Abstract
It has been recognized for a long time that the secretory granules of mast cells are acidic, but the functional importance of maintaining an acidic pH in the mast cell granules is not fully understood. Here we addressed this issue by examining the effects of raising the pH of the mast cell secretory granules. Mast cells were incubated with bafilomycin A1, an inhibitor of the vacuolar-type ATPase proton pump. Supporting a role of vacuolar-type ATPase in mast cell granule acidification, bafilomycin A1 treatment caused a robust increase in granule pH. This was accompanied by marked effects on mast cell granules, including swelling and acquisition of vacuole-like morphology. Moreover, bafilomycin A1 caused extensive, yet selective effects on the granule content. These included aberrant processing of pro-carboxypeptidase A3 and a reduction in the level of intracellular histamine, the latter being accompanied by an increase in extracellular histamine. In contrast, the storage of β-hexosaminidase, a prototype lysosomal hydrolase known to be stored in mast cell granules, was not affected by abrogation of granule acidification. Moreover, bafilomycin A1 caused a reduction of tryptase enzymatic activity and appearance of tryptase degradation products. Tryptase inhibition prevented the formation of such degradation products, suggesting that the pH elevation causes tryptase to undergo autoproteolysis. Taken together, our findings reveal that mast cell secretory granule homeostasis is critically dependent on an acidic milieu.
Highlights
Mast cells are hematopoietic cells with a large impact on various immune reactions including, in particular, allergic conditions but mast cells have been implicated in numerous additional pathologies such as arthritis and cancer.[1]
These are filled with large amounts of different bioactive compounds such as various amines, cytokines, proteoglycans of serglycin type and different mast cell-restricted proteases, the latter including chymase, tryptase and CPA3.2,3 When mast cells undergo degranulation, typically induced by IgE receptor crosslinking[4] or by engagement of the MRGPRX2/MRGPRB2 receptor,[5] the contents of the granules are expelled to the exterior and can cause a profound inflammatory reaction.[2]
As V-ATPase is known to have a prominent role in the acidification of lysosome-like organelles in numerous other cell types,[10,11] we reasoned that it was likely that this ATPase has an analogous function in the regulation of pH in mast cell secretory granules
Summary
Gunnar Pejler*,1,2,3, Jun Mei Hu Frisk[1,3], Daniel Sjöström[1,3], Aida Paivandy[1] and Helena Öhrvik*,1 It has been recognized for a long time that the secretory granules of mast cells are acidic, but the functional importance of maintaining an acidic pH in the mast cell granules is not fully understood. The acidic pH optimum of the lysosomal enzymes ensures that their action is largely confined to this cellular compartment, whereas the neutral pH milieu of the extracellular space or the cytosol will cause their inactivation and confer protection from their potentially damaging effects It has been known for a long time that the mast cell granules have a low pH,[7,8,9] the impact of the acidic pH milieu on mast cell granule functionality and homeostasis is not fully understood. This investigation reveals a prominent role of acidic pH in regulating the homeostasis on mast cell secretory granules
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