Nanoscale coupling of junctophilin-2 and ryanodine receptors regulates vascular smooth muscle cell contractility

Pritchard, Harry A. T.; Griffin, Caoimhin S.; Yamasaki, Evan; Thakore, Pratish; Lane, Conor; Greenstein, Adam; Earley, Scott

doi:10.1073/pnas.1911304116

Cited by 39 publications

(58 citation statements)

References 40 publications

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“…To investigate this hypothesis, we utilized immunofluorescence labeling in conjunction with ground state depletion followed by individual molecule return (GSDIM) superresolution microscopy (34)(35)(36) in native BSMCs to examine the subcellular nanoscale colocalization of TRPML1 channel clusters with RyR2s and the LEL membrane-bound protein, Lamp-1 (lysosomal-associated membrane protein 1). Using DNA origamibased nanorulers, we have previously shown that the lateral resolution of our GSDIM system is 20 to 40 nm (37)(38)(39).…”

Section: Nanoscale Localization Of Trpml1-positive Lels To Ryr2 Clustmentioning

confidence: 96%

The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

Griffin

Alvarado

Yamasaki

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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TRPML1 (transient receptor potential mucolipin 1) is a Ca2+-permeable, nonselective cation channel that is predominantly localized to the membranes of late endosomes and lysosomes (LELs). Intracellular release of Ca2+through TRPML1 is thought to be pivotal for maintenance of intravesicular acidic pH as well as the maturation, fusion, and trafficking of LELs. Interestingly, genetic ablation of TRPML1 in mice (Mcoln1−/−) induces a hyperdistended/hypertrophic bladder phenotype. Here, we investigated this phenomenon further by exploring an unconventional role for TRPML1 channels in the regulation of Ca2+-signaling activity and contractility in bladder and urethral smooth muscle cells (SMCs). Four-dimensional (4D) lattice light-sheet live-cell imaging showed that the majority of LELs in freshly isolated bladder SMCs were essentially immobile. Superresolution microscopy revealed distinct nanoscale colocalization of LEL-expressing TRPML1 channels with ryanodine type 2 receptors (RyR2) in bladder SMCs. Spontaneous intracellular release of Ca2+from the sarcoplasmic reticulum (SR) through RyR2 generates localized elevations of Ca2+(“Ca2+sparks”) that activate plasmalemmal large-conductance Ca2+-activated K+(BK) channels, a critical negative feedback mechanism that regulates smooth muscle contractility. This mechanism was impaired inMcoln1−/−mice, which showed diminished spontaneous Ca2+sparks and BK channel activity in bladder and urethra SMCs. Additionally, ex vivo contractility experiments showed that loss of Ca2+spark–BK channel signaling inMcoln1−/−mice rendered both bladder and urethra smooth muscle hypercontractile. Voiding activity analyses revealed bladder overactivity inMcoln1−/−mice. We conclude that TRPML1 is critically important for Ca2+spark signaling, and thus regulation of contractility and function, in lower urinary tract SMCs.

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Section: Nanoscale Localization Of Trpml1-positive Lels To Ryr2 Clustmentioning

confidence: 96%

The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

Griffin

Alvarado

Yamasaki

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…In further studies, single SMCs from cerebral arteries isolated from control and Stim1-smKO mice were enzymatically dispersed, immunolabeled with an anti-STIM1 primary antibody, and imaged using a GSDIM (ground state depletion followed by individual molecule return) superresolution microscopy system, which we previously showed using DNA-origami-based nanorulers has a lateral resolution of 20-40 nm (49)(50)(51). VSMCs from control mice exhibited punctate STIM1 protein clusters (Figure 1B).…”

Section: Stim1-smko Mice Lack Stim1 Protein Expression In Vsmcsmentioning

confidence: 98%

Peripheral Coupling Sites Formed by STIM1 Govern the Contractility of Vascular Smooth Muscle Cells

Krishnan

Ali

et al. 2021

Preprint

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Peripheral coupling between the sarcoplasmic reticulum (SR) and plasma membrane (PM) forms signaling complexes that regulate the membrane potential and contractility of vascular smooth muscle cells (VSMCs), although the mechanisms responsible for these membrane interactions are poorly understood. In many cells, STIM1 (stromal interaction molecule 1), a single transmembrane-domain protein that resides in the endoplasmic reticulum (ER), transiently moves to ER-PM junctions in response to depletion of ER Ca2+ stores and initiates store-operated Ca2+ entry (SOCE). Fully differentiated VSMCs express STIM1 but exhibit only marginal SOCE activity. We hypothesized that STIM1 is constitutively active in contractile VSMCs and maintains peripheral coupling. In support of this concept, we found that the number and size of SR-PM interacting sites were decreased and SR-dependent Ca2+ signaling processes were disrupted in freshly isolated cerebral artery SMCs from tamoxifen-inducible, SMC specific STIM1-knockout (Stim1-smKO) mice. VSMCs from Stim1-smKO mice also exhibited a reduction in nanoscale colocalization between Ca2+-release sites on the SR and Ca2+-activated ion channels on the PM, accompanied by diminished channel activity. Stim1-smKO mice were hypotensive and resistance arteries isolated from them displayed blunted contractility. These data suggest that STIM1 – independent of SR Ca2+ store depletion – is critically important for stable peripheral coupling in contractile VSMCs.

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“…Importantly, others have confirmed the regional distribution of SERCA and RyRs not only in pulmonary arterial smooth muscles [ 137 ], but in airway smooth muscles [ 138 ], which may provide for the high degree of variability in kinetics of spontaneous SR calcium release in myocytes from the ileum [ 189 ] and hepatic portal vein [ 126 ]. It is also notable that previous studies have identified RyR1, RyR2 and RyR3 expression in cerebral arterial myocytes [ 139 ], which must at some stage inform current models on calcium signalling in these cells too, which for some reason rely solely on a consideration of RyR2 clusters, whether one considers PM-SR nanojunctions [ 230 , 231 ] or more recent “comparative” assessment of the role of lysosome-SR nanojunctions in this cell type [ 129 ].…”

Section: Reviewing the Situation: Got To Pick A Pocket Or Two Boymentioning

confidence: 99%

On a Magical Mystery Tour with 8-Bromo-Cyclic ADP-Ribose: From All-or-None Block to Nanojunctions and the Cell-Wide Web

Evans

2020

Molecules

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A plethora of cellular functions are controlled by calcium signals, that are greatly coordinated by calcium release from intracellular stores, the principal component of which is the sarco/endooplasmic reticulum (S/ER). In 1997 it was generally accepted that activation of various G protein-coupled receptors facilitated inositol-1,4,5-trisphosphate (IP3) production, activation of IP3 receptors and thus calcium release from S/ER. Adding to this, it was evident that S/ER resident ryanodine receptors (RyRs) could support two opposing cellular functions by delivering either highly localised calcium signals, such as calcium sparks, or by carrying propagating, global calcium waves. Coincidentally, it was reported that RyRs in mammalian cardiac myocytes might be regulated by a novel calcium mobilising messenger, cyclic adenosine diphosphate-ribose (cADPR), that had recently been discovered by HC Lee in sea urchin eggs. A reputedly selective and competitive cADPR antagonist, 8‑bromo‑cADPR, had been developed and was made available to us. We used 8‑bromo‑cADPR to further explore our observation that S/ER calcium release via RyRs could mediate two opposing functions, namely pulmonary artery dilation and constriction, in a manner seemingly independent of IP3Rs or calcium influx pathways. Importantly, the work of others had shown that, unlike skeletal and cardiac muscles, smooth muscles might express all three RyR subtypes. If this were the case in our experimental system and cADPR played a role, then 8‑bromo‑cADPR would surely block one of the opposing RyR-dependent functions identified, or the other, but certainly not both. The latter seemingly implausible scenario was confirmed. How could this be, do cells hold multiple, segregated SR stores that incorporate different RyR subtypes in receipt of spatially segregated signals carried by cADPR? The pharmacological profile of 8‑bromo‑cADPR action supported not only this, but also indicated that intracellular calcium signals were delivered across intracellular junctions formed by the S/ER. Not just one, at least two. This article retraces the steps along this journey, from the curious pharmacological profile of 8‑bromo‑cADPR to the discovery of the cell-wide web, a diverse network of cytoplasmic nanocourses demarcated by S/ER nanojunctions, which direct site-specific calcium flux and may thus coordinate the full panoply of cellular processes.

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Nanoscale coupling of junctophilin-2 and ryanodine receptors regulates vascular smooth muscle cell contractility

Cited by 39 publications

References 40 publications

The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

Peripheral Coupling Sites Formed by STIM1 Govern the Contractility of Vascular Smooth Muscle Cells

On a Magical Mystery Tour with 8-Bromo-Cyclic ADP-Ribose: From All-or-None Block to Nanojunctions and the Cell-Wide Web

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Nanoscale coupling of junctophilin-2 and ryanodine receptors regulates vascular smooth muscle cell contractility

Cited by 39 publications

References 40 publications

The intracellular Ca 2+ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

The intracellular Ca 2+ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

Peripheral Coupling Sites Formed by STIM1 Govern the Contractility of Vascular Smooth Muscle Cells

On a Magical Mystery Tour with 8-Bromo-Cyclic ADP-Ribose: From All-or-None Block to Nanojunctions and the Cell-Wide Web

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The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility

The intracellular Ca ²⁺ release channel TRPML1 regulates lower urinary tract smooth muscle contractility