Sphingosine-1-Phosphate Induces ATP Release via Volume-Regulated Anion Channels in Breast Cell Lines

Abstract

High interstitial level of ATP and its lysate adenosine in the cancer microenvironment are considered a halo mark of cancer. Adenosine acts as a strong immune suppressor. However, the source of ATP release is unclear. We clarified the release of ATP via volume-regulated anion channels (VRACs) in breast cell lines using an ATP luminescence imaging system. We detected a slowly rising diffuse pattern of ATP release that was only observed in undifferentiated cells, not in differentiated primary cultured cells. This was confirmed by suppression with DCPIB, a blocker of VRACs, and shRNA for LRRC8A, an indispensable subunit of VRACs. We herein demonstrated that the inflammatory mediator sphingosine-1-phosphate (S1P), which exists abundantly in the cancer microenvironment, induced a diffuse pattern of ATP release isovolumetrically. The response was dose-dependent and suppressed by the knock-down of LRRC8A. It was also suppressed by blockers of S1P receptor 1 and 2 (W146 and JTE013, respectively). RTqPCR demonstrated the prominent presence of S1PR1 and S1PR2 mRNAs. We discussed the roles of S1P-induced ATP release in the cancer microenvironment.

Properties, Structures, and Physiological Roles of Three Types of Anion Channels Molecularly Identified in the 2010’s

“…Actually, a large number of studies demonstrated VSOR/VRAC-mediated glutamate release and Maxi-Cl-mediated release of glutamate and ATP (see Reviews: Okada et al, 2018 , 2021a ). VSOR/VRAC-mediated ATP release was observed in some but not all types of cells ( Hisadome et al, 2002 ; Dutta et al, 2004 ; Dunn et al, 2020 ; Furuya et al, 2021 ). Such cell type-dependent ATP release capability of VSOR/VRAC may be explained by the fact that ATP conductivity via VSOR/VRAC depends on the subunit composition of the LRRC8 hexamer ( Gaitán-Peñas et al, 2016 ; Syeda et al, 2016 ).…”

Physiology of the volume-sensitive/regulatory anion channel VSOR/VRAC: part 2: its activation mechanisms and essential roles in organic signal release

“…ATP release induced by mechanical stimulation was found to be sensitive to VSOR/VRAC blockers but insensitive to Lrrc8a knockdown in rat astrocytes [72]. In contrast, gene knockdown of LRRC8A was shown to suppress ATP release induced by hypoosmotic stimulation in HEK293 cells [73], HeLa cells [74], and mouse microglial BV-2 cells [74] and that induced by application of S1P in mouse microglial BV-2 cells [74,75] and in human breast cancer MCF7 and MDA-MB-231 cells [76]. Collectively, it appears that VSOR/VRAC can mediate ATP release in many, but not all, cell types.…”