P2X7-mediated ATP secretion is accompanied by depletion of cytosolic ATP
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P2X7-mediated ATP secretion is accompanied by depletion of cytosolic ATP. / Johnsen, Bjarne; Kaschubowski, Klaus Eric; Nader, Sorush; Schneider, Enja; Nicola, Jan-Andrei; Fliegert, Ralf; Wolf, Insa M A; Guse, Andreas H; Nikolaev, Viacheslav O; Koch-Nolte, Friedrich; Haag, Friedrich.
in: PURINERG SIGNAL, Jahrgang 15, Nr. 2, 06.2019, S. 155-166.Publikationen: SCORING: Beitrag in Fachzeitschrift/Zeitung › SCORING: Zeitschriftenaufsatz › Forschung › Begutachtung
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T1 - P2X7-mediated ATP secretion is accompanied by depletion of cytosolic ATP
AU - Johnsen, Bjarne
AU - Kaschubowski, Klaus Eric
AU - Nader, Sorush
AU - Schneider, Enja
AU - Nicola, Jan-Andrei
AU - Fliegert, Ralf
AU - Wolf, Insa M A
AU - Guse, Andreas H
AU - Nikolaev, Viacheslav O
AU - Koch-Nolte, Friedrich
AU - Haag, Friedrich
PY - 2019/6
Y1 - 2019/6
N2 - ATP and its metabolites are important extracellular signal transmitters acting on purinergic P2 and P1 receptors. Most cells can actively secrete ATP in response to a variety of external stimuli such as gating of the P2X7 receptor. We used Yac-1 murine lymphoma cells to study P2X7-mediated ATP release. These cells co-express P2X7 and ADP-ribosyltransferase ARTC2, permitting gating of P2X7 by NAD+-dependent ADP-ribosylation without the need to add exogenous ATP. Yac-1 cells released ATP into the extracellular space within minutes after stimulation with NAD+. This was blocked by pre-incubation with the inhibitory P2X7-specific nanobody 13A7. Gating of P2X7 for 3 h significantly decreased intracellular ATP levels in living cells, but these had returned to normal by 20 h. P2X7-mediated ATP release was dependent on a rise in cytosolic calcium and the depletion of intracellular potassium, but was not blocked by inhibitors of pannexins or connexins. We used genetically encoded FRET-based ATP sensors targeted to the cytosol to image P2X7-mediated changes in the distribution of ATP in 3T3 fibroblasts co-expressing P2X7 and ARTC2 and in Yac-1 cells. In response to NAD+, we observed a marked depletion of ATP in the cytosol. This study demonstrates the potential of ATP sensors as tools to study regulated ATP release by other cell types under other conditions.
AB - ATP and its metabolites are important extracellular signal transmitters acting on purinergic P2 and P1 receptors. Most cells can actively secrete ATP in response to a variety of external stimuli such as gating of the P2X7 receptor. We used Yac-1 murine lymphoma cells to study P2X7-mediated ATP release. These cells co-express P2X7 and ADP-ribosyltransferase ARTC2, permitting gating of P2X7 by NAD+-dependent ADP-ribosylation without the need to add exogenous ATP. Yac-1 cells released ATP into the extracellular space within minutes after stimulation with NAD+. This was blocked by pre-incubation with the inhibitory P2X7-specific nanobody 13A7. Gating of P2X7 for 3 h significantly decreased intracellular ATP levels in living cells, but these had returned to normal by 20 h. P2X7-mediated ATP release was dependent on a rise in cytosolic calcium and the depletion of intracellular potassium, but was not blocked by inhibitors of pannexins or connexins. We used genetically encoded FRET-based ATP sensors targeted to the cytosol to image P2X7-mediated changes in the distribution of ATP in 3T3 fibroblasts co-expressing P2X7 and ARTC2 and in Yac-1 cells. In response to NAD+, we observed a marked depletion of ATP in the cytosol. This study demonstrates the potential of ATP sensors as tools to study regulated ATP release by other cell types under other conditions.
U2 - 10.1007/s11302-019-09654-5
DO - 10.1007/s11302-019-09654-5
M3 - SCORING: Journal article
C2 - 31016551
VL - 15
SP - 155
EP - 166
JO - PURINERG SIGNAL
JF - PURINERG SIGNAL
SN - 1573-9538
IS - 2
ER -