsis; DNA-Binding Proteins; Clathrin; Arabidopsis Proteins; Microscopy, Confocal; Chromatography, Liquid; Reverse Transcriptase Polymerase Chain Reaction; Immunoprecipitation; Endocytosis; Mutagenesis; Protein Binding; Phosphorylation; Light; Molecular Sequence Data; Tandem Mass Spectrometry; Phototropins
Subjects: Q Science > QH Natural history > QH301 Biology
Divisions: Faculty of Science and Health
Faculty of Science and Health > Life Sciences, School of
SWORD Depositor: Elements
Depositing User: Elements
Date Deposited: 12 Jan 2013 12:00
Last Modified: 06 Jan 2022 13:30
URI: http://repository.essex.ac.uk/id/eprint/5080

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