Chirality Probe of Twisted Bilayer Graphene in the Linear Transport Regime.

We propose minimal transport experiments in the coherent regime that can probe the chirality of twisted moiré structures. We show that only with a third contact and in the presence of an in-plane magnetic field (or another time-reversal symmetry breaking effect) a chiral system may display nonreciprocal transport in the linear regime. We then propose to use the third lead as a voltage probe and show that opposite enantiomers give rise to different voltage drops on the third lead. Additionally, in the scenario of layer-discriminating contacts, the third lead can serve as a current probe capable of detecting different handedness even in the absence of a magnetic field. In a complementary configuration, applying opposite voltages on the two layers of the third lead gives rise to a chiral (super)current in the absence of a source-drain voltage whose direction is determined by its chirality.

An optogenetic method for the controlled release of single molecules.

We developed a system for optogenetic release of single molecules in cells. We confined soluble and transmembrane proteins to the Golgi apparatus via a photocleavable protein and released them by short pulses of light. Our method allows for a light dose-dependent delivery of functional proteins to the cytosol and plasma membrane in amounts compatible with single-molecule imaging, greatly simplifying access to single-molecule microscopy of any protein in live cells. We were able to reconstitute ion conductance by delivering BK and LRRC8/volume-regulated anion channels to the plasma membrane. Finally we were able to induce NF-kB signaling in T lymphoblasts stimulated by interleukin-1 by controlled release of a signaling protein that had been knocked out. We observed light-induced formation of functional inflammatory signaling complexes that triggered phosphorylation of the inhibitor of nuclear factor kappa-B kinase only in activated cells. We thus developed an optogenetic method for the reconstitution and investigation of cellular function at the single-molecule level.

Physiological Functions of the Volume-Regulated Anion Channel VRAC/LRRC8 and the Proton-Activated Chloride Channel ASOR/TMEM206.

Volume-regulated anion channels (VRACs) and the acid-sensitive outwardly rectifying anion channel (ASOR) mediate flux of chloride and small organic anions. Although known for a long time, they were only recently identified at the molecular level. VRACs are heteromers consisting of LRRC8 proteins A to E. Combining the essential LRRC8A with different LRRC8 paralogues changes key properties of VRAC such as conductance or substrate selectivity, which is how VRACs are involved in multiple physiological functions including regulatory volume decrease, cell proliferation and migration, cell death, purinergic signalling, fat and glucose metabolism, insulin signalling, and spermiogenesis. VRACs are also involved in pathological conditions, such as the neurotoxic release of glutamate and aspartate. Certain VRACs are also permeable to larger, organic anions, including antibiotics and anti-cancer drugs, making them an interesting therapeutic target. ASOR, also named proton-activated chloride channel (PAC), is formed by TMEM206 homotrimers on the plasma membrane and on endosomal compartments where it mediates chloride flux in response to extracytosolic acidification and plays a role in the shrinking and maturation of macropinosomes. ASOR has been shown to underlie neuronal swelling which causes cell death after stroke as well as promoting the metastasis of certain cancers, making them intriguing therapeutic targets as well.

Impaired Autophagic Clearance with a Gain-of-Function Variant of the Lysosomal Cl-/H+ Exchanger ClC-7.

ClC-7 is a ubiquitously expressed voltage-gated Cl-/H+ exchanger that critically contributes to lysosomal ion homeostasis. Together with its ß-subunit Ostm1, ClC-7 localizes to lysosomes and to the ruffled border of osteoclasts, where it supports the acidification of the resorption lacuna. Loss of ClC-7 or Ostm1 leads to osteopetrosis accompanied by accumulation of storage material in lysosomes and neurodegeneration. Interestingly, not all osteopetrosis-causing CLCN7 mutations from patients are associated with a loss of ion transport. Some rather result in an acceleration of voltage-dependent ClC-7 activation. Recently, a gain-of-function variant, ClC-7Y715C, that yields larger ion currents upon heterologous expression, was identified in two patients with neurodegeneration, organomegaly and albinism. However, neither the patients nor a mouse model that carried the equivalent mutation developed osteopetrosis, although expression of ClC-7Y715C induced the formation of enlarged intracellular vacuoles. Here, we investigated how, in transfected cells with mutant ClC-7, the substitution of this tyrosine impinged on the morphology and function of lysosomes. Combinations of the tyrosine mutation with mutations that either uncouple Cl- from H+ counter-transport or strongly diminish overall ion currents were used to show that increased ClC-7 Cl-/H+ exchange activity is required for the formation of enlarged vacuoles by membrane fusion. Degradation of endocytosed material was reduced in these compartments and resulted in an accumulation of lysosomal storage material. In cells expressing the ClC-7 gain-of-function mutant, autophagic clearance was largely impaired, resulting in a build-up of autophagic material.

Trends in volume-regulated anion channel (VRAC) research: visualization and bibliometric analysis from 2014 to 2022.

Objective: In this study, we utilized bibliometric methods to assess the worldwide scientific output and identify hotspots related to the research on the volume-regulated anion channel (VRAC) from 2014 to 2022. Methods: From Web of Science, we obtained studies related to VRAC published from 2014 to 2022. To analyzed the data, we utilized VOSviewer, a tool for visualizing network, to create networks based on the collaboration between countries, institutions, and authors. Additionally, we performed an analysis of journal co-citation, document citation, and co-occurrence of keywords. Furthermore, we employed CiteSpace (6.1. R6 Advanced) to analyzed keywords and co-cited references with the strongest burst. Results: The final analysis included a total of 278 related articles and reviews, covering the period from 2014 to 2022. The United States emerged as the leading country contributing to this field, while the University of Copenhagen stood out as the most prominent institution. The author with most publications and most citations was Thomas J. Jentsch. Among the cited references, the article by Voss et al. published in Science (2014) gained significant attention for its identification of LRRC8 heteromers as a crucial component of the volume-regulated anion channel VRAC. Pflügers Archiv European Journal of Physiology and Journal of Physiology-London were the leading journals in terms of the quantity of associated articles and citations. Through the analysis of keyword co-occurrence, it was discovered that VRAC is involved in various physiological processes including cell growth, migration, apoptosis, swelling, and myogenesis, as well as anion and organic osmolyte transport including chloride, taurine, glutamate and ATP. VRAC is also associated with related ion channels such as TMEM16A, TMEM16F, pannexin, and CFTR, and associated with various diseases including epilepsy, leukodystrophy, atherosclerosis, hypertension, cerebral edema, stroke, and different types of cancer including gastric cancer, glioblastoma and hepatocellular carcinoma. Furthermore, VRAC is involved in anti-tumor drug resistance by regulating the uptake of platinum-based drugs and temozolomide. Additionally, VRAC has been studied in the context of pharmacology involving DCPIB and flavonoids. Conclusion: The aim of this bibliometric analysis is to provide an overall perspective for research on VRAC. VRAC has become a topic of increasing interest, and our analysis shows that it continues to be a prominent area. This study offers insights into the investigation of VRAC channel and may guide researchers in identifying new directions for future research.

Functional analysis of two SLC9A6 frameshift variants in lymphoblastoid cells from patients with Christianson syndrome.

BACKGROUND: Christianson syndrome (CS) is caused by mutations in SLC9A6 and is characterized by global developmental delay, epilepsy, hyperkinesis, ataxia, microcephaly, and behavioral disorder. However, the molecular mechanism by which these SLC9A6 mutations cause CS in humans is not entirely understood, and there is no objective method to determine the pathogenicity of single SLC9A6 variants. METHODS: Trio-based whole exome sequencing (WES) was carried out on two individuals with suspicion of CS. qRT-PCR, western blot analysis, filipin staining, lysosomal enzymatic assays, and electron microscopy examination, using EBV-LCLs established from the two patients, were performed. RESULTS: Trio-based WES identified a hemizygous SLC9A6 c.1560dupT, p.T521Yfs*23 variant in proband 1 and a hemizygous SLC9A6 c.608delA, p.H203Lfs*10 variant in proband 2. Both children exhibited typical phenotypes associated with CS. Expression analysis in EBV-LCLs derived from the two patients showed a significant decrease in mRNA levels and no detectable normal NHE6 protein. EBV-LCLs showed a statistically significant increase in unesterified cholesterol in patient 1, but only non-significant increase in patient 2 when stained with filipin. Activities of lysosomal enzymes (ß-hexosaminidase A, ß-hexosaminidase A + B, ß-galactosidase, galactocerebrosidase, arylsulfatase A) of EBV-LCLs did not significantly differ between the two patients and six controls. Importantly, by electron microscopy we detected an accumulation of lamellated membrane structures, deformed mitochondria, and lipid droplets in the patients' EBV-LCLs. CONCLUSIONS: The SLC9A6 p.T521Yfs*23 and p.H203Lfs*10 variants in our patients result in loss of NHE6. Alterations of mitochondria and lipid metabolism may play a role in the pathogenesis of CS. Moreover, the combination of filipin staining with electron microscopy examination of patient lymphoblastoid cells can serve as a useful complementary diagnostic method for CS.

CLCN7, a gene shared by autosomal recessive and autosomal dominant osteopetrosis.

After the discovery of abundant v-ATPase complexes in the osteoclast ruffled membrane it was obvious that in parallel a negative counter-ion needs to be transported across this membrane to allow for efficient transport of protons into the resorption lacuna. While different candidate proteins were discussed the osteopetrosis phenotype of Clcn7 knockout mice suggested that the chloride/proton-exchanger ClC-7 might be responsible for transporting the negative charge. In the following, individuals with autosomal recessive osteopetrosis (ARO) were found to carry biallelic CLCN7 pathogenic variants. Shortly thereafter, heterozygous pathogenic variants were identified as the exclusive cause of autosomal dominant osteopetrosis type 2 (ADO2). Since in most cell types other than osteoclasts ClC-7 resides in late endosomes and lysosomes, it took some time until the electrophysiological properties of ClC-7 were elucidated. Whereas most missense variants lead to reduced chloride currents, several variants with accelerated kinetics have been identified. Evidence for folding problems is also known for several missense variants. Paradoxically, a heterozygous activating variant in ClC-7 was described to cause lysosomal alteration, pigmentation defects, and intellectual disability without osteopetrosis. The counter-intuitive 2 Cl-/H+ exchange function of ClC-7 was shown to be physiologically important for intravesicular ion homeostasis. The lysosomal function of ClC-7 is also the reason why individuals with CLCN7-ARO can develop a storage disorder and neurodegeneration, a feature that is variable and difficult to predict. Furthermore, the low penetrance of heterozygous pathogenic CLCN7 variants and the clinical variability of ADO2 are incompletely understood. We aim to give an overview not only of the current knowledge about ClC-7 and its related pathologies, but also of the scientists and clinicians that paved the way for these discoveries.

Ion Channels and Transporters in Muscle Cell Differentiation.

Investigations on ion channels in muscle tissues have mainly focused on physiological muscle function and related disorders, but emerging evidence supports a critical role of ion channels and transporters in developmental processes, such as controlling the myogenic commitment of stem cells. In this review, we provide an overview of ion channels and transporters that influence skeletal muscle myoblast differentiation, cardiac differentiation from pluripotent stem cells, as well as vascular smooth muscle cell differentiation. We highlight examples of model organisms or patients with mutations in ion channels. Furthermore, a potential underlying molecular mechanism involving hyperpolarization of the resting membrane potential and a series of calcium signaling is discussed.

The expanding toolbox to study the LRRC8-formed volume-regulated anion channel VRAC.

The volume-regulated anion channel (VRAC) is activated upon cell swelling and facilitates the passive movement of anions across the plasma membrane in cells. VRAC function underlies many critical homeostatic processes in vertebrate cells. Among them are the regulation of cell volume and membrane potential, glutamate release and apoptosis. VRAC is also permeable for organic osmolytes and metabolites including some anti-cancer drugs and antibiotics. Therefore, a fundamental understanding of VRAC's structure-function relationships, its physiological roles, its utility for therapy of diseases, and the development of compounds modulating its activity are important research frontiers. Here, we describe approaches that have been applied to study VRAC since it was first described more than 30 years ago, providing an overview of the recent methodological progress. The diverse applications reflecting a compromise between the physiological situation, biochemical definition, and biophysical resolution range from the study of VRAC activity using a classic electrophysiology approach, to the measurement of osmolytes transport by various means and the investigation of its activation using a novel biophysical approach based on fluorescence resonance energy transfer.

The molecular and phenotypic spectrum of CLCN4-related epilepsy.

OBJECTIVE: This study was undertaken to expand the phenotypic and genetic spectrum of CLCN4-related epilepsy and to investigate genotype-phenotype correlations. METHODS: We systematically reviewed the phenotypic and genetic spectrum of newly diagnosed and previously reported patients with CLCN4-related epilepsy. Three novel variants identified in four patients reported in this study were evaluated through in silico prediction and functional analysis by Western blot, immunofluorescence, and electrophysiological measurements. RESULTS: Epilepsy was diagnosed in 54.55% (24/44) of individuals with CLCN4-related disorders and was drug-resistant in most cases. Of 24 patients, 15 had epileptic encephalopathy and four died at an early age; 69.57% of patients had seizure onset within the first year of life. Myoclonic seizures are the most common seizure type, and 56.25% of patients presented multiple seizure types. Notably, seizure outcome was favorable in individuals with only one seizure type. All patients showed intellectual disability, which was severe in 65.22% of patients. Additional common features included language delay, behavioral disorders, and dysmorphic features. Five patients benefitted from treatment with lamotrigine. Most variants, which were mainly missense (79.17%), were inherited (70.83%). Whereas frameshift, intragenic deletion, or inherited variants were associated with milder phenotypes, missense or de novo variants led to more severe phenotypes. All evaluated CLCN4 variants resulted in loss of function with reduced ClC-4 currents. Nonetheless, genotype-phenotype relationships for CLCN4-related epilepsy are not straightforward, as phenotypic variability was observed in recurrent variants and within single families. SIGNIFICANCE: Pathogenic CLCN4 variants contribute significantly to the genetic etiology of epilepsy. The phenotypic spectrum of CLCN4-related epilepsy includes drug-resistant seizures, cognitive and language impairment, behavioral disorders, and congenital anomalies. Notably, the mutation type and the number of seizure types correlate with the severity of the phenotype, suggesting its use for clinical prognosis. Lamotrigine can be considered a therapeutic option.

Efficient generation of osteoclasts from human induced pluripotent stem cells and functional investigations of lethal CLCN7-related osteopetrosis.

Human induced pluripotent stem cells (hiPSCs) hold great potential for modeling human diseases and the development of innovative therapeutic approaches. Here, we report on a novel, simplified differentiation method for forming functional osteoclasts from hiPSCs. The three-step protocol starts with embryoid body formation, followed by hematopoietic specification, and finally osteoclast differentiation. We observed continuous production of monocyte-like cells over a period of up to 9 weeks, generating sufficient material for several osteoclast differentiations. The analysis of stage-specific gene and surface marker expression proved mesodermal priming, the presence of monocyte-like cells, and of terminally differentiated multinucleated osteoclasts, able to form resorption pits and trenches on bone and dentine in vitro. In comparison to peripheral blood mononuclear cell (PBMC)-derived osteoclasts hiPSC-derived osteoclasts were larger and contained a higher number of nuclei. Detailed functional studies on the resorption behavior of hiPSC-osteoclasts indicated a trend towards forming more trenches than pits and an increase in pseudoresorption. We used hiPSCs from an autosomal recessive osteopetrosis (ARO) patient (BIHi002-A, ARO hiPSCs) with compound heterozygous missense mutations p.(G292E) and p.(R403Q) in CLCN7, coding for the Cl- /H+ -exchanger ClC-7, for functional investigations. The patient's leading clinical feature was a brain malformation due to defective neuronal migration. Mutant ClC-7 displayed residual expression and retained lysosomal co-localization with OSTM1, the gene coding for the osteopetrosis-associated transmembrane protein 1, but only ClC-7 harboring the mutation p.(R403Q) gave strongly reduced ion currents. An increased autophagic flux in spite of unchanged lysosomal pH was evident in undifferentiated ARO hiPSCs. ARO hiPSC-derived osteoclasts showed an increased size compared to hiPSCs of healthy donors. They were not able to resorb bone, underlining a loss-of-function effect of the mutations. In summary, we developed a highly reproducible, straightforward hiPSC-osteoclast differentiation protocol. We demonstrated that osteoclasts differentiated from ARO hiPSCs can be used as a disease model for ARO and potentially also other osteoclast-related diseases. © 2021 The Authors. Journal of Bone and Mineral Research published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research (ASBMR).

Neurodegeneration Upon Dysfunction of Endosomal/Lysosomal CLC Chloride Transporters.

The regulation of luminal ion concentrations is critical for the function of, and transport between intracellular organelles. The importance of the acidic pH in the compartments of the endosomal-lysosomal pathway has been well-known for decades. Besides the V-ATPase, which pumps protons into their lumen, a variety of ion transporters and channels is involved in the regulation of the organelles' complex ion homeostasis. Amongst these are the intracellular members of the CLC family, ClC-3 through ClC-7. They localize to distinct but overlapping compartments of the endosomal-lysosomal pathway, partially with tissue-specific expression. Functioning as 2Cl-/H+ exchangers, they can support the vesicular acidification and accumulate luminal Cl-. Mutations in the encoding genes in patients and mouse models underlie severe phenotypes including kidney stones with CLCN5 and osteopetrosis or hypopigmentation with CLCN7. Dysfunction of those intracellular CLCs that are expressed in neurons lead to neuronal defects. Loss of endosomal ClC-3, which heteromerizes with ClC-4, results in neurodegeneration. Mutations in ClC-4 are associated with epileptic encephalopathy and intellectual disability. Mice lacking the late endosomal ClC-6 develop a lysosomal storage disease with reduced pain sensitivity. Human gene variants have been associated with epilepsy, and a gain-of-function mutation causes early-onset neurodegeneration. Dysfunction of the lysosomal ClC-7 leads to a lysosomal storage disease and neurodegeneration in mice and humans. Reduced luminal chloride, as well as altered calcium regulation, has been associated with lysosomal storage diseases in general. This review discusses the properties of endosomal and lysosomal Cl-/H+ exchange by CLCs and how various alterations of ion transport by CLCs impact organellar ion homeostasis and function in neurodegenerative disorders.

West Syndrome Caused By a Chloride/Proton Exchange-Uncoupling CLCN6 Mutation Related to Autophagic-Lysosomal Dysfunction.

Vesicular chloride/proton exchangers of the CLC family are critically involved in the function of the endosomal-lysosomal pathway. Their dysfunction leads to severe disorders including intellectual disability and epilepsy for ClC-4, Dent's disease for ClC-5, and lysosomal storage disease and osteopetrosis for ClC-7. Here, we report a de novo variant p.Glu200Ala (p.E200A; c.599A>C) of the late endosomal ClC-6, encoded by CLCN6, in a patient with West syndrome (WS), severe developmental delay, autism, movement disorder, microcephaly, facial dysmorphism, and visual impairment. Mutation of this conserved glutamate uncouples chloride transport from proton antiport by ClC-6. This affects organellar ion homeostasis and was shown to be deleterious for other CLCs. In this study, we found that upon heterologous expression, the ClC-6 E200A variant caused autophagosome accumulation and impaired the clearance of autophagosomes by blocking autophagosome-lysosome fusion. Our study provides clinical and functional support for an association between CLCN6 variants and WS. Our findings also provide novel insights into the molecular mechanisms underlying the pathogenesis of WS, suggesting an involvement of autophagic-lysosomal dysfunction.

Plasmon-Enhanced Near-Field Chirality in Twisted van der Waals Heterostructures.

It is shown that chiral plasmons, characterized by a longitudinal magnetic moment accompanying the longitudinal charge plasmon, lead to electromagnetic near-fields that are also chiral. For twisted bilayer graphene, we estimate that the near-field chirality of screened plasmons can be several orders of magnitude larger than that of the related circularly polarized light. The chirality also manifests itself in a deflection angle that is formed between the direction of the plasmon propagation and its Poynting vector. Twisted van der Waals heterostructures might thus provide a novel platform to promote enantiomer-selective physio-chemical processes in chiral molecules without the application of a magnetic field or external nanopatterning that break time-reversal, mirror plane, or inversion symmetry, respectively.

LRRC8 channel activation and reduction in cytosolic chloride concentration during early differentiation of C2C12 myoblasts.

Leucine-rich repeat containing family 8 (LRRC8) proteins form the volume-regulated anion channel (VRAC). Recently, they were shown to be required for normal differentiation and fusion of C2C12 myoblasts, by promoting membrane hyperpolarization and intracellular Ca2+ signals. However, the mechanism by which they are involved remained obscure. Here, using a FRET-based sensor for VRAC activity, we show temporary activation of VRAC within the first 2 h of myogenic differentiation. During this period, we also observed a significant decrease in the intracellular Cl- concentration that was abolished by the VRAC inhibitor carbenoxolone. However, lowering the intracellular Cl- concentration by extracellular Cl- depletion did not promote differentiation as judged by the percentage of myogenin-positive nuclei or total myogenin levels in C2C12 cells. Instead, it inhibited myosin expression and myotube formation. Together, these data suggest that VRAC is activated and mediates Cl- efflux early on during myogenic differentiation, and a moderate intracellular Cl- concentration is necessary for myoblast fusion.

Chiral Plasmons with Twisted Atomic Bilayers.

van der Waals heterostructures of atomically thin layers with rotational misalignments, such as twisted bilayer graphene, feature interesting structural moiré superlattices. Because of the quantum coupling between the twisted atomic layers, light-matter interaction is inherently chiral; as such, they provide a promising platform for chiral plasmons in the extreme nanoscale. However, while the interlayer quantum coupling can be significant, its influence on chiral plasmons still remains elusive. Here we present the general solutions from full Maxwell equations of chiral plasmons in twisted atomic bilayers, with the consideration of interlayer quantum coupling. We find twisted atomic bilayers have a direct correspondence to the chiral metasurface, which simultaneously possesses chiral and magnetic surface conductivities, besides the common electric surface conductivity. In other words, the interlayer quantum coupling in twisted van der Waals heterostructures may facilitate the construction of various (e.g., bi-anisotropic) atomically-thin metasurfaces. Moreover, the chiral surface conductivity, determined by the interlayer quantum coupling, determines the existence of chiral plasmons and leads to a unique phase relationship (i.e., ±π/2 phase difference) between their transverse-electric (TE) and transverse-magnetic (TM) wave components. Importantly, such a unique phase relationship for chiral plasmons can be exploited to construct the missing longitudinal spin of plasmons, besides the common transverse spin of plasmons.

Nano-photocurrent Mapping of Local Electronic Structure in Twisted Bilayer Graphene.

We report a combined nano-photocurrent and infrared nanoscopy study of twisted bilayer graphene (TBG) enabling access to the local electronic phenomena at length scales as short as 20 nm. We show that the photocurrent changes sign at carrier densities tracking the local superlattice density of states of TBG. We use this property to identify domains of varying local twist angle by local photothermoelectric effect. Consistent with the photocurrent study, infrared nanoimaging experiments reveal optical conductivity features dominated by twist-angle-dependent interband transitions. Our results provide a fast and robust method for mapping the electronic structure of TBG and suggest that similar methods can be broadly applied to probe electronic inhomogeneities of Moiré superlattices in other van der Waals heterostructures.

Absolute Protein Amounts and Relative Abundance of Volume-regulated Anion Channel (VRAC) LRRC8 Subunits in Cells and Tissues Revealed by Quantitative Immunoblotting.

The volume-regulated anion channel (VRAC) plays an important role in osmotic cell volume regulation. In addition, it is involved in various physiological processes such as insulin secretion, glia-neuron communication and purinergic signaling. VRAC is formed by hetero-hexamers of members of the LRRC8 protein family, which consists of five members, LRRC8A-E. LRRC8A is an essential subunit for physiological functionality of VRAC. Its obligate heteromerization with at least one of its paralogues, LRRC8B-E, determines the biophysical properties of VRAC. Moreover, the subunit composition is of physiological relevance as it largely influences the activation mechanism and especially the substrate selectivity. However, the endogenous tissue-specific subunit composition of VRAC is unknown. We have now developed and applied a quantitative immunoblot study of the five VRAC LRRC8 subunits in various mouse cell lines and tissues, using recombinant protein for signal calibration. We found tissue-specific expression patterns of the subunits, and generally relative low expression of the essential LRRC8A subunit. Immunoprecipitation of LRRC8A also co-precipitates an excess of the other subunits, suggesting that non-LRRC8A subunits present the majority in hetero-hexamers. With this, we can estimate that in the tested cell lines, the number of VRAC channels per cell is in the order of 10,000, which is in agreement with earlier calculations from the comparison of single-channel and whole-cell currents.

A Mathematical Model of Lysosomal Ion Homeostasis Points to Differential Effects of Cl- Transport in Ca2+ Dynamics.

The establishment and maintenance of ion gradients between the interior of lysosomes and the cytosol are crucial for numerous cellular and organismal functions. Numerous ion transport proteins ensure the required variation in luminal concentrations of the different ions along the endocytic pathway to fit the needs of the organelles. Failures in keeping proper ion homeostasis have pathological consequences. Accordingly, several human diseases are caused by the dysfunction of ion transporters. These include osteopetrosis, caused by the dysfunction of Cl-/H+ exchange by the lysosomal transporter ClC-7. To better understand how chloride transport affects lysosomal ion homeostasis and how its disruption impinges on lysosomal function, we developed a mathematical model of lysosomal ion homeostasis including Ca2+ dynamics. The model recapitulates known biophysical properties of ClC-7 and enables the investigation of its differential activation kinetics on lysosomal ion homeostasis. We show that normal functioning of ClC-7 supports the acidification process, is associated with increased luminal concentrations of sodium, potassium, and chloride, and leads to a higher Ca2+ uptake and release. Our model highlights the role of ClC-7 in lysosomal acidification and shows the existence of differential Ca2+ dynamics upon perturbations of Cl-/H+ exchange and its activation kinetics, with possible pathological consequences.