Expression and comparative characterization of Gq-coupled invertebrate visual pigments and melanopsin.
A non-visual pigment melanopsin, which is localized in photosensitive retinal ganglion cells and is involved in the circadian photoentrainment and pupillary responses in mammals, is phylogenetically close to the visual pigments of invertebrates, such as insects and cephalopods. Recent studies suggested that melanopsin is a bistable pigment and drives a Gq-mediated signal transduction cascade, like the invertebrate visual pigments. Because detailed electrophysiological properties are somewhat different between the visual cells and the photosensitive ganglion cells, we here expressed and purified the invertebrate visual pigment and melanopsin to comparatively investigate their Gq-activation abilities. We successfully expressed and purified UV and blue light-sensitive visual pigments of the honeybee as well as the amphioxus melanopsin. Although the purified UV-sensitive pigment and the melanopsin lost their bistable nature during purification, reconstitution of the pigments in lipid vesicles resulted in return of the bistable nature. The light-dependent Gq-activation abilities among these reconstituted pigments are similar, suggesting that the electrophysiological differences do not depend on the Gq-activation step but rather on the other signal transduction steps and/or on cell properties. Our findings are also important in that this is the first report describes a heterologous large-scale expression of the Gq-coupled invertebrate visual pigments in cultured cells.
Terakita
Akihisa
A
Department of Biophysics, Graduate School of Science, Kyoto University, Kyoto, Japan. terakita@sci.osaka-cu.ac.jp
Tsukamoto
Hisao
H
Koyanagi
Mitsumasa
M
Sugahara
Michio
M
Yamashita
Takahiro
T
Shichida
Yoshinori
Y
D000818:Animals; D001516:Bees; D002478:Cells, Cultured; D005260:Female; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism*; D015151:Immunoblotting; D007448:Invertebrates / Q000166:cytology / Q000378:metabolism*; D055537:Light Signal Transduction / Q000502:physiology; D008563:Membrane Lipids / Q000378:metabolism; D009448:Neurochemistry / Q000379:methods; D017956:Photoreceptor Cells, Invertebrate / Q000378:metabolism* / Q000528:radiation effects; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism*; D012168:Retinal Pigments / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D017299:Rod Opsins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D013057:Spectrum Analysis; D014466:Ultraviolet Rays; D000099210:Melanopsin
D043802:GTP-Binding Protein alpha Subunits, Gq-G11; D008563:Membrane Lipids; D043562:Receptors, G-Protein-Coupled; D012168:Retinal Pigments; D017299:Rod Opsins; D000099210:Melanopsin