Functional characterization of a constitutively active kinase variant of Arabidopsis phototropin 1

Petersen, J., Inoue, S.-i., Kelly, S. M. , Sullivan, S., Kinoshita, T. and Christie, J. M. (2017) Functional characterization of a constitutively active kinase variant of Arabidopsis phototropin 1. Journal of Biological Chemistry, 292(33), pp. 13843-13852. (doi: 10.1074/jbc.M117.799643) (PMID:28663371) (PMCID:PMC5566536)

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Abstract

Phototropins (phots) are plasma membrane-associated serine/threonine kinases that coordinate a range of processes linked to optimizing photosynthetic efficiency in plants. These photoreceptors contain two light-, oxygen- or voltage-sensing (LOV) domains within their N-terminus, with each binding one molecule of flavin mononucleotide (FMN) as a UV/blue light absorbing chromophore. Although phots contain two LOV domains, light-induced activation of the C-terminal kinase domain and subsequent receptor autophosphorylation is controlled primarily by the A′α-LOV2-Jα photosensory module. Mutations that disrupt interactions between the LOV2-core and its flanking helical segments can uncouple this mode of light regulation. Yet, the impact of these mutations on phot function in Arabidopsis has not been explored. Here, we report that histidine substitution of Arg-472 located within the A′α-helix of Arabidopsis phot1 to histidine results in constitutively activates kinas activity in vitro without affecting LOV2 photochemistry. Expression analysis of phot1 R472H in the phot-deficient mutant confirmed that it is autophosphorylated in darkness in vivo, but was unable to initiate phot1 signaling in the absence of light. Instead, we found that the phot1 R472H mutant is poorly functional under low-light conditions, but can restore phototropism, chloroplast accumulation, stomatal opening, and leaf positioning and expansion at higher light intensities. Our findings suggest that Arabidopsis can adapt to the elevated phosphorylation status of the phot1 R472H mutant by in part reducing its stability, whereas the activity the mutant under high-light conditions can be attributed to additional increases in LOV2-mediated photoreceptor autophosphorylation.

Item Type:Articles
Status:Published
Refereed:Yes
Glasgow Author(s) Enlighten ID:Kelly, Dr Sharon and Christie, Professor John and Sullivan, Dr Stuart and Petersen, Dr Jan
Authors: Petersen, J., Inoue, S.-i., Kelly, S. M., Sullivan, S., Kinoshita, T., and Christie, J. M.
College/School:College of Medical Veterinary and Life Sciences > Institute of Molecular Cell and Systems Biology
Journal Name:Journal of Biological Chemistry
Publisher:American Society for Biochemistry and Molecular Biology, Inc.
ISSN:0021-9258
ISSN (Online):1083-351X
Published Online:29 June 2017
Copyright Holders:Copyright © 2017 The American Society for Biochemistry and Molecular Biology, Inc
First Published:First published in Journal of Biological Chemistry 292(33):13843-13852
Publisher Policy:Reproduced under a Creative Commons License

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Project CodeAward NoProject NamePrincipal InvestigatorFunder's NameFunder RefLead Dept
581491Regulation of Auxin Fluxes Required For Phototropic GrowthJohn ChristieBiotechnology and Biological Sciences Research Council (BBSRC)BB/J016047/1RI MOLECULAR CELL & SYSTEMS BIOLOGY
659801Photoreceptor Engineering to Modulate Plant GrowthJohn ChristieBiotechnology and Biological Sciences Research Council (BBSRC)BB/M002128/1RI MOLECULAR CELL & SYSTEMS BIOLOGY