Direct Recording of Trans-Plasma Membrane Electron Currents Mediated by a Member of the Cytochrome b561 Family of Soybean.
Trans-plasma membrane electron transfer is achieved by b-type cytochromes of different families, and plays a fundamental role in diverse cellular processes involving two interacting redox couples that are physically separated by a phospholipid bilayer, such as iron uptake and redox signaling. Despite their importance, no direct recordings of trans-plasma membrane electron currents have been described in plants. In this work, we provide robust electrophysiological evidence of trans-plasma membrane electron flow mediated by a soybean (Glycine max) cytochrome b561 associated with a dopamine β-monooxygenase redox domain (CYBDOM), which localizes to the plasma membrane in transgenic Arabidopsis (Arabidopsis thaliana) plants and CYBDOM complementary RNA-injected Xenopus laevis oocytes. In oocytes, two-electrode voltage clamp experiments showed that CYBDOM-mediated currents were activated by extracellular electron acceptors in a concentration- and type-specific manner. Current amplitudes were voltage dependent, strongly potentiated in oocytes preinjected with ascorbate (the canonical electron donor for cytochrome b561), and abolished by mutating a highly conserved His residue (H292L) predicted to coordinate the cytoplasmic heme b group. We believe that this unique approach opens new perspectives in plant transmembrane electron transport and beyond.
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.).
Cristiana
C
Picco
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.).
Joachim
http://orcid.org/0000-0002-7376-6635
J
Scholz-Starke
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.).
Margherita
http://orcid.org/0000-0003-4990-7554
M
Festa
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.).
Alex
http://orcid.org/0000-0002-2628-1176
A
Costa
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.).
Francesca
http://orcid.org/0000-0002-2103-4870
F
Sparla
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.) paolo.trost@unibo.it carpaneto@ge.ibf.cnr.it.
Paolo
http://orcid.org/0000-0002-6347-8701
P
Trost
Institute of Biophysics, Consiglio National Research Council, 16149 Genova, Italy (C.P., J.S.-S., M.F., A.Ca.);Department of Biosciences, University of Milan, 20133 Milan, Italy (A.Co.);Institute of Biophysics, Consiglio Nazionale delle Ricerche, 20133 Milan, Italy (A.Co.); andDepartment of Pharmacy and Biotechnology (FaBiT), University of Bologna, 40126 Bologna, Italy (F.S., P.T.) paolo.trost@unibo.it carpaneto@ge.ibf.cnr.it.
Armando
http://orcid.org/0000-0002-5060-3657
A
Carpaneto
D000818:Animals; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D001205:Ascorbic Acid / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D003573:Cytochrome b Group / Q000235:genetics / Q000378:metabolism*; D004299:Dopamine beta-Hydroxylase / Q000235:genetics / Q000378:metabolism; D004579:Electron Transport; D055724:Electrophysiological Phenomena / Q000502:physiology; D005292:Ferricyanides / Q000494:pharmacology; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D030821:Plants, Genetically Modified; D017434:Protein Structure, Tertiary; D013025:Glycine max / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis / Q000378:metabolism
D003573:Cytochrome b Group; D005292:Ferricyanides; D010940:Plant Proteins; C023646:cytochrome b561; C007931:hexacyanoferrate III; D004299:Dopamine beta-Hydroxylase; D001205:Ascorbic Acid