Functional expression of TRPM7 as a Ca<sup>2+</sup>influx pathway in adipocytes

Inoue, Hirokazu; Inazu, Masato; Konishi, Masato; Yokoyama, Utako

doi:10.14814/phy2.14272

Cited by 8 publications

(9 citation statements)

References 64 publications

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“…5 G), indicating a reduction in the total Ca 2+ mobilization due a faster inactivation kinetics after stimulation. Similarly, Inoue et al, [50] showed that naltriben (50 μM) increases a TRPM7-like current and [Ca 2+ ] cyt in adipocytes, that they associated with changes in Ca 2+ clearance. This suggests that naltriben (50-100 μM) may affect clearance mechanisms (PMCAs or Na + /Ca 2+ exchangers).…”

Section: Discussionmentioning

confidence: 79%

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

Bomfim

Costiniti

et al. 2020

Cell Calcium

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Calcium (Ca 2+) release-activated Ca 2+ (CRAC) channels mediated by STIM1/2 and ORAI (ORAI1-3) proteins form the dominant store-operated Ca 2+ entry (SOCE) pathway in a variety of cells. Among these, the enamel-forming cells known as ameloblasts rely on CRAC channel function to enable Ca 2+ influx, which is important for enamel mineralization. This key role of the CRAC channel is supported by human mutations and animal models lacking STIM1 and ORAI1, which results in enamel defects and hypomineralization. A number of recent reports have highlighted the role of the chanzyme TRPM7 (transient receptor potential melastanin 7), a transmembrane protein containing an ion channel permeable to divalent cations (Mg 2+ , Ca 2+), as a modulator of SOCE. This raises the question as to whether TRPM7 should be considered an alternative route for Ca 2+ influx, or if TRPM7 modifies CRAC channel activity in enamel cells. To address these questions, we monitored Ca 2+ influx mediated by SOCE using the pharmacological TRPM7 activator naltriben and the inhibitor NS8593 in rat primary enamel cells and in the murine ameloblast cell line LS8 cells stimulated with thapsigargin. We also measured Ca 2+ dynamics in ORAI1/2-deficient (shOrai1/2) LS8 cells and in cells with siRNA knock-down of Trpm7. We found that primary enamel cells stimulated with theTRPM7 activator potentiated Ca 2+ influx via SOCE compared to control cells. However, blockade of TRPM7 with NS8593 did not decrease the SOCE peak. Furthermore, activation of TRPM7 in shOrai1/2 LS8 cells lacking SOCE failed to elicit Ca 2+ influx, and Trpm7 knock-down had no effect on SOCE. Taken together, our data suggest that TRPM7 is a positive modulator of SOCE potentiating Ca 2+ influx in enamel cells, but its function is fully dependent on the prior activation of the ORAI channels.

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Section: Discussionmentioning

confidence: 79%

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

Bomfim

Costiniti

et al. 2020

Cell Calcium

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“…In adipocytes, cytosolic Ca 2+ regulates insulin responses and the secretion of adipokines. Inoue et al [ 118 ] investigated whether TRPM7 contributes to Ca 2+ influx in freshly isolated white adipocytes and in 3T3-L1 adipocytes differentiated from 3T3-L1 pre-adipocyte cells. The authors used NS8593 together with FTY720, naltriben and siRNA techniques to show that the TRPM7 channel is functionally expressed in adipocytes.…”

Section: Ns8593 As a Tool To Investigate The Function Of Trpm7 Curmentioning

confidence: 99%

“…The authors used NS8593 together with FTY720, naltriben and siRNA techniques to show that the TRPM7 channel is functionally expressed in adipocytes. The authors conclude that TRPM7 plays a role as a Ca 2+ influx pathway in adipocytes [ 118 ].…”

Section: Ns8593 As a Tool To Investigate The Function Of Trpm7 Curmentioning

confidence: 99%

Mapping TRPM7 Function by NS8593

Chubanov¹,

Gudermann²

2020

IJMS

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The transient receptor potential cation channel, subfamily M, member 7 (TRPM7) is a ubiquitously expressed membrane protein, which forms a channel linked to a cytosolic protein kinase. Genetic inactivation of TRPM7 in animal models uncovered the critical role of TRPM7 in early embryonic development, immune responses, and the organismal balance of Zn2+, Mg2+, and Ca2+. TRPM7 emerged as a new therapeutic target because malfunctions of TRPM7 have been associated with anoxic neuronal death, tissue fibrosis, tumour progression, and giant platelet disorder. Recently, several laboratories have identified pharmacological compounds allowing to modulate either channel or kinase activity of TRPM7. Among other small molecules, NS8593 has been defined as a potent negative gating regulator of the TRPM7 channel. Consequently, several groups applied NS8593 to investigate cellular pathways regulated by TRPM7. Here, we summarize the progress in this research area. In particular, two notable milestones have been reached in the assessment of TRPM7 druggability. Firstly, several laboratories demonstrated that NS8593 treatment reliably mirrors prominent phenotypes of cells manipulated by genetic inactivation of TRPM7. Secondly, it has been shown that NS8593 allows us to probe the therapeutic potential of TRPM7 in animal models of human diseases. Collectively, these studies employing NS8593 may serve as a blueprint for the preclinical assessment of TRPM7-targeting drugs.

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“…Because the extracellular Mg 2+ produces voltage-dependent pore block, the inward current is very small in the presence of physiological extracellular divalent cations. Therefore, in our previous studies on overexpressed mouse TRPM7 (mTRPM7) current in HEK293 cells and an endogenous TRPM7 current in mouse white adipocytes, the extracellular divalent cations were eliminated to investigate the effect of oxidative stress that was induced by H 2 O 2 on both the outward and inward currents (Inoue et al, 2014; Inoue et al, 2019). H 2 O 2 was shown to inhibit TRPM7 current in a voltage-independent and an intracellular free Mg 2+ concentration ([Mg 2+ ] i )-dependent manner.…”

Section: Resultsmentioning

confidence: 99%

Zinc-binding motif acts as an oxidative stress sensor to regulate TRPM7 channel activity

Inoue¹,

Murayama

Kobayashi

et al. 2020

Preprint

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TRPM7 channel activity is negatively regulated by intracellular Mg2+. We previously reported that TRPM7 was inhibited by oxidative stress due to an enhancement of the inhibition by intracellular Mg2+. In the present study, we aimed to clarify the precise mechanism underlying the TRPM7 inhibition by oxidative stress induced by hydrogen peroxide (H2O2). Site-directed mutagenesis on full-length TRPM7 revealed that none of the cysteines other than C1809 and C1813 within the zinc-binding motif of the TRPM7 kinase domain was involved in the H2O2-induced TRPM7 inhibition. When C1809 or C1813 was mutated, full-length TRPM7 failed to be expressed on the plasmamembrane. We therefore developed a novel approach in which the full-length TRPM7 is functionally reconstituted by co-expressing the TRPM7 channel domain (M7cd) and the TRPM7 kinase domain (M7kd) as separate individual proteins in HEK293 cells. When M7cd was expressed alone, the current was inhibited by intracellular Mg2+ stronger than in full-length TRPM7. Co-expression of M7cd and M7kd attenuated the current inhibition by intracellular Mg2+, and the current was sensitive to oxidative stress, indicating a successful reconstitution of a full-length TRPM7-like current. A similar current reconstitution was observed when M7cd co-expressed with the kinase inactive mutant M7kd-K1645R. Thus, it was suggested that the kinase activity might not be essential for the reconstitution. Co-expression of M7cd and M7kd carrying a mutation at C1809 or C1813 failed to restore the full-length TRPM7-like current. No reconstitution was also observed with M7kd carrying a mutation at H1750 and H1807, which are involved in the zinc-binding motif formation together with C1809 and C1813. These data suggest that the zinc-binding motif is essential for the intracellular Mg2+-dependent regulation of the TRPM7 channel activity by M7kd, and the cysteines in the zinc-binding motif might play a role in the oxidative stress response of TRPM7.

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Functional expression of TRPM7 as a Ca²⁺influx pathway in adipocytes

Cited by 8 publications

References 64 publications

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

Mapping TRPM7 Function by NS8593

Zinc-binding motif acts as an oxidative stress sensor to regulate TRPM7 channel activity

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Functional expression of TRPM7 as a Ca2+influx pathway in adipocytes

Cited by 8 publications

References 64 publications

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

TRPM7 activation potentiates SOCE in enamel cells but requires ORAI

Mapping TRPM7 Function by NS8593

Zinc-binding motif acts as an oxidative stress sensor to regulate TRPM7 channel activity

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Functional expression of TRPM7 as a Ca²⁺influx pathway in adipocytes