Subunit 2 (or beta) of retinal rod cGMP-gated cation channel is a component of the 240-kDa channel-associated protein and mediates Ca(2+)-calmodulin modulation.
The cGMP-gated cation channel mediating visual transduction in retinal rods was recently found to comprise at least two subunits, 1 and 2 (or alpha and beta). SDS gels of the purified channel show, in addition to a 63-kDa protein band (subunit 1), a 240-kDa protein band that binds Ca(2+)-calmodulin, a modulator of the channel. To examine any connection between subunit 2 and the 240-kDa protein, cGMP-gated channels formed from the expressed cloned subunits in human embryonic kidney (HEK) 293 cells were tested for Ca(2+)-calmodulin effect. Homooligomeric channels formed by subunit 1 alone showed no sensitivity to Ca(2+)-calmodulin, and neither did heterooligomeric channels formed by subunit 1 and the short alternatively spliced form of subunit 2 (2a). By contrast, the cGMP half-activation constant (K1/2) for heterooligomeric channels formed from subunit 1 and the long form of subunit 2 (2b) was increased 1.5- to 2-fold by Ca(2+)-calmodulin, similar to the increase observed for the native channel. In Western blots of rod outer segment membranes, a subunit 2-specific antibody also recognized the 240-kDa protein. Finally, amino acid sequences derived from peptide fragments of the bovine 240-kDa protein showed approximately 80% identity to regions of subunit 2b of the human channel. These results together suggest that subunit 2b of the rod channel is a component of the 240-kDa protein and that it mediates the Ca(2+)-calmodulin modulation of the channel.
Chen
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TY
Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205.
Illing
M
M
Molday
L L
LL
Hsu
Y T
YT
Yau
K W
KW
Molday
R S
RS
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D002417:Cattle; D054815:Cyclic Nucleotide-Gated Cation Channels; D005136:Eye Proteins / Q000737:chemistry* / Q000502:physiology*; D006801:Humans; D015640:Ion Channel Gating; D007473:Ion Channels / Q000737:chemistry / Q000502:physiology*; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011994:Recombinant Proteins; D012374:Rod Cell Outer Segment / Q000737:chemistry*; D014162:Transfection
D002147:Calmodulin; D054815:Cyclic Nucleotide-Gated Cation Channels; D005136:Eye Proteins; D007473:Ion Channels; D046911:Macromolecular Substances; D011994:Recombinant Proteins; D002118:Calcium