title: Single expressed glycine receptor domains reconstitute functional ion channels without subunit-specific desensitization behavior.
abstract: Cys loop receptors are pentameric arrangements of independent subunits that assemble into functional ion channels. Each subunit shows a domain architecture. Functional ion channels can be reconstituted even from independent, nonfunctional subunit domains, as shown previously for GlyRα1 receptors. Here, we demonstrate that this reconstitution is not restricted to α1 but can be transferred to other members of the Cys loop receptor family. A nonfunctional GlyR subunit, truncated at the intracellular TM3-4 loop by a premature stop codon, can be complemented by co-expression of the missing tail portion of the receptor. Compared with α1 subunits, rescue by domain complementation was less efficient when GlyRα3 or the GABAA/C subunit ρ1 was used. If truncation disrupted an alternative splicing cassette within the intracellular TM3-4 loop of α3 subunits, which also regulates receptor desensitization, functional rescue was not possible. When α3 receptors were restored by complementation using domains with and without the spliced insert, no difference in desensitization was found. In contrast, desensitization properties could even be transferred between α1/α3 receptor chimeras harboring or lacking the α3 splice cassette proving that functional rescue depends on the integrity of the alternative splicing cassette in α3. Thus, an intact α3 splicing cassette in the TM3-4 loop environment is indispensable for functional rescue, and the quality of receptor restoration can be assessed from desensitization properties.
authors.affiliation: From Bioinformatics and.
authors.forename: Heike
authors.initials: H
authors.lastname: Meiselbach
authors.affiliation: Institute of Biochemistry, Emil Fischer Center, Friedrich-Alexander University Erlangen-Nürnberg, Fahrstrasse 17, 91054 Erlangen, Germany.
authors.forename: Nico
authors.initials: N
authors.lastname: Vogel
authors.affiliation: the Institute for Clinical Neurobiology, University of Würzburg, Versbacherstrasse 5, 97078 Würzburg, Germany, and.
authors.forename: Georg
authors.initials: G
authors.lastname: Langlhofer
authors.affiliation: Institute of Biochemistry, Emil Fischer Center, Friedrich-Alexander University Erlangen-Nürnberg, Fahrstrasse 17, 91054 Erlangen, Germany.
authors.forename: Sabine
authors.initials: S
authors.lastname: Stangl
authors.affiliation: the Institute for Clinical Neurobiology, University of Würzburg, Versbacherstrasse 5, 97078 Würzburg, Germany, and.
authors.forename: Barbara
authors.initials: B
authors.lastname: Schleyer
authors.affiliation: the Biochemistry Department, German University Cairo, New Cairo City, Cairo 11835, Egypt.
authors.forename: Lamia'a
authors.initials: L
authors.lastname: Bahnassawy
authors.affiliation: From Bioinformatics and.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Sticht
authors.affiliation: the Biochemistry Department, German University Cairo, New Cairo City, Cairo 11835, Egypt.
authors.forename: Hans-Georg
authors.initials: HG
authors.lastname: Breitinger
authors.affiliation: Institute of Biochemistry, Emil Fischer Center, Friedrich-Alexander University Erlangen-Nürnberg, Fahrstrasse 17, 91054 Erlangen, Germany.
authors.forename: Cord-Michael
authors.initials: CM
authors.lastname: Becker
authors.affiliation: Institute of Biochemistry, Emil Fischer Center, Friedrich-Alexander University Erlangen-Nürnberg, Fahrstrasse 17, 91054 Erlangen, Germany, the Institute for Clinical Neurobiology, University of Würzburg, Versbacherstrasse 5, 97078 Würzburg, Germany, and Villmann_C@ukw.de.
authors.forename: Carmen
authors.initials: C
authors.lastname: Villmann
mesh_terms: D017398:Alternative Splicing; D000595:Amino Acid Sequence; D019921:Biotinylation; D003545:Cysteine / Q000737:chemistry; D005816:Genetic Complementation Test; D005998:Glycine / Q000737:chemistry; D057809:HEK293 Cells; D006801:Humans; D007473:Ion Channels / Q000737:chemistry*; D008969:Molecular Sequence Data; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D018009:Receptors, Glycine / Q000737:chemistry*; D017386:Sequence Homology, Amino Acid
chemical_list: D007473:Ion Channels; D018009:Receptors, Glycine; C113965:glycine receptor alpha3 subunit; D003545:Cysteine; D005998:Glycine
keywords: Desensitization; Domain Architecture; Glycine Receptor; Glycine Receptors; Ion Channel; Ion Channels; Neurobiology; Neurophysiology; Neurotransmitter Receptor; Patch Clamp