title: The contribution of the SPCA1 Ca2+ pump to the Ca2+ accumulation in the Golgi apparatus of HeLa cells assessed via RNA-mediated interference.
abstract: The secretory-pathway Ca(2+)-ATPase SPCA1 is a thapsigargin-insensitive intracellular Ca(2+) pump found mostly in the Golgi compartment. We have explored the contribution of this Ca(2+) pump to cytosolic Ca(2+) signaling in HeLa cells by using RNA-mediated interference to disrupt its expression. Removal of SPCA1 was confirmed by immunofluorescence with specific anti-SPCA1 antibodies. Measurements of the free Ca(2+) concentration in the lumen of the Golgi apparatus by specifically targeting the Ca(2+)-sensitive luminescent protein aequorin to this organelle revealed that endogenous SPCA1 was responsible for Ca(2+) uptake in a subfraction of the Golgi apparatus. HeLa cells lacking SPCA1 could still set up baseline Ca(2+) spiking when stimulated with histamine, indicating that the SPCA1-containing Ca(2+) store was not absolutely needed to set up these oscillations. However, baseline Ca(2+) oscillations occurred less frequently than in control cells, pointing to a contribution of SPCA1 in the shaping of the cytosolic Ca(2+) signal in HeLa cells.
authors.affiliation: Laboratorium voor Fysiologie, K.U. Leuven Campus Gasthuisberg O/N, Herestraat 49, B-3000, Leuven, Belgium.
authors.forename: Kurt
authors.initials: K
authors.lastname: Van Baelen
authors.forename: Jo
authors.initials: J
authors.lastname: Vanoevelen
authors.forename: Geert
authors.initials: G
authors.lastname: Callewaert
authors.forename: Jan B
authors.initials: JB
authors.lastname: Parys
authors.forename: Humbert
authors.initials: H
authors.lastname: De Smedt
authors.forename: Luc
authors.initials: L
authors.lastname: Raeymaekers
authors.forename: Rosario
authors.initials: R
authors.lastname: Rizzuto
authors.forename: Ludwig
authors.initials: L
authors.lastname: Missiaen
authors.forename: Frank
authors.initials: F
authors.lastname: Wuytack
mesh_terms: D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors; D006056:Golgi Apparatus / Q000378:metabolism*; D006367:HeLa Cells; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D007700:Kinetics; D008856:Microscopy, Fluorescence; D034622:RNA Interference; D011994:Recombinant Proteins / Q000378:metabolism; D019284:Thapsigargin / Q000494:pharmacology; D014162:Transfection
chemical_list: D004791:Enzyme Inhibitors; D011994:Recombinant Proteins; D019284:Thapsigargin; C403624:ATP2C1 protein, human; D000252:Calcium-Transporting ATPases; D002118:Calcium