Increased Red Cell KCNN4 Activity in Sporadic Hereditary Xerocytosis Associated With Enhanced Single Channel Pressure Sensitivity of PIEZO1 Mutant V598M

Gnanasambandam, Radhakrishnan; Rivera, Alicia; Vandorpe, David H.; Shmukler, Boris E.; Brugnara, Carlo; Snyder, Lois; Andolfo, Immacolata; Iolascon, Achille; Silveira, Paulo Augusto Achucarro; Hamerschlak, Nelson; Gottlieb, Philip A.; Alper, Seth L.

doi:10.1097/hs9.0000000000000055

Cited by 12 publications

(12 citation statements)

References 10 publications

(25 reference statements)

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“…Although HEK293 cells constitute a powerful expression system extensively used to study stretch-activated channels (and channels in general), these above considerations together with the results presented here question the relevance of using HEK293 cells to study the gating of mechano-sensitive channel from RBC. Of note, in a previous study, the pressure sensitivity of V598M PIEZO1 mutant expressed in HEK293 cells was shown to slightly increase compared to WT (Gnanasambandam et al, 2018). This discrepancy with our present results would suggest an even more complex regulation of PIEZO1 activity depending on the different experimental conditions among diverse laboratories, such as (i) different cellular clones, (ii) stages of cellular cycle, (iii) cellular senescence, and (iv) number of replicates.…”

Section: Discussioncontrasting

confidence: 99%

“…Hence, these PIEZO1 mutants are likely to induce a constitutive Na + , K + , and Ca 2+ leak in RBC. Moreover, in previously published patch-clamp experiments on RBC carrying the V598M PIEZO1 mutant (in whole cell and cellattached configurations), a constitutive cation leak, increased conductance and increased spontaneous activity were observed (Rapetti-Mauss et al, 2017;Gnanasambandam et al, 2018). To characterize mutant PIEZO1 features, they were expressed in HEK293 cells.…”

Section: Discussionmentioning

confidence: 99%

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Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells

et al. 2021

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Hereditary Xerocytosis, a rare hemolytic anemia, is due to gain of function mutations in PIEZO1, a non-selective cation channel activated by mechanical stress. How these PIEZO1 mutations impair channel function and alter red blood cell (RBC) physiology, is not completely understood. Here, we report the characterization of mutations in the N-terminal part of the protein (V598M, F681S and the double mutation G782S/R808Q), a part of the channel that was subject of many investigations to decipher its role in channel gating. Our data show that the electrophysiological features of these PIEZO1 mutants expressed in HEK293T cells are different from previously characterized PIEZO1 mutations that are located in the pore or at the C-terminal extracellular domain of the protein. Although RBC with PIEZO1 mutations showed a dehydrated phenotype, the activity of V598M, F681S or R808Q in response to stretch was not significantly different from the WT channels. In contrast, the G782S mutant showed larger currents compared to the WT PIEZO1. Interestingly, basal activity of all the mutated channels was not significantly altered at the opposite of what was expected according to the decreased water and cation contents of resting RBC. In addition, the features of mutant PIEZO1 expressed in HEK293 cells do not always correlate with the observation in RBC where PIEZO1 mutations induced a cation leak associated with an increased conductance. Our work emphasizes the role of the membrane environment in PIEZO1 activity and the need to characterize RBC permeability to assess pathogenicity to PIEZO1 mutants associated with erythrocyte diseases.

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Section: Discussioncontrasting

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells

et al. 2021

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“…The V598M mutation has also previously been reported in an unrelated HX case with a typical phenotype associating hemolysis, iron overload, and neonatal ascites, with functional studies revealing a lower PIEZO1 activation threshold and increased K + influx sensitive to Gardos channel inhibition. 53 These data suggest that mutations involving amino-acid 598 may be associated with a higher risk of PE development.…”

Section: Piezo1-hx and Perinatal Edemamentioning

confidence: 89%

“…Thus, in DHS1, RBC dehydration is largely due to inappropriate activation of the Gardos channel (see Section 2 above) and can be reverted in vitro using the selective Gardos inhibitor Senicapoc (Figure 2(B)). 31,53 Senicapoc may thus represent an attractive treatment option for symptomatic HX, especially as the molecule has been shown to be safe in clinical trials for the treatment of sickle‐cell disease, although its efficacy on dehydration and hemolysis parameters was not shown to reduce the frequency of vaso‐occlusive crises 54,55 . Strikingly, Gardos channelopathies and DHS1 are not entirely comparable, differing in the severity of hematological phenotype, occurrence of perinatal edema, thrombotic risk after splenectomy, and perhaps iron overload risk 28,50,56 .…”

Section: Erythroid Cell Defects In Piezo1‐hxmentioning

confidence: 99%

Recent advances in the pathophysiology of PIEZO1‐related hereditary xerocytosis

et al. 2021

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Hereditary xerocytosis is a rare red blood cell disease related to gain-of-function mutations in the FAM38A gene, encoding PIEZO1, in 90% of cases; PIEZO1 is a broadly expressed mechano-transducer that plays a major role in many cell systems and tissues that respond to mechanical stress. In erythrocytes, PIEZO1 adapts the intracellular ionic content and cell hydration status to the mechanical constraints induced by the environment. Until recently, the pathophysiology of hereditary xerocytosis was mainly believed to be based on the "PIEZO1-Gardos channel axis" in erythrocytes, according to which PIEZO1-activating mutations induce a calcium influx that secondarily activates the Gardos channel, leading to potassium and water efflux and subsequently to red blood cell dehydration. However, recent studies have demonstrated additional roles for PIEZO1 during early erythropoiesis and reticulocyte maturation, as well as roles in other tissues and cells such as lymphatic vessels, hepatocytes, macrophages and platelets that may affect the pathophysiology of the disease. These findings, presented and discussed in this review, broaden our understanding of hereditary xerocytosis beyond that of primarily being a red blood cell disease and identify potential therapeutic targets. | INTRODUCTIONThe PIEZO proteins are a family of mechano-transducers first described in a neuron derived cell line in 2010. 1 The two members are PIEZO1, encoded by FAM38A on chromosome 16, and PIEZO2, encoded by FAM38B on chromosome 18. They are expressed in many tissues. 1,2 The structure of PIEZO1 has been recently elucidated. A first description of the murine protein revealed its three-dimensional structure, consisting of a three-bladed homotrimeric transmembrane helix completed by an extracellular cap. An intra-cytoplasmic domain, split into three parts, called "beams," extends from the transmembrane domain. The C-terminal end of each monomer has an extracellular domain (ECD), which forms the cap, and an intracellular domain (CTD), which appears to be connected to the paddles by the beams and closes the inner ion pore (Figure 1). The assembly provides a hydrophilic central transmembrane duct bordered by six helices that allows the passage of ions. Thus PIEZO1 constitutes a mechanosensitive ion channel due to the flexibility of the three blades. 1 Deformation of the plasma membrane subsequent to mechanical stimulation induces rotation of the PIEZO1 blades, transmitting such deformation to the ECD and CTD domains, allowing the ion channel to open using a lever-like mechanism. 2 When a compressive force is applied parallel to the axis of the ion channel, it induces lateral membrane tension, resulting in flattening and widening of PIEZO1, which causes the pore to open and allows the passage of ions. Such deformations are reversible, and PIEZO1 returns rapidly to its original conformation. [3][4][5] After opening, PIEZO1 acts as a passive channel for mono/divalent cations depending on their extracellular-intracellular Nicolas Jankovsky and Alexis Caulier...

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“…Mutated KCNN4 Channels have alterations of kinetic of open‐closure of the channel similar to PIEZO1 mutants (Rapetti‐Mauss et al , ; Rivera et al , ). Recent studies demonstrate that chronic KCNN4‐driven red cell dehydration and intracellular cation imbalance (also caused by PIEZO1 mutations) can lead to functional inactivation of KCNN4 activity in DHS RBCs (Gnanasambandam et al , ; Rivera et al , ).…”

Section: Red Blood Cell Membrane Disorders: Classification and Pathogmentioning

confidence: 99%

Advances in understanding the pathogenesis of red cell membrane disorders

2019

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Summary Hereditary erythrocyte membrane disorders are caused by mutations in genes encoding various transmembrane or cytoskeletal proteins of red blood cells. The main consequences of these genetic alterations are decreased cell deformability and shortened erythrocyte survival. Red blood cell membrane defects encompass a heterogeneous group of haemolytic anaemias caused by either (i) altered membrane structural organisation (hereditary spherocytosis, hereditary elliptocytosis, hereditary pyropoikilocytosis and Southeast Asian ovalocytosis) or (ii) altered membrane transport function (overhydrated hereditary stomatocytosis, dehydrated hereditary stomatocytosis or xerocytosis, familial pseudohyperkalaemia and cryohydrocytosis). Herein we provide a comprehensive review of the recent literature on the molecular genetics of erythrocyte membrane defects and their reported clinical consequences. We also describe the effect of low‐expression genetic variants on the high inter‐ and intra‐familial phenotype variability of erythrocyte structural defects.

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Increased Red Cell KCNN4 Activity in Sporadic Hereditary Xerocytosis Associated With Enhanced Single Channel Pressure Sensitivity of PIEZO1 Mutant V598M

Abstract: Supplemental Digital Content is available in the text

Cited by 12 publications

References 10 publications

Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells

Hereditary Xerocytosis: Differential Behavior of PIEZO1 Mutations in the N-Terminal Extracellular Domain Between Red Blood Cells and HEK Cells

Recent advances in the pathophysiology of PIEZO1‐related hereditary xerocytosis

Advances in understanding the pathogenesis of red cell membrane disorders

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