Ca2+ accumulation into acidic organelles mediated by Ca2+- and vacuolar H+-ATPases in human platelets

Abstract

Most physiological agonists increase cytosolic free [Ca2+]c (cytosolic free Ca2+ concentration) to regulate a variety of cellular processes. How different stimuli evoke distinct spatiotemporal Ca2+ responses remains unclear, and the presence of separate intracellular Ca2+ stores might be of great functional relevance. Ca2+ accumulation into intracellular compartments mainly depends on the activity of Ca2+- and H+-ATPases. Platelets present two separate Ca2+ stores differentiated by the distinct sensitivity to thapsigargin and TBHQ [2,5-di-(t-butyl)-1,4-hydroquinone]. Although one store has long been identified as the dense tubular system, the nature of the TBHQ-sensitive store remains uncertain. Treatment of platelets with GPN (glycylphenylalanine-2-naphthylamide) impaired Ca2+ release by TBHQ and reduced that evoked by thrombin. In contrast, GPN did not modify Ca2+ mobilization stimulated by ADP or AVP ([arginine]vasopressin). Treatment with nigericin, a proton carrier, and bafilomycin A1, an inhibitor of the vacuolar H+-ATPase, to dissipate the proton gradient into acidic organelles induces a transient increase in [Ca2+]c that was abolished by previous treatment with the SERCA (sarcoplasmic/endoplasmic-reticulum Ca2+-ATPase) 3 inhibitor TBHQ. Depleted acidic stores after nigericin or bafilomycin A1 were refilled by SERCA 3. Thrombin, but not ADP or AVP, reduces the rise in [Ca2+]c evoked by nigericin and bafilomycin A1. Our results indicate that the TBHQ-sensitive store in human platelets is an acidic organelle whose Ca2+ accumulation is regulated by both Ca2+- and vacuolar H+-ATPases.

Ca2+/H+exchange by acidic organelles regulates cell migration in vivo

“…CAXs identified here join SER CA3 and TMEM165 as credible candidates for filling acidic organelles with Ca 2+ (López et al, 2005;Demaegd et al, 2013). Importantly, Ca 2+ /H + transport emerges as a novel way to control cellular migration and likely other Ca 2+ -dependent events in animals.…”

STIM1 and STIM2 Are Located in the Acidic Ca2+ Stores and Associates with Orai1 upon Depletion of the Acidic Stores in Human Platelets

“…It has long been known that Ca 2ϩ is stored in the endoplasmic reticulum (ER), which is the best-studied agonistreleasable Ca 2ϩ store; however, a number of studies have revealed that Ca 2ϩ is also dynamically accumulated in a number of acidic organelles (2), including secretory granules, lysosomes and lysosome-related organelles, endosomes, and vesicles of the Golgi complex (2)(3)(4)(5)(6). ER Ca 2ϩ store depletion has been reported to be sensed by STIM1 (stromal interaction molecule-1) (7-9).…”

“…ER Ca 2ϩ store depletion has been reported to be sensed by STIM1 (stromal interaction molecule-1) (7-9). Although STIM1 was originally identified as a surface membrane protein in stromal cells (10) 4 The abbreviations used are: SOCE, store-operated calcium entry; ER, endoplasmic reticulum; HBS, HEPES-buffered saline; hTRPC1 and hTRPC6, human canonical TRP1 and TRP6, respectively; IP 3 , inositol 1,4,5-trisphosphate; TBHQ, 2,5-di-(tert-butyl)-1,4-hydroquinone; TG, thapsigargin; SERCA, sarco/endoplasmic reticulum Ca 2ϩ -ATPase; V-ATPase, vacuolar proton-ATPase; NAADP, nicotinic acid adenine dinucleotide phosphate; DTS, dense tubular system; AM, acetoxymethyl ester; BAPTA-AM, 1,2-bis(2-aminophenoxy)ethane-N,N,NЈ,NЈ-tetraacetic acid tetrakis(acetoxymethyl ester); GPN, glycyl-1-phenylalanine 2-naphthylamide; PDI, protein-disulfide isomerase. …”

“…Store-operated calcium entry (SOCE), 4 a Ca 2ϩ influx mechanism regulated by the filling state of the intracellular Ca 2ϩ stores, is a major mechanism for Ca 2ϩ influx in non-excitable cells (1). It has long been known that Ca 2ϩ is stored in the endoplasmic reticulum (ER), which is the best-studied agonistreleasable Ca 2ϩ store; however, a number of studies have revealed that Ca 2ϩ is also dynamically accumulated in a number of acidic organelles (2), including secretory granules, lysosomes and lysosome-related organelles, endosomes, and vesicles of the Golgi complex (2)(3)(4)(5)(6).…”

Calcium dynamics in the secretory granules of neuroendocrine cells

“…The sarco-endoplasmic reticulum Ca 2+ -ATPase (SERCA) inhibitor 2,5-diterbutyl benzohydroquinone (BHQ) has been shown to inhibit Ca 2+ uptake into isolated synaptic vesicles [39]. In platelets, the acidic Ca 2+ pool was controlled both by a Ca 2+ /H + exchange and by a BHQ-sensitive SERCA 3 [40,41]. Similarly, loading with Ca 2+ of dense-core insulin storage granules of mouse pancreatic ␤-cells was sensitive to both SERCA inhibitors BHQ and thapsigargin [42].…”