These findings establish glycosylation-mechanics coupling as a determinant of neuronal force sensing and suggest that, by facilitating Piezo1 recruitment, defective glycosylation may increase cortical vulnerability to mechanical stress, potentially contributing to head trauma-triggered neurological episodes in phosphomannomutase 2 deficiency (PMM2-CDG).
Abstract
Piezo1 is a mechanically activated cation channel whose N-linked glycans support protein maturation and plasma membrane trafficking, but their contribution to mechanical gating is unknown. We asked whether hypoglycosylation alters Piezo1 mechanosensitivity and cortical neuronal mechanotransduction, with potential relevance to neurological manifestations of congenital disorders of glycosylation (CDG). Human Piezo1 was studied in HEK293 cells after mutation of two conserved cap-domain N-glycosylation sites or inhibition of N-glycan maturation with swainsonine or kifunensine. Mechanically activated currents were recorded by cell-attached patch-clamp during incremental negative-pressure pulses, whereas Ca2+ responses were measured during uniaxial stretch. Piezo1 abundance, synaptic localisation and stretch-evoked Ca2+ signals were also examined in primary mouse cortical neurons. On poly-L-lysine, N2293Q or N2330Q shifted the pressure-response relationship towards lower activating pressures without changing maximal current or inactivation kinetics. This effect was absent on collagen. Swainsonine and kifunensine reduced mature Piezo1 glycosylation and lowered the mechanical activation threshold. Hypoglycosylation enhanced Ca2+ entry during submaximal stretch in HEK293 cells. In cortical neurons, inhibition of glycan maturation increased somatic Piezo1 immunoreactivity without changing its association with synaptic markers, and potentiated Ca2+ responses to both the Piezo1 activator Yoda1 and submaximal stretch. Thus, mature N-glycans and the extracellular adhesive environment jointly set Piezo1’s mechanical activation threshold rather than merely regulating biosynthesis and trafficking. These findings establish glycosylation-mechanics coupling as a determinant of neuronal force sensing and suggest that, by facilitating Piezo1 recruitment, defective glycosylation may increase cortical vulnerability to mechanical stress, potentially contributing to head trauma-triggered neurological episodes in phosphomannomutase 2 deficiency (PMM2-CDG). Key points Piezo1 channels convert mechanical forces into electrical and calcium signals. N-linked glycans support channel trafficking to the plasma membrane, but whether they tune the force needed for Piezo1 activation was unknown. Mutating either of two conserved N-glycosylation sites in Piezo1 cap domain, or pharmacologically restricting N-glycan maturation, lowered channel’s mechanical activation threshold without changing maximal current or inactivation. This sensitisation depended on the adhesive substrate (occurred on poly-L-lysine but not collagen), and was most evident during submaximal stretch, showing that glycosylation and the extracellular mechanical environment jointly determine Piezo1 force sensing. In mouse cortical neurons, impaired N-glycan maturation increased somatic Piezo1 abundance and enhanced Ca2+ responses to its chemical activator Yoda1 and stretch, without changing synaptic localisation. By allowing weak mechanical inputs to recruit Piezo1 more effectively, defective glycosylation may increase cortical responses to mechanical stress and help explain susceptibility to head trauma-triggered neurological episodes in phosphomannomutase 2 deficiency (PMM2-CDG).
By explaining how a single amino acid change produces a hypomorphic PIEZO2 allele, the findings broaden the clinical spectrum of PIEZO2 disorders and offer structural insight into mechanotransduction.
Alec R. Nickolls, Eric M. Mulhall, Daniel J. Orlin et al.· Neuron· 0 citations
Mechanosensitive Piezo1 and Piezo2 channels convert mechanical forces into intracellular signals, playing essential roles in both physiological homeostasis and disease pathogenesis. This review synthesizes current evidence on their involvement in major neurological disorders, including stroke, Alzheimer's disease, traumatic brain injury, and glioma. Piezo1 primarily contributes to neuroinflammation, blood-brain barrier disruption, and tumor mechanosignaling, whereas Piezo2 dysfunction leads to sensory deficits, mechanical allodynia, and impaired proprioception. Notably, the functional consequences of Piezo activation are context-dependent: Piezo1 exacerbates ischemic brain damage but promotes amyloid-β clearance in Alzheimer's disease. Pharmacological modulators such as GsMTx4 and Yoda1 show promise in preclinical models, yet challenges remain regarding subtype selectivity and blood-brain barrier penetration. Targeting Piezo channels represents a promising therapeutic frontier, but success will require precise, cell-specific modulation based on disease stage and molecular context. This review highlights key pathogenic mechanisms, evaluates current pharmacological strategies, and outlines future directions for translating Piezo-targeted interventions into clinical practice.
Lu Zhao, Chao Wang, Yu-Dan Zhu et al.· Current opinion in pharmacol...· 0 citations
AIMS
Mitral regurgitation (MR) pathophysiology involves increased serotonin (5-HT) receptor (HTR) expression and signaling, together with reduced 5-HT transporter (SLC6A4) expression in mitral valve interstitial cells (MVIC). The mechanosensitive calcium channels, PIEZO1 and PIEZO2, regulate cellular responses to a variety of mechanical conditions; however, PIEZO1 and PIEZO2 have not been studied in MR. We investigated the hypothesis that PIEZO1 and PIEZO2 mediate, through 5-HT mechanisms both homeostasis in normal mitral valves (nMV) and in the pathophysiology of MR.
METHODS AND RESULTS
Immunofluorescence and Western blots demonstrated the presence of PIEZO1 and PIEZO2 in samples of explanted human nMV and MR. MVIC derived from nMV and MR leaflets also expressed PIEZO1 and PIEZO2. Single-cell RNA sequencing (scRNA-seq) analyses of nMV and MR leaflet samples from 9 MR and 5 nMV cases identified 7 different cell types: Endothelial cells, smooth muscle cells, T-cells, macrophages, and three distinct MVIC phenotypes, VIC1, VIC2, and VIC3. MVIC together comprised 97% of all valve cells, irrespective of valve pathology. In addition, scRNA-seq of nMV and MR leaflets demonstrated MVIC PIEZO1 and PIEZO2 expression, together with HTR2B and SLC6A4. MVIC cultures, with no added 5-HT, demonstrated that PIEZO1 activation with its agonist, Yoda1, increased MVIC Ca2+ uptake. This was inhibited by PIEZO1-siRNA, and Dooku1, a Yoda1 antagonist, but not PIEZO2-siRNA or noncoding-RNA. Additional MVIC studies revealed that in static MVIC cultures, PIEZO1 activation with Yoda1 downregulated SLC6A4, upregulated HTR2B, and increased 5-HT levels and protein synthesis. Cyclic tension MVIC bioreactor studies showed that either PIEZO1-siRNA or PIEZO2-siRNA mitigated tension induced SLC6A4 downregulation and increased collagen synthesis; only PIEZO2-siRNA mitigated both static and cyclic tension-induced MVIC 5-HT production.
CONCLUSIONS
Our findings have identified a novel link between mechano-transduction, 5-HT production and receptor signaling, and collagen synthesis, suggesting PIEZO1, PIEZO2, and downstream 5-HT pathways as potential therapeutic targets for mitigating MR progression.
Stanley J. Stachelek, Emily J. Hauber, Xiao-Qiu Yang et al.· Cardiovascular Research· 0 citations
Every known life form senses and reacts to mechanical forces. These mechanical stimuli can be converted into electrical signals by mechanically gated ion channels, a transduction cascade pivotal to numerous physiological functions including touch, hearing, mechanical pain, circulation, gastrointestinal function, and mechanical loading in various tissues. Despite continuous efforts, numerous mechanically gated ion channels with the mechanotransduction process underlying these physiological functions remain unidentified. Here, we focused on the transmembrane channel-like (TMC) protein family expressed in the cultured cells to identify those with potential mechanosensitive activity. Remarkably, in contrast to human TMC1/2 (HsTMC1/2), human TMC3-8 (HsTMC3-8) proteins are localized to the plasma membrane when heterologously expressed in the cultured cells. Further experiments revealed that mechanical poking stimuli can effectively activate HsTMC3-8. In addition, HsTMC3-8 induced stretch-activated currents and elicited well-resolved single-channel activities in response to negative pressure stimulation. The mutants near the putative pore region altered reversal potentials (Erev) of HsTMC3-8, suggesting that TMC3-8 are likely pore-forming subunits of ion channels. In summary, we proposed that TMC proteins are the largest mammalian mechanically gated ion channel family.
Songdi Fu, Jianying Dong, Xing Luo et al.· bioRxiv· 0 citations
Immune cells operate within a dynamic mechanical environment. Shear stress, matrix stiffness, membrane tension, and cellular traction continuously shape their fate and function. Mechanosensitive ion channels (MSICs) are the convergent molecular transducers of these forces. Five families dominate immune mechanotransduction: Piezo, TRPV4, K2P/TREK, TMEM63/OSCA, and ENaC. Each converts mechanical input into Ca2+ and K+ fluxes that drive the transcriptional and effector programs governing immune cell behavior. Two organizing principles that have emerged from the past decade of work structure this review. First, MSICs act as mechanical immune checkpoints. Their gating state controls T-cell, NK-cell, and dendritic-cell function in stiff tumor stroma. This parallels the chemical checkpoints exploited by current immunotherapies. Second, MSICs display context-dependent functional polarity. The same channel produces opposite outputs depending on the magnitude, geometry, and time scale of the mechanical input. PIEZO1 has been reported to promote T-cell activation under physiological shear in some experimental settings but restrain cytotoxicity in stiff stroma in others. It drives a pro-inflammatory macrophage phenotype in atherosclerosis but a tissue-reparative phenotype in sepsis. It restrains ILC2s acutely but drives type-2 pathology chronically. We integrate these principles with the structural biophysics of MSIC gating, contrasting force-from-lipid and force-from-filament mechanisms. We then trace MSIC function across macrophages, microglia, T cells, NK cells, dendritic cells, neutrophils, B cells, and innate lymphoid cells. We connect this biology to the emerging mechanomedicine toolkit: sonogenetics, magnetogenetics, force-responsive nanoparticles, organ-on-chip platforms, and engineered cellular therapies. Together, mechanical immune checkpoints and context-dependent polarity reframe immunity as a force-programmable system. This view positions MSICs as translational substrates for next-generation immunotherapies in cancer, autoimmunity, fibrosis, and chronic inflammation.
Mechanosensation is fundamentally viewed as a plasma membrane phenomenon. We challenge this paradigm by introducing intracellular mechanosensation in intestinal smooth muscle. We hypothesized that a distinct, organelle-based signalling axis exists to amplify mechanotransduction from the inside out. To test this we investigated whether Piezo1, a canonical plasma membrane (PM) mechanosensor, also operates within the cell. Using tissue-level wire myography, high-resolution confocal microscopy, proximity ligation assays and patch-clamp electrophysiology on freshly dissociated cells, we identified a previously uncharacterized Piezo1-RyR-BKCa signalling axis in small intestinal smooth muscle cells (SMC). This intracellular mechanism relies on a nanoscale signalling complex (<40 nm) comprising an intracellular sensor (intra-Piezo1) and an amplifier (ryanodine receptor, RyR), coupled with a PM effector (large-conductance, Ca2+-activated K+ channels, i.e., BKCa channels). Activating this intracellular complex generated paxilline-sensitive outward currents independent of extracellular Ca2+ and dependent on internal SR Ca2+ stores, consistent with intrinsic organellar mechanotransduction. Within this complex intra-Piezo1 and RyR are positioned to operate as a coupled SR Ca2+ release unit that activates BK channels at SR-PM junctions, driving potent membrane hyperpolarization that reduces smooth muscle contractility, revealing the intra-Piezo1 complex as a molecular brake on excitation. These findings support a model in which mechanotransduction is not confined to the cell surface. Instead a specialized sensor-amplifier-effector complex originating at intracellular organelles amplifies cellular sensitivity to physical force, providing a critical gain-control system that restrains smooth muscle excitability and regulates gastrointestinal (GI) motility. KEY POINTS: This study advances a framework in which intracellular organelles contribute to mechanosensory signalling in gastrointestinal (GI) smooth muscle cells, complementing plasma membrane mechanisms. Piezo1 is predominantly intracellular in intestinal smooth muscle, where this pool, intra-Piezo1, forms a nanoscale signalling complex on the sarcoplasmic reticulum (SR) that positions it within <40 nm of RyR and of sarcolemmal large-conductance, Ca2+-activated K+ (BKCa) channels. Electrophysiological recordings show that this 'sensor-amplifier-effector' mechanism generates potent paxilline-sensitive hyperpolarizing currents that depend on SR Ca2+ release and persist without extracellular Ca2+, amplifying the cell's response to mechanical stress when intra-Piezo1 is activated from the inside out. Activation of this intra-Piezo1-mediated axis significantly dampens smooth muscle contractility, acting as a molecular 'brake' that supports the stretch-induced-relaxation feedback mechanism essential for intestinal function.
Geoanna M. Bautista, Declan Manning, Emily C. Lieu et al.· Journal of Physiology· 0 citations
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