title: The glutamic acid-rich protein is a gating inhibitor of cyclic nucleotide-gated channels.
abstract: The cyclic nucleotide-gated (CNG) cation channel of rod photoreceptors is a heterotetramer consisting of homologous CNGA1 and CNGB1a subunits. While CNGA1 is indispensable for channel activation, the specific role of CNGB1a in this process has remained elusive. Here, we show that the N-terminal glutamic acid-rich protein (GARP) domain of CNGB1a and soluble GARP2, which corresponds to the proximal portion of the GARP domain, act as autoinhibitory domains that decrease the opening probability of the CNG channel. In the presence of mutations that structurally impair the cyclic nucleotide-binding domain (CNBD) of CNGB1a, the GARP domain completely abolishes channel activity. In agreement with an inhibitory function of GARP, the activity of mutant CNG channels could be fully restored by deletion of the GARP domain. We identified two sequences within the GARP domain that confer most of the inhibitory effect and demonstrate that the profound inhibition imposed by the GARP domain is caused by direct and autonomous protein-protein interaction with the CNG channel complex. In wild-type rod CNG channels, this inhibitory effect can be relieved by binding of cGMP to the CNBD of CNGB1a. In conclusion, we propose that the N terminus of CNGB1a and soluble GARPs act as molecular gate keepers that control the activation of heteromeric rod CNG channels. Our results suggest that the GARP domain has evolved in rod photoreceptors to reduce current noise resulting from openings of CNG channels in the absence of cGMP.
authors.affiliation: Munich Center for Integrated Protein Science, Ludwig-Maximilians-Universität München, D-81377 München, Germany.
authors.forename: Stylianos
authors.initials: S
authors.lastname: Michalakis
authors.forename: Xiangang
authors.initials: X
authors.lastname: Zong
authors.forename: Elvir
authors.initials: E
authors.lastname: Becirovic
authors.forename: Verena
authors.initials: V
authors.lastname: Hammelmann
authors.forename: Thomas
authors.initials: T
authors.lastname: Wein
authors.forename: Klaus T
authors.initials: KT
authors.lastname: Wanner
authors.forename: Martin
authors.initials: M
authors.lastname: Biel
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D055592:Biophysical Phenomena / Q000187:drug effects / Q000235:genetics; D001710:Biotin / Q000031:analogs & derivatives / Q000378:metabolism; D019921:Biotinylation; D002461:Cell Line, Transformed; D000242:Cyclic AMP / Q000494:pharmacology; D006152:Cyclic GMP / Q000494:pharmacology; D054815:Cyclic Nucleotide-Gated Cation Channels / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D004558:Electric Stimulation; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005786:Gene Expression Regulation / Q000187:drug effects / Q000235:genetics; D006801:Humans; D047468:Immunoprecipitation / Q000379:methods; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D008164:Luminescent Proteins / Q000235:genetics; D008564:Membrane Potentials / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D051379:Mice; D018345:Mice, Knockout; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D009419:Nerve Tissue Proteins / Q000235:genetics / Q000378:metabolism*; D018408:Patch-Clamp Techniques / Q000379:methods; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000235:genetics / Q000502:physiology; D013388:Succinimides / Q000378:metabolism; D014162:Transfection / Q000379:methods
chemical_list: C512817:CA11 protein, human; C518243:CNGA1 protein, human; D054815:Cyclic Nucleotide-Gated Cation Channels; D008164:Luminescent Proteins; D009419:Nerve Tissue Proteins; D013388:Succinimides; C065130:sulfo-N-hydroxysuccinimide-biotin; D001710:Biotin; D000242:Cyclic AMP; D006152:Cyclic GMP