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Store-operated Ca²⁺ entry in primary murine lung fibroblasts is independent of classical transient receptor potential (TRPC) channels and contributes to cell migration.

ABSTRACT: Stromal interaction molecules (STIM1, 2) are acting as sensors for Ca²⁺ in intracellular stores and activate Orai channels at the plasma membrane for store-operated Ca²⁺ entry (SOCE), while classical transient receptor potential (TRPC) channel mediate receptor-operated Ca²⁺ entry (ROCE). Several reports, however, indicate a role for TRPC in SOCE in certain cell types. Here, we analyzed Ca²⁺ influx and cell function in TRPC1/6-deficient (TRPC1/6^-/-) and STIM1/2- deficient (STIM1/2^?pmLF) primary murine lung fibroblasts (pmLF). As expected, SOCE was decreased in STIM1/2- deficient pmLF and ROCE was decreased in TRPC1/6^-/- pmLF compared to control cells. By contrast, SOCE was not significantly different in TRPC1/6^-/- pmLF and ROCE was similar in STIM1/2-deficient pmLF compared to Wt cells. Most interestingly, cell proliferation, migration and nuclear localization of nuclear factor of activated T-cells (NFATc1 and c3) were decreased after ablation of STIM1/2 proteins in pmLF. In conclusion, TRPC1/6 channels are not involved in SOCE and STIM1/2 deficiency resulted in decreased cell proliferation and migration in pmLF.

SUBMITTER: Bendiks L

PROVIDER: S-EPMC7176639 | biostudies-literature | 2020 Apr

REPOSITORIES: biostudies-literature

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Store-operated Ca2+ entry in primary murine lung fibroblasts is independent of classical transient receptor potential (TRPC) channels and contributes to cell migration.

Bendiks Larissa L Geiger Fabienne F Gudermann Thomas T Feske Stefan S Dietrich Alexander A

Scientific reports 20200422 1

Stromal interaction molecules (STIM1, 2) are acting as sensors for Ca2+ in intracellular stores and activate Orai channels at the plasma membrane for store-operated Ca2+ entry (SOCE), while classical transient receptor potential (TRPC) channel mediate receptor-operated Ca2+ entry (ROCE). Several reports, however, indicate a role for TRPC in SOCE in certain cell types. Here, we analyzed Ca2+ influx and cell function in TRPC1/6-deficient (TRPC1/6-/- ...[more]

PMID: 32321939

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Project description:The endoplasmic reticulum (ER) is at the epicenter of astrocyte Ca(2+) signaling. We sought to identify the molecular mechanism underlying store-operated calcium entry that replenishes ER stores in mouse Müller cells. Store depletion, induced through blockade of sequestration transporters in Ca(2+)-free saline, induced synergistic activation of canonical transient receptor potential 1 (TRPC1) and Orai channels. Store-operated TRPC1 channels were identified by their electrophysiological properties, pharmacological blockers, and ablation of the Trpc1 gene. Ca(2+) release-activated currents (ICRAC) were identified by ion permeability, voltage dependence, and sensitivity to selective Orai antagonists Synta66 and GSK7975A. Depletion-evoked calcium influx was initiated at the Müller end-foot and apical process, triggering centrifugal propagation of Ca(2+) waves into the cell body. EM analysis of the end-foot compartment showed high-density ER cisternae that shadow retinal ganglion cell (RGC) somata and axons, protoplasmic astrocytes, vascular endothelial cells, and ER-mitochondrial contacts at the vitreal surface of the end-foot. The mouse retina expresses transcripts encoding both Stim and all known Orai genes; Müller glia predominantly express stromal interacting molecule 1 (STIM1), whereas STIM2 is mainly confined to the outer plexiform and RGC layers. Elimination of TRPC1 facilitated Müller gliosis induced by the elevation of intraocular pressure, suggesting that TRPC channels might play a neuroprotective role during mechanical stress. By characterizing the properties of store-operated signaling pathways in Müller cells, these studies expand the current knowledge about the functional roles these cells play in retinal physiology and pathology while also providing further evidence for the complexity of calcium signaling mechanisms in CNS astroglia.Significance statementStore-operated Ca(2+) signaling represents a major signaling pathway and source of cytosolic Ca(2+) in astrocytes. Here, we show that the store-operated response in Müller cells, radial glia that perform key structural, signaling, osmoregulatory, and mechanosensory functions within the retina, is mediated through synergistic activation of transient receptor potential and Orai channels. The end-foot disproportionately expresses the depletion sensor stromal interacting molecule 1, which contains an extraordinarily high density of endoplasmic reticulum cisternae that shadow neuronal, astrocytic, vascular, and axonal structures; interface with mitochondria; but also originate store-operated Ca(2+) entry-induced transcellular Ca(2+) waves that propagate glial excitation into the proximal retina. These results identify a molecular mechanism that underlies complex interactions between the plasma membrane and calcium stores, and contributes to astroglial function, regulation, and response to mechanical stress.

Dataset Information

Store-operated Ca²⁺ entry in primary murine lung fibroblasts is independent of classical transient receptor potential (TRPC) channels and contributes to cell migration.

Publications

Store-operated Ca<sup>2+</sup> entry in primary murine lung fibroblasts is independent of classical transient receptor potential (TRPC) channels and contributes to cell migration.

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