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BK channel agonist represents a potential therapeutic approach for lysosomal storage diseases

View Article: PubMed Central - PubMed

ABSTRACT

Efficient lysosomal Ca2+ release plays an essential role in lysosomal trafficking. We have recently shown that lysosomal big conductance Ca2+-activated potassium (BK) channel forms a physical and functional coupling with the lysosomal Ca2+ release channel Transient Receptor Potential Mucolipin-1 (TRPML1). BK and TRPML1 forms a positive feedback loop to facilitate lysosomal Ca2+ release and subsequent lysosome membrane trafficking. However, it is unclear whether the positive feedback mechanism is common for other lysosomal storage diseases (LSDs) and whether BK channel agonists rescue abnormal lysosomal storage in LSDs. In this study, we assessed the effect of BK agonist, NS1619 and NS11021 in a number of LSDs including NPC1, mild cases of mucolipidosis type IV (ML4) (TRPML1-F408∆), Niemann-Pick type A (NPA) and Fabry disease. We found that TRPML1-mediated Ca2+ release was compromised in these LSDs. BK activation corrected the impaired Ca2+ release in these LSDs and successfully rescued the abnormal lysosomal storage of these diseases by promoting TRPML1-mediated lysosomal exocytosis. Our study suggests that BK channel activation stimulates the TRPML1-BK positive reinforcing loop to correct abnormal lysosomal storage in LSDs. Drugs targeting BK channel represent a potential therapeutic approach for LSDs.

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Activation of BK channels by NS1619.(A) Bath application of NS1619 (15 μM), a BK activator, significantly increased BK currents, but not inwardly rectifying ML1 currents in lysosomes isolated from Cos1 cells expressing BK-GFP. Bath application of 100 μM Ca2+ was employed to maximally activate BK currents. (B) NS1619 (50 μM) markedly increased outwardly rectifying BK currents but not inwardly rectifying ML1 currents in lysosomes isolated from human skin fibroblasts. ML-SA (20 μM) was used to induce TRPML1 currents. (C) Bath NS1619 (50 μM) did not induce outward current in lysosomes isolated from BK knockout human fibroblasts, whereas ML-SA (20 μM) induced significant TRPML1 currents. (Upper) representative BK currents in response to a voltage ramp (from −120 mV to 120 mV, vholding = 0 mV). (Bottom) Normalized currents (normalized to the basal current amplitude under 100 nM Ca2+) measured at 105 mV.
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f2: Activation of BK channels by NS1619.(A) Bath application of NS1619 (15 μM), a BK activator, significantly increased BK currents, but not inwardly rectifying ML1 currents in lysosomes isolated from Cos1 cells expressing BK-GFP. Bath application of 100 μM Ca2+ was employed to maximally activate BK currents. (B) NS1619 (50 μM) markedly increased outwardly rectifying BK currents but not inwardly rectifying ML1 currents in lysosomes isolated from human skin fibroblasts. ML-SA (20 μM) was used to induce TRPML1 currents. (C) Bath NS1619 (50 μM) did not induce outward current in lysosomes isolated from BK knockout human fibroblasts, whereas ML-SA (20 μM) induced significant TRPML1 currents. (Upper) representative BK currents in response to a voltage ramp (from −120 mV to 120 mV, vholding = 0 mV). (Bottom) Normalized currents (normalized to the basal current amplitude under 100 nM Ca2+) measured at 105 mV.

Mentions: The rescue effect of NS1619 on NPC1 cellular phenotypes could be attributed to activating either BK or TRPML1. To tease apart these two possibilities, we directly studied NS1619’s effect on lysosomal BK and TRPML1 channel activity. Under whole-lysosome recording mode, TRPML1 displays strongly inward rectification while BK shows strongly outward rectification7. In Cos1 cells expressing BK-GFP, 15 μM NS1619 significantly increased lysosomal BK currents by ~5 fold (at 105 mV) (Fig. 2A). In wild type human skin fibroblasts, 50 μM NS1619 also significantly increased endogenous lysosomal BK currents by ~4 fold (at 105 mV) (Fig. 2B). In contrast, NS1619 (50 μM) induced little BK current in human fibroblasts with BK deleted using CRISPR-Cas9 gene editing method7 (Fig. 2C), suggesting that NS1619 specifically activates BK to induce the outwardly rectifying currents. On the other hand, NS1619 (50 μM) had no effect on both endogenous (Fig. 2B,C) and heterologous (data not shown) TRPML1 currents, which could be activated by the ML1 agonist, mucolipin synthetic agonist 1 (ML-SA1).


BK channel agonist represents a potential therapeutic approach for lysosomal storage diseases
Activation of BK channels by NS1619.(A) Bath application of NS1619 (15 μM), a BK activator, significantly increased BK currents, but not inwardly rectifying ML1 currents in lysosomes isolated from Cos1 cells expressing BK-GFP. Bath application of 100 μM Ca2+ was employed to maximally activate BK currents. (B) NS1619 (50 μM) markedly increased outwardly rectifying BK currents but not inwardly rectifying ML1 currents in lysosomes isolated from human skin fibroblasts. ML-SA (20 μM) was used to induce TRPML1 currents. (C) Bath NS1619 (50 μM) did not induce outward current in lysosomes isolated from BK knockout human fibroblasts, whereas ML-SA (20 μM) induced significant TRPML1 currents. (Upper) representative BK currents in response to a voltage ramp (from −120 mV to 120 mV, vholding = 0 mV). (Bottom) Normalized currents (normalized to the basal current amplitude under 100 nM Ca2+) measured at 105 mV.
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Related In: Results  -  Collection

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f2: Activation of BK channels by NS1619.(A) Bath application of NS1619 (15 μM), a BK activator, significantly increased BK currents, but not inwardly rectifying ML1 currents in lysosomes isolated from Cos1 cells expressing BK-GFP. Bath application of 100 μM Ca2+ was employed to maximally activate BK currents. (B) NS1619 (50 μM) markedly increased outwardly rectifying BK currents but not inwardly rectifying ML1 currents in lysosomes isolated from human skin fibroblasts. ML-SA (20 μM) was used to induce TRPML1 currents. (C) Bath NS1619 (50 μM) did not induce outward current in lysosomes isolated from BK knockout human fibroblasts, whereas ML-SA (20 μM) induced significant TRPML1 currents. (Upper) representative BK currents in response to a voltage ramp (from −120 mV to 120 mV, vholding = 0 mV). (Bottom) Normalized currents (normalized to the basal current amplitude under 100 nM Ca2+) measured at 105 mV.
Mentions: The rescue effect of NS1619 on NPC1 cellular phenotypes could be attributed to activating either BK or TRPML1. To tease apart these two possibilities, we directly studied NS1619’s effect on lysosomal BK and TRPML1 channel activity. Under whole-lysosome recording mode, TRPML1 displays strongly inward rectification while BK shows strongly outward rectification7. In Cos1 cells expressing BK-GFP, 15 μM NS1619 significantly increased lysosomal BK currents by ~5 fold (at 105 mV) (Fig. 2A). In wild type human skin fibroblasts, 50 μM NS1619 also significantly increased endogenous lysosomal BK currents by ~4 fold (at 105 mV) (Fig. 2B). In contrast, NS1619 (50 μM) induced little BK current in human fibroblasts with BK deleted using CRISPR-Cas9 gene editing method7 (Fig. 2C), suggesting that NS1619 specifically activates BK to induce the outwardly rectifying currents. On the other hand, NS1619 (50 μM) had no effect on both endogenous (Fig. 2B,C) and heterologous (data not shown) TRPML1 currents, which could be activated by the ML1 agonist, mucolipin synthetic agonist 1 (ML-SA1).

View Article: PubMed Central - PubMed

ABSTRACT

Efficient lysosomal Ca2+ release plays an essential role in lysosomal trafficking. We have recently shown that lysosomal big conductance Ca2+-activated potassium (BK) channel forms a physical and functional coupling with the lysosomal Ca2+ release channel Transient Receptor Potential Mucolipin-1 (TRPML1). BK and TRPML1 forms a positive feedback loop to facilitate lysosomal Ca2+ release and subsequent lysosome membrane trafficking. However, it is unclear whether the positive feedback mechanism is common for other lysosomal storage diseases (LSDs) and whether BK channel agonists rescue abnormal lysosomal storage in LSDs. In this study, we assessed the effect of BK agonist, NS1619 and NS11021 in a number of LSDs including NPC1, mild cases of mucolipidosis type IV (ML4) (TRPML1-F408∆), Niemann-Pick type A (NPA) and Fabry disease. We found that TRPML1-mediated Ca2+ release was compromised in these LSDs. BK activation corrected the impaired Ca2+ release in these LSDs and successfully rescued the abnormal lysosomal storage of these diseases by promoting TRPML1-mediated lysosomal exocytosis. Our study suggests that BK channel activation stimulates the TRPML1-BK positive reinforcing loop to correct abnormal lysosomal storage in LSDs. Drugs targeting BK channel represent a potential therapeutic approach for LSDs.

No MeSH data available.


Related in: MedlinePlus