title: DOC2A and DOC2B are sensors for neuronal activity with unique calcium-dependent and kinetic properties.
abstract: Elevation of the intracellular calcium concentration ([Ca2+]i) to levels below 1 microm alters synaptic transmission and induces short-term plasticity. To identify calcium sensors involved in this signalling, we investigated soluble C2 domain-containing proteins and found that both DOC2A and DOC2B are modulated by submicromolar calcium levels. Fluorescent-tagged DOC2A and DOC2B translocated to plasma membranes after [Ca2+]i elevation. DOC2B translocation preceded DOC2A translocation in cells co-expressing both isoforms. Half-maximal translocation occurred at 450 and 175 nm[Ca2+]i for DOC2A and DOC2B, respectively. This large difference in calcium sensitivity was accompanied by a modest kinetic difference (halftimes, respectively, 2.6 and 2.0 s). The calcium sensitivity of DOC2 isoforms can be explained by predicted topologies of their C2A domains. Consistently, neutralization of aspartates D218 and D220 in DOC2B changed its calcium affinity. In neurones, both DOC2 isoforms were reversibly recruited to the plasma membrane during trains of action potentials. Consistent with its higher calcium sensitivity, DOC2B translocated at lower depolarization frequencies. Styryl dye uptake experiments in hippocampal neurones suggest that the overexpression of mutated DOC2B alters the synaptic activity. We conclude that both DOC2A and DOC2B are regulated by neuronal activity, and hypothesize that their calcium-dependent translocation may regulate synaptic activity.
authors.affiliation: Department of Functional Genomics, Center for Neurogenomics and Cognition Research, Vrije Universiteit (VU) and VU Medical Centre, Amsterdam, the Netherlands. sander@cncr.vu.nl
authors.forename: Alexander J A
authors.initials: AJ
authors.lastname: Groffen
authors.forename: Reut
authors.initials: R
authors.lastname: Friedrich
authors.forename: Elisabeth C
authors.initials: EC
authors.lastname: Brian
authors.forename: Uri
authors.initials: U
authors.lastname: Ashery
authors.forename: Matthijs
authors.initials: M
authors.lastname: Verhage
mesh_terms: D000818:Animals; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002135:Calcium-Binding Proteins / Q000378:metabolism*; D002415:Cats; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002478:Cells, Cultured; D002614:Chelating Agents / Q000494:pharmacology; D019439:Chromaffin Cells; D006224:Cricetinae; D003952:Diagnostic Imaging / Q000379:methods; D004305:Dose-Response Relationship, Drug; D004307:Dose-Response Relationship, Radiation; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D004558:Electric Stimulation / Q000379:methods; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology; D006801:Humans; D007476:Ionophores / Q000494:pharmacology; D008954:Models, Biological; D008958:Models, Molecular; D016296:Mutagenesis / Q000502:physiology; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism* / Q000528:radiation effects; D011189:Potassium Chloride / Q000494:pharmacology; D017434:Protein Structure, Tertiary / Q000502:physiology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D011726:Pyridinium Compounds / Q000493:pharmacokinetics; D000644:Quaternary Ammonium Compounds / Q000493:pharmacokinetics; D051381:Rats; D016415:Sequence Alignment / Q000379:methods; D013997:Time Factors; D014162:Transfection / Q000379:methods
chemical_list: D002135:Calcium-Binding Proteins; D002614:Chelating Agents; C091524:DOC2A protein, human; C490420:DOC2B protein, human; C092350:FM 4-64; D007476:Ionophores; D009419:Nerve Tissue Proteins; D011726:Pyridinium Compounds; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins; D000001:Calcimycin; D004533:Egtazic Acid; D011189:Potassium Chloride; C025603:1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid; D002118:Calcium