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The skin is a complex organ that acts as a protective layer against the external environment. It protects the internal tissues from harmful agents, dehydration, ultraviolet radiation and physical injury as well as conferring thermoregulatory control, sensation, immunological surveillance and various biochemical functions. The diverse cell types that make up the skin include 1) keratinocytes, which form the bulk of the protective outer layer; 2) melanocytes, which protect the body from ultraviolet radiation by secreting the pigment melanin; and 3) cells that form the secretory appendages: eccrine and apocrine sweat glands, and the sebaceous gland. Emerging evidence suggests that store-operated Ca2+ entry (SOCE), whereby depletion of intracellular Ca2+ stores triggers Ca2+ influx across the plasma membrane, is central to the normal physiology of these cells and thus skin function. Numerous skin pathologies including dermatitis, anhidrotic ectodermal dysplasia, hyperhidrosis, hair loss and cancer are now linked to dysfunction in SOCE proteins. Principal amongst these are the stromal interaction molecules (STIMs) that sense Ca2+ depletion and Orai channels that mediate Ca2+ influx. In this review, the roles of STIM, Orai and other store-operated channels are discussed in the context of keratinocyte differentiation, melanogenesis, and eccrine sweat secretion. We explore not only STIM1-Orai1 as drivers of SOCE, but also independent actions of STIM, and emerging signal cascades stemming from their activities. Roles are discussed for the elusive transient receptor potential canonical channel (TRPC) complex in keratinocytes, Orai channels in Ca2+-cyclic AMP signal crosstalk in melanocytes, and Orai isoforms in eccrine sweat gland secretion.

Background and Purpose: The canonical Kir6.2/SUR2A ventricular KATP channel is highly ATP‐sensitive and remains closed under normal physiological conditions. These channels activate only when prolonged metabolic compromise causes significant ATP depletion and then shortens the action potential to reduce contractile activity. Pharmacological activation of KATP channels is cardioprotective, but physiologically, it is difficult to understand how these channels protect the heart if they only open under extreme metabolic stress. The presence of a second KATP channel population could help explain this. Here, we characterise the biophysical and pharmacological behaviours of a constitutively active Kir6.1‐containing KATP channel in ventricular cardiomyocytes. Experimental Approach: Patch‐clamp recordings from rat ventricular myocytes in combination with well‐defined pharmacological modulators was used to characterise these newly identified K+ channels. Action potential recording, calcium (Fluo‐4) fluorescence measurements and video edge detection of contractile function were used to assess functional consequences of channel modulation. Key Results: Our data show a ventricular K+ conductance whose biophysical characteristics and response to pharmacological modulation were consistent with Kir6.1‐containing channels. These Kir6.1‐containing channels lack the ATP‐sensitivity of the canonical channels and are constitutively active. Conclusion and Implications: We conclude there are two functionally distinct populations of ventricular KATP channels: constitutively active Kir6.1‐containing channels that play an important role in fine‐tuning the action potential and Kir6.2/SUR2A channels that activate with prolonged ischaemia to impart late‐stage protection against catastrophic ATP depletion. Further research is required to determine whether Kir6.1 is an overlooked target in Comprehensive in vitro Proarrhythmia Assay (CiPA) cardiac safety screens. [ABSTRACT FROM AUTHOR]

Calmodulin (CaM) is a highly conserved mediator of calcium (Ca2+)‐dependent signalling and modulates various cardiac ion channels. Genotyping has revealed several CaM mutations associated with long QT syndrome (LQTS). LQTS patients display prolonged ventricular recovery times (QT interval), increasing their risk of incurring life‐threatening arrhythmic events. Loss‐of‐function mutations to Kv7.1 (which drives the slow delayed rectifier potassium current, IKs, a key ventricular repolarising current) are the largest contributor to congenital LQTS (>50% of cases). CaM modulates Kv7.1 to produce a Ca2+‐sensitive IKs, but little is known about the consequences of LQTS‐associated CaM mutations on Kv7.1 function. Here, we present novel data characterising the biophysical and modulatory properties of three LQTS‐associated CaM variants (D95V, N97I and D131H). We showed that mutations induced structural alterations in CaM and reduced affinity for Kv7.1, when compared with wild‐type (WT). Using HEK293T cells expressing Kv7.1 channel subunits (KCNQ1/KCNE1) and patch‐clamp electrophysiology, we demonstrated that LQTS‐associated CaM variants reduced current density at systolic Ca2+ concentrations (1 μm), revealing a direct QT‐prolonging modulatory effect. Our data highlight for the first time that LQTS‐associated perturbations to CaM's structure impede complex formation with Kv7.1 and subsequently result in reduced IKs. This provides a novel mechanistic insight into how the perturbed structure–function relationship of CaM variants contributes to the LQTS phenotype. Key points: Calmodulin (CaM) is a ubiquitous, highly conserved calcium (Ca2+) sensor playing a key role in cardiac muscle contraction.Genotyping has revealed several CaM mutations associated with long QT syndrome (LQTS), a life‐threatening cardiac arrhythmia syndrome.LQTS‐associated CaM variants (D95V, N97I and D131H) induced structural alterations, altered binding to Kv7.1 and reduced IKs.Our data provide a novel mechanistic insight into how the perturbed structure–function relationship of CaM variants contributes to the LQTS phenotype. [ABSTRACT FROM AUTHOR]

Skin is the largest organ in the human body with ~95% of its surface made up of keratinocytes. These cells maintain a healthy skin barrier through regulated differentiation driven by Ca2+-transcriptional coupling. Many important skin conditions arise from disruption of this process although not all stages are fully understood. We know that elevated extracellular Ca2+ at the skin surface is detected by keratinocyte Gαq-coupled receptors that signal to empty endoplasmic reticulum Ca2+ stores. Orai channel store-operated Ca2+ entry (SOCE) and Ca2+ influx via "canonical" transient receptor potential (TRPC)-composed channels then activates transcription factors that drive differentiation. While STIM-mediated activation of Orai channels following store depletion is well defined, how TRPC channels are activated is less clear. Multiple modes of TRPC channel activation have been proposed, including 1) independent TRPC activation by STIM, 2) formation of Orai-TRPC-STIM complexes, and 3) the insertion of constitutively-active TRPC channels into the membrane during SOCE. To help distinguish between these models, we used high-resolution microscopy of intact keratinocyte (HaCaT) cells and immunogold transmission electron microscopy (TEM) of HaCaT plasma membrane sheets. Our data shows no evidence of significant insertion of Orai1 or TRPC subunits into the membrane during SOCE. Analysis of transmission electron microscopy data shows that during store-depletion and SOCE, Orai1 and TRPC subunits form separate membrane-localized clusters that migrate towards each other. This clustering of TRPC channel subunits in keratinocytes may support the formation of TRPC-STIM interactions at ER-plasma membrane junctions that are distinct from Orai-STIM junctions. [ABSTRACT FROM AUTHOR]

The small-angle X-ray scattering and NMR spectroscopy are used for examining the conformation of the PDZ domain region of SAP97 PDZ1-3. The restricted dynamics and preferential Kir2.1 binding to PDZ2 are features that have enabled SAP97 to function as a scaffold protein, thus allowing other proteins to bind to the other two PDZ domains in sufficient proximity to produce productive channelosomes.