PMID:17148436

Citation	Fiser, R, Masín, J, Basler, M, Krusek, J, Spuláková, V, Konopásek, I and Sebo, P (2007) Third activity of Bordetella adenylate cyclase (AC) toxin-hemolysin. Membrane translocation of AC domain polypeptide promotes calcium influx into CD11b+ monocytes independently of the catalytic and hemolytic activities. J. Biol. Chem. 282:2808-20
Abstract	The Bordetella adenylate cyclase toxin-hemolysin (CyaA) targets phagocytes expressing the alpha(M)beta2 integrin (CD11b/CD18), permeabilizes their membranes by forming small cation-selective pores, and delivers into cells a calmodulin-activated adenylate cyclase (AC) enzyme that dissipates cytosolic ATP into cAMP. We describe here a third activity of CyaA that yields elevation of cytosolic calcium concentration ([Ca2+]i) in target cells. The CyaA-mediated [Ca2+]i increase in CD11b+ J774A.1 monocytes was inhibited by extracellular La3+ ions but not by nifedipine, SK&F 96365, flunarizine, 2-aminoethyl diphenylborinate, or thapsigargin, suggesting that influx of Ca2+ into cells was not because of receptor signaling or opening of conventional calcium channels by cAMP. Compared with intact CyaA, a CyaA-AC- toxoid unable to generate cAMP promoted a faster, albeit transient, elevation of [Ca2+]i. This was not because of cell permeabilization by the CyaA hemolysin pores, because a mutant exhibiting a strongly enhanced pore-forming activity (CyaA-E509K/E516K), but unable to deliver the AC domain into cells, was also unable to elicit a [Ca2+]i increase. Further mutations interfering with AC translocation into cells, such as proline substitutions of glutamate residues 509 or 570 or deletion of the AC domain as such, reduced or ablated the [Ca2+]i-elevating capacity of CyaA. Moreover, structural alterations within the AC domain, because of insertion of various oligopeptides, differently modulated the kinetics and extent of Ca2+ influx elicited by the respective AC- toxoids. Hence, the translocating AC polypeptide itself appears to participate in formation of a novel type of membrane path for calcium ions, contributing to action of CyaA in an unexpected manner.
Links	PubMed Online version:10.1074/jbc.M609979200
Keywords	Adenosine Triphosphate/metabolism; Adenylate Cyclase/metabolism; Adenylate Cyclase Toxin/chemistry; Adenylate Cyclase Toxin/genetics; Adenylate Cyclase Toxin/isolation & purification; Adenylate Cyclase Toxin/metabolism; Amino Acid Substitution; Animals; Antigens, CD11b/physiology; Biological Transport; Calcium/metabolism; Catalysis; Cell Line; Cell Membrane/physiology; Cyclic AMP/metabolism; Hemolysis; Macrophages/physiology; Mice; Monocytes/physiology; Mutagenesis, Site-Directed; Polymerase Chain Reaction; Recombinant Proteins/chemistry; Recombinant Proteins/isolation & purification; Recombinant Proteins/metabolism; Sheep
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
BORPE:CYAA		GO:0007204: elevation of cytosolic calcium ion concentration	ECO:0000314:		P		Figure 1.	complete CACAO 4613
BORPE:CYAA		GO:2001170: negative regulation of ATP biosynthetic process	ECO:0000314:		P		Figure 3.	complete CACAO 4617
BORPE:CYAA		GO:0004016: adenylate cyclase activity	ECO:0000314:		F		Table 1.	complete CACAO 4618
BORPE:CYAA		GO:0044179: hemolysis in other organism	ECO:0000314:		P		Table 1.	complete CACAO 4619
BORPE:CYAA	involved_in	GO:0044179: hemolysis in other organism	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete
BORPE:CYAA	enables	GO:0004016: adenylate cyclase activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
BORPE:CYAA	involved_in	GO:0007204: positive regulation of cytosolic calcium ion concentration	ECO:0000314: direct assay evidence used in manual assertion		P		Seeded From UniProt	complete

References

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