Journal
CELL CALCIUM
Volume 54, Issue 5, Pages 350-361Publisher
ELSEVIER SCI LTD
DOI: 10.1016/j.ceca.2013.08.004
Keywords
Calcium homeostasis; ATPase; Ion exchanger; Aequorin; Calcium transporters
Categories
Funding
- NSF [DBI/0722494]
- American Heart Association [09BGIA2330036]
- OCAST [HR12-167]
- [NIH P30 DK 089507]
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Pseudomonas aeruginosa is an opportunistic human pathogen causing severe acute and chronic infections. Earlier we have shown that calcium (Ca2+) induces P. aeruginosa biofilm formation and production of virulence factors. To enable further studies of the regulatory role of Ca2+, we characterized Ca2+ homeostasis in P. aeruginosa PAO1 cells. By using Ca2+-binding photoprotein aequorin, we determined that the concentration of free intracellular Ca2+ ([Ca2+](in)) is 0.14 +/- 0.05 mu M. In response to external Ca2+, the [Ca2+](in) quickly increased at least 13-fold followed by a multi-phase decline by up to 73%. Growth at elevated Ca2+ modulated this response. Treatment with inhibitors known to affect Ca2+ channels, monovalent cations gradient, or P-type and F-type ATPases impaired [Ca2+](in) response, suggesting the importance of the corresponding mechanisms in Ca2+ homeostasis. To identify Ca2+ transporters maintaining this homeostasis, bioinformatic and LC-MS/MS-based membrane proteomic analyses were used. [Ca2+](in) homeostasis was monitored for seven Ca2+-affected and eleven bioinformatically predicted transporters by using transposon insertion mutants. Disruption of P-type ATPases PA2435, PA3920, and ion exchanger PA2092 significantly impaired Ca2+ homeostasis. The lack of PA3920 and vanadate treatment abolished Ca2+-induced swarming, suggesting the role of the P-type ATPase in regulating P. aeruginosa response to Ca2+. (C) 2013 Elsevier Ltd. All rights reserved.
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