title: The ataxia related G1107D mutation of the plasma membrane Ca2+ATPase isoform 3 affects its interplay with calmodulin and the autoinhibition process.
abstract: The plasma membrane Ca2+ATPases (PMCA pumps) have a long, cytosolic C-terminal regulatory region where a calmodulin-binding domain (CaM-BD) is located. Under basal conditions (low Ca2+), the C-terminal tail of the pump interacts with autoinhibitory sites proximal to the active center of the enzyme. In activating conditions (i.e., high Ca2+), Ca2+-bound CaM displaces the C-terminal tail from the autoinhibitory sites, restoring activity. We have recently identified a G1107D replacement within the CaM-BD of isoform 3 of the PMCA pump in a family affected by X-linked congenital cerebellar ataxia. Here, we investigate the effects of the G1107D replacement on the interplay of the mutated CaM-BD with both CaM and the pump core, by combining computational, biochemical and functional approaches. We provide evidence that the affinity of the isolated mutated CaM-BD for CaM is significantly reduced with respect to the wild type (wt) counterpart, and that the ability of CaM to activate the pump in vitro is thus decreased. Multiscale simulations support the conclusions on the detrimental effect of the mutation, indicating reduced stability of the CaM binding. We further show that the G1107D replacement impairs the autoinhibition mechanism of the PMCA3 pump as well, as the introduction of a negative charge perturbs the contacts between the CaM-BD and the pump core. Thus, the mutation affects both the ability of the pump to optimally transport Ca2+in the activated state, and the autoinhibition mechanism in its resting state.
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Tito
authors.initials: T
authors.lastname: Calì
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Martina
authors.initials: M
authors.lastname: Frizzarin
authors.affiliation: Department of Biosciences, University of Milano, Italy.
authors.forename: Laura
authors.initials: L
authors.lastname: Luoni
authors.affiliation: Shanghai Institute of Advanced Immunochemical Studies, ShanghaiTech University, Shanghai, China.
authors.forename: Francesco
authors.initials: F
authors.lastname: Zonta
authors.affiliation: Institut Pasteur de Montevideo, Uruguay.
authors.forename: Sergio
authors.initials: S
authors.lastname: Pantano
authors.affiliation: Aggeu Magalhães Research Center - CpqAM, Oswaldo Cruz Foundation - FioCruz, Rio de Janeiro, Brazil; Institut Pasteur de Montevideo, Uruguay.
authors.forename: Carlos
authors.initials: C
authors.lastname: Cruz
authors.affiliation: Department of Biosciences, University of Milano, Italy.
authors.forename: Maria Cristina
authors.initials: MC
authors.lastname: Bonza
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Ilenia
authors.initials: I
authors.lastname: Bertipaglia
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Maria
authors.initials: M
authors.lastname: Ruzzene
authors.affiliation: Department of Biosciences, University of Milano, Italy.
authors.forename: Maria Ida
authors.initials: MI
authors.lastname: De Michelis
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Nunzio
authors.initials: N
authors.lastname: Damiano
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Oriano
authors.initials: O
authors.lastname: Marin
authors.affiliation: Department of Neurosciences, Bambino Gesù Children's Hospital, IRCCS, Rome, Italy.
authors.forename: Ginevra
authors.initials: G
authors.lastname: Zanni
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy.
authors.forename: Giuseppe
authors.initials: G
authors.lastname: Zanotti
authors.affiliation: Department of Biology, University of Padova, Italy.
authors.forename: Marisa
authors.initials: M
authors.lastname: Brini
authors.affiliation: Department of Biomedical Sciences, University of Padova, Italy. Electronic address: raffaele.lopreiato@unipd.it.
authors.forename: Raffaele
authors.initials: R
authors.lastname: Lopreiato
authors.affiliation: Venetian Institute of Molecular Medicine, Padova, Italy. Electronic address: ernesto.carafoli@unipd.it.
authors.forename: Ernesto
authors.initials: E
authors.lastname: Carafoli
mesh_terms: D001259:Ataxia / Q000235:genetics* / Q000378:metabolism; D020013:Calcium Signaling; D002147:Calmodulin / Q000378:metabolism*; D006801:Humans; D008958:Models, Molecular; D053499:Plasma Membrane Calcium-Transporting ATPases / Q000235:genetics* / Q000378:metabolism; D017354:Point Mutation
chemical_list: D002147:Calmodulin; D053499:Plasma Membrane Calcium-Transporting ATPases
keywords: Autoinhibition; Calcium signaling; Calmodulin; Plasma membrane calcium ATPases; X-linked cerebellar ataxia