Auxin-responsive genes AIR12 code for a new family of plasma membrane b-type cytochromes specific to flowering plants.
We report here on the identification of the major plasma membrane (PM) ascorbate-reducible b-type cytochrome of bean (Phaseolus vulgaris) and soybean (Glycine max) hypocotyls as orthologs of Arabidopsis (Arabidopsis thaliana) AIR12 (for auxin induced in root cultures). Soybean AIR12, which is glycosylated and glycosylphosphatidylinositol-anchored to the external side of the PM in vivo, was expressed in Pichia pastoris in a recombinant form, lacking the glycosylphosphatidylinositol modification signal and purified from the culture medium. Recombinant AIR12 is a soluble protein predicted to fold into a beta-sandwich domain and belonging to the DOMON (for dopamine beta-monooxygenase N terminus) domain superfamily. It is shown to be a b-type cytochrome with a symmetrical alpha-band at 561 nm, fully reduced by ascorbate, and fully oxidized by monodehydroascorbate radical. AIR12 is a high-potential cytochrome b showing a wide bimodal dependence from the redox potential between +80 mV and +300 mV. Optical absorption and electron paramagnetic resonance analysis indicate that AIR12 binds a single, highly axial low-spin heme, likely coordinated by methionine-91 and histidine-76, which are strongly conserved in AIR12 sequences. Phylogenetic analyses reveal that the auxin-responsive genes AIR12 represent a new family of PM b-type cytochromes specific to flowering plants. Circumstantial evidence suggests that AIR12 may interact with other redox partners within the PM to constitute a redox link between cytoplasm and apoplast.
Preger
Valeria
V
Laboratory of Molecular Plant Physiology, Department of Experimental Evolutionary Biology, University of Bologna, Bologna 40126, Italy. valeria.preger@unibo.it
Tango
Nunzio
N
Marchand
Christophe
C
Lemaire
Stéphane D
SD
Carbonera
Donatella
D
Di Valentin
Marilena
M
Costa
Alex
A
Pupillo
Paolo
P
Trost
Paolo
P
D000595:Amino Acid Sequence; D001205:Ascorbic Acid / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism*; D003573:Cytochrome b Group / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D035264:Flowers / Q000187:drug effects / Q000378:metabolism*; D017343:Genes, Plant; D006418:Heme / Q000378:metabolism; D018546:Hypocotyl / Q000187:drug effects / Q000378:metabolism; D007210:Indoleacetic Acids / Q000494:pharmacology*; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction / Q000187:drug effects; D027805:Phaseolus / Q000235:genetics / Q000378:metabolism*; D010802:Phylogeny; D010843:Pichia; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013025:Glycine max / Q000235:genetics / Q000378:metabolism*; D013045:Species Specificity; D013113:Spin Labels
D003573:Cytochrome b Group; D007210:Indoleacetic Acids; D010940:Plant Proteins; D013113:Spin Labels; D006418:Heme; D001205:Ascorbic Acid