title: Function coupling of otoferlin with GAD65 acts to modulate GABAergic activity.
abstract: Otoferlin, an integral membrane protein implicated in a late stage of exocytosis, has been reported to play a critical role in hearing although the underlying mechanisms remain elusive. However, its widespread tissue distribution infers a more ubiquitous role in synaptic vesicle trafficking. Glutamate, an excitatory neurotransmitter, is converted to its inhibitory counterpart, γ-aminobutyric acid (GABA), by L-glutamic acid decarboxylase (GAD), which exists in soluble (GAD67) and membrane-bound (GAD65) forms. For the first time, we have revealed a close association between otoferlin and GAD65 in both HEK293 and neuronal cells, including SH-SY5Y neuroblastoma and primary rat hippocampus cells, showing a direct interaction between GAD65 and otoferlin's C2 domains. In primary rat hippocampus cells, otoferlin and GAD65 co-localized in a punctate pattern within the cell body, as well as in the axon along the path of vesicular traffic. Significantly, GABA is virtually abolished in otoferlin-knockdown neuronal cells whereas otoferlin overexpression markedly increases endogenous GABA. GABA attenuation in otoferlin-knockdown primary cells is correlated with diminished L-type calcium current. This previously unknown and close correlation demonstrates that otoferlin, through GAD65, modulates GABAergic activity. The discovery of otoferlin-GAD65 functional coupling provides a new avenue for understanding the molecular mechanism by which otoferlin functions in neurological pathways.
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Wu
authors.initials: W
authors.lastname: Wu
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Mona N
authors.initials: MN
authors.lastname: Rahman
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Jun
authors.initials: J
authors.lastname: Guo
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Natalie
authors.initials: N
authors.lastname: Roy
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Lihua
authors.initials: L
authors.lastname: Xue
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada Present address: Department of Anesthesiology & Perioperative Care, University of California Irvine, Irvine, CA, USA.
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Cahill
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada.
authors.forename: Shetuan
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Department of Biomedical and Molecular Sciences, Queen's University, Kingston, Ontario K7L 3N6, Canada jia@queensu.ca.
authors.forename: Zongchao
authors.initials: Z
authors.lastname: Jia
mesh_terms: D000818:Animals; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D059330:GABAergic Neurons / Q000502:physiology*; D005968:Glutamate Decarboxylase / Q000502:physiology*; D057809:HEK293 Cells; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D008565:Membrane Proteins / Q000502:physiology*; D021381:Protein Transport; D017207:Rats, Sprague-Dawley; D005680:gamma-Aminobutyric Acid / Q000096:biosynthesis
chemical_list: D020746:Calcium Channels, L-Type; D008565:Membrane Proteins; C533993:Otof protein, rat; D005680:gamma-Aminobutyric Acid; D005968:Glutamate Decarboxylase; C401141:glutamate decarboxylase 2
keywords: GABA; GAD65; exocytosis; neurotransmitter; otoferlin