Light-driven translocation of the protein phosphatase 2A complex regulates light/dark dephosphorylation of phosducin and rhodopsin.
In steps of protein purification of bovine retinal protein phosphatase 2A (PP2A), phosducin dephosphorylation activity peaks coelute with a PP2A enzyme complex, shown by peptide sequence analysis to contain a B' subunit, B56 epsilon. Other PP2A complexes with a slightly larger (56.5 kDa) B' subunit (sequenced to be B56 alpha) or with the B alpha regulatory subunit have no phosducin dephosphorylation activity. Upon exposure to light, a significant increase in the immunoreactive protein level of the A, C, and B56 epsilon PP2A subunits is observed in the cytosolic fraction of mouse retina, the phosducin dephosphorylation of which occurs rapidly. During dark exposure, these subunits translocate to the membrane fraction where rhodopsin is slowly dephosphorylated. This PP2A redistribution occurs in less than 1.5 min and is dependent upon light and not upon an intrinsic circadian rhythm. Forty times more of the A subunit (approximately 20 ng/mouse retina) and 9 times more of the C subunit (approximately 4 ng/mouse retina) than of the B56 epsilon subunit (approximately 0.45 ng/mouse retina) redistribute, which suggests that the predominant form of the PP2A enzyme complex on the membrane in the dark is a dimer, consisting of only A and C subunits. We observe that the dimer favors phosphorylated opsin as a substrate, while the trimer, particularly the enzyme complex with the B56 epsilon subunit, greatly prefers phosphorylated phosducin, with an activity several hundred times those of other substrates that were tested. This light-driven PP2A translocation provides a potential mechanism for efficient dephosphorylation of two critical photoreceptor transduction proteins, cytosolic phosducin and membrane-bound rhodopsin, by the same enzyme.
Brown
Bruce M
BM
The Mary D. Allen Laboratory for Vision Research, Doheny Eye Institute, and Department of Cell and Neurobiology, Keck School of Medicine of the University of Southern California, Los Angeles, California 90089-9112, USA.
Carlson
Brian L
BL
Zhu
Xuemei
X
Lolley
Richard N
RN
Craft
Cheryl M
CM
D000818:Animals; D019393:Arrestin / Q000378:metabolism; D002417:Cattle; D003624:Darkness; D005136:Eye Proteins / Q000378:metabolism*; D020555:GTP-Binding Protein Regulators; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D015151:Immunoblotting; D008027:Light; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010446:Peptide Fragments / Q000737:chemistry; D010749:Phosphoprotein Phosphatases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011494:Protein Kinases / Q000378:metabolism*; D054648:Protein Phosphatase 2; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D021381:Protein Transport / Q000502:physiology*; D012160:Retina / Q000201:enzymology / Q000528:radiation effects; D012243:Rhodopsin / Q000378:metabolism*; D017299:Rod Opsins / Q000378:metabolism; D013379:Substrate Specificity
D019393:Arrestin; D005136:Eye Proteins; D020555:GTP-Binding Protein Regulators; D010446:Peptide Fragments; D010750:Phosphoproteins; D021122:Protein Subunits; D017299:Rod Opsins; C065631:phosducin; D012243:Rhodopsin; D011494:Protein Kinases; D010749:Phosphoprotein Phosphatases; D054648:Protein Phosphatase 2