Explore the vast range of titles available.

Gnathodiaphyseal dysplasia (GDD) is a rare autosomal dominant genetic disease, mainly characterized by enlargement of the mandible, osteosclerosis, and frequent fracture of tubular bone. GDD is caused by heterozygous mutations in Anoctamin 5 (ANO5). We have previously generated an Ano5 knockout (KO) mice model and validated the phenotypes consistent with GDD patients, including enhanced bone formation and alkaline phosphatase (ALP) activity. Experiments have identified that Ano5 deficiency elevated the osteogenesis of calvaria-derived osteoblasts (mCOBs). In this study, we found that Ano5 deficiency notably inhibited miR-34c-5p expression. Krüppel-Like Factor 4 (Klf4), a target gene of miR-34c-5p confirmed by dual luciferase reporter assay, was up-regulated in Ano5−/− mCOBs, accompanied by activated downstream canonical Wnt/β-catenin signaling and increased expression of β-catenin. Overexpression of miR-34c-5p in Ano5−/− mCOBs inhibited osteogenic capacity by suppressing proliferative capacity, osteoblast-related factor levels, ALP activity, and matrix calcification through regulating KLF4/β-catenin signaling axis. Furthermore, miR-34c-5p adeno-associated virus (AAV) treatment in vivo rescued the abnormally thickened cortical bone and enhanced biomechanical properties in Ano5−/− mice. Importantly, the serum level of P1NP, a marker of bone formation, was also significantly declined. We conclude that dysregulation of miR-34c-5p contributes to the enhanced osteogenesis in GDD by excessive activation of KLF4/β-catenin signaling axis under Ano5-deficient conditions. This study elucidates the pathogenesis of GDD and provides novel insights into the therapeutic strategies.

TMEM16B (ANO2) is the Ca2+-activated chloride channel expressed in multiple brain regions, including the amygdala. Here we report that Ano2 knockout mice exhibit impaired anxiety-related behaviors and context-independent fear memory, thus implicating TMEM16B in anxiety modulation. We found that TMEM16B is expressed in somatostatin-positive (SOM+) GABAergic neurons of the central lateral amygdala (CeL), and its activity modulates action potential duration and inhibitory postsynaptic current (IPSC). We further provide evidence for TMEM16B actions not only in the soma but also in the presynaptic nerve terminals of GABAergic neurons. Our study reveals an intriguing role for TMEM16B in context-independent but not context-dependent fear memory, and supports the notion that dysfunction of the amygdala contributes to anxiety-related behaviors.

Mutations in the anoctamin-5 (ANO5) gene can lead to musculoskeletal disorders, with monoallelic (autosomal dominant) mutations typically presenting as skeletal abnormalities known as gnathodiaphyseal dysplasia (GDD). Clinically, GDD is characterized by thickened cortices of long bones and mandibles, narrowed medullary cavities, and increased bone fragility. While autophagy is necessary in regulating bone formation, the specific relationship between ANO5 and autophagy remains poorly understood. In this study, we demonstrated that Ano5 deficiency activates autophagy in mouse cranial osteoblasts (mCOBs), leading to enhanced osteogenic capacity in Ano5-/- mCOBs. The application of 3-methyladenine (3-MA) and chloroquine (CQ) reversed the excessive osteogenesis observed in Ano5-/- mCOBs. Further analysis revealed that Ano5 deficiency upregulated the expression of ATG9A, and silencing ATG9A significantly reduced both autophagy and osteogenic activity in Ano5-/- mCOBs. Additionally, AMP-activated protein kinase (AMPK) was found to positively regulate ATG9A, and inhibiting AMPK reduced ATG9A expression, which in turn mitigated excessive osteogenesis of Ano5-/- mCOBs. Moreover, in vivo experiments confirmed that treatment with 3-MA alleviated the bone phenotype abnormalities in Ano5-/- mice. These findings suggest that Ano5 negatively regulates autophagy, contributing to illuminate pathogenesis of GDD. Meanwhile, this research highlights potential therapeutic strategies targeting autophagy to pave the way for the clinical manifestations of GDD.

Purinergic P2X 7 receptors (P2X 7 R) are fundamental to innate immune response. In macrophages, transient stimulation of P2X 7 R activates several transport mechanisms and induces the scrambling of phospholipids with subsequent membrane blebbing and apoptosis. These processes support phagocytosis and subsequent killing of phagocytosed bacteria. Here we demonstrate that the stimulation of P2X 7 receptors activates anoctamin 6 (ANO6, TMEM16F), a protein that functions as Ca 2+ dependent phospholipid scramblase and Ca 2+ -activated Cl − channel. Inhibition or knockdown of ANO6 attenuates ATP-induced cell shrinkage, cell migration and phospholipid scrambling. In mouse macrophages, Ano6 produces large ion currents by stimulation of P2X 7 receptors and contributes to ATP-induced membrane blebbing and apoptosis, which is largely reduced in macrophages from Ano6 −/− mice. ANO6 supports bacterial phagocytosis and killing by mouse and human THP-1 macrophages. Our data demonstrate that anoctamin 6 is an essential component of the immune defense by macrophages. Activation of purinergic P2X 7 receptors is important for phagocytosis and bacterial killing. Here the authors show that a phospholipid scramblase, Anoctamin 6, is activated downstream of P2X 7 R and is a critical mediator of bacterial internalization and killing by macrophages.

Calcium-activated chloride currents are thought to amplify the olfactory signal during the transduction process. Here the authors identify Ano2 as the critical channel in both main olfactory epithelium and in the vomeronasal organ. Surprisingly, disruption of Ano2 did not reduce performance in an olfactory behavioral task, suggesting that olfaction does not actually depend on these channels. Canonical olfactory signal transduction involves the activation of cyclic AMP–activated cation channels that depolarize the cilia of receptor neurons and raise intracellular calcium. Calcium then activates Cl − currents that may be up to tenfold larger than cation currents and are believed to powerfully amplify the response. We identified Anoctamin2 (Ano2, also known as TMEM16B) as the ciliary Ca 2+ -activated Cl − channel of olfactory receptor neurons. Ano2 is expressed in the main olfactory epithelium (MOE) and in the vomeronasal organ (VNO), which also expresses the related Ano1 channel. Disruption of Ano2 in mice virtually abolished Ca 2+ -activated Cl − currents in the MOE and VNO. Ano2 disruption reduced fluid-phase electro-olfactogram responses by only ∼40%, did not change air-phase electro-olfactograms and did not reduce performance in olfactory behavioral tasks. In contrast with the current view, cyclic nucleotide–gated cation channels do not need a boost by Cl − channels to achieve near-physiological levels of olfaction.

Inherited ataxias are heterogeneous disorders affecting both children and adults, with over 40 different causative genes, making molecular genetic diagnosis challenging. Although recent advances in next-generation sequencing have significantly improved mutation detection, few treatments exist for patients with inherited ataxia. In two patients with adult-onset cerebellar ataxia and coenzyme Q 10 (CoQ 10 ) deficiency in muscle, whole exome sequencing revealed mutations in ANO10 , which encodes anoctamin 10, a member of a family of putative calcium-activated chloride channels, and the causative gene for autosomal recessive spinocerebellar ataxia-10 (SCAR10). Both patients presented with slowly progressive ataxia and dysarthria leading to severe disability in the sixth decade. Epilepsy and learning difficulties were also present in one patient, while retinal degeneration and cataract were present in the other. The detection of mutations in ANO10 in our patients indicate that ANO10 defects cause secondary low CoQ 10 and SCAR10 patients may benefit from CoQ 10 supplementation.

This study examined the associations of 25-hydroxyvitamin D and specific host genetic variants that affect vitamin D levels or its effects on immune function, with the risk of TB or mortality in children. A case-cohort sample of 466 South African infants enrolled in P1041 trial (NCT00080119) underwent 25-hydroxyvitamin D testing by chemiluminescent immunoassay. Single nucleotide polymorphisms (SNPs) that alter the effect of vitamin D [e.g. vitamin D receptor (VDR)], vitamin D levels [e.g. vitamin D binding protein (VDBP)], or toll like receptor (TLR) expression (SIGIRR including adjacent genes PKP3 and TMEM16J) were identified by real-time PCR. Outcomes were time to TB, and to the composite of TB or death by 192 weeks of follow-up. Effect modification between vitamin D status and SNPs for outcomes was assessed. Median age at 25-hydroxyvitamin D determination was 8 months; 11% were breastfed, 51% were HIV-infected and 26% had low 25-hydroxyvitamin D (<32ng/mL). By 192 weeks, 138 incident TB cases (43 definite/probable, and 95 possible) and 26 deaths occurred. Adjusting for HIV status and potential confounders, low 25-hydroxyvitamin D was associated with any TB (adjusted hazard ratio [aHR] 1.76, 95% CI 1.01-3.05; p = 0.046) and any TB or death (aHR 1.76, 95% CI 1.03-3.00; p = 0.038). Children with low 25-hydroxyvitamin D and TMEM 16J rs7111432-AA or PKP3 rs10902158-GG were at increased risk for probable/definite TB or death (aHR 8.12 and 4.83, p<0.05) and any TB or death (aHR 4.78 and 3.26, p<0.005) respectively; SNPs in VDBP, VDR, and vitamin D precursor or hydroxylation genes were not. There was significant interaction between low 25-hydroxyvitamin D and, TMEM 16J rs7111432-AA (p = 0.04) and PKP3 rs10902158-GG (p = 0.02) SNPs. Two novel SNPs, thought to be associated with innate immunity, in combination with low vitamin D levels were identified as increasing a young child's risk of developing TB disease or death. Identifying high-risk children and providing targeted interventions such as vitamin D supplementation may be beneficial. ClinicalTrials.gov NCT00080119.

The role of calcium-activated chloride channels for renal function is unknown. By immunohistochemistry we demonstrate dominant expression of the recently identified calcium-activated chloride channels, Anoctamin 1 (Ano1, TMEM16A) in human and mouse proximal tubular epithelial (PTE) cells, with some expression in podocytes and other tubular segments. Ano1-null mice had proteinuria and numerous large reabsorption vesicles in PTE cells. Selective knockout of Ano1 in podocytes (Ano1-/-/Nphs2-Cre) did not impair renal function, whereas tubular knockout in Ano1-/-/Ksp-Cre mice increased urine protein excretion and decreased urine electrolyte concentrations. Purinergic stimulation activated calcium-dependent chloride currents in isolated proximal tubule epithelial cells from wild-type but not from Ano1-/-/Ksp-Cre mice. Ano1 currents were activated by acidic pH, suggesting parallel stimulation of Ano1 chloride secretion with activation of the proton–ATPase. Lack of calcium-dependent chloride secretion in cells from Ano1-/-/Ksp-Cre mice was paralleled by attenuated proton secretion and reduced endosomal acidification, which compromised proximal tubular albumin uptake. Tubular knockout of Ano1 enhanced serum renin and aldosterone concentrations, probably leading to enhanced compensatory distal tubular reabsorption, thus maintaining normal blood pressure levels. Thus, Ano1 has a role in proximal tubular proton secretion and protein reabsorption. The results correspond to regulation of the proton–ATPase by the Ano1-homolog Ist2 in yeast.

Anoctamin 6 (Ano6) belongs to a conserved gene family (TMEM16) predicted to code for eight transmembrane proteins with putative Ca2+-activated chloride channel (CaCC) activity. Recent work revealed that disruption of ANO6 leads to a blood coagulation defect and impaired skeletal development. However, its function in skeletal muscle cells remains to be determined. By using a RNA interference mediated (RNAi) loss-of-function approach, we show that Ano6 regulates C2C12 myoblast proliferation. Ano6 is highly expressed in C2C12 myoblasts and its expression decreases upon differentiation. Knocking down Ano6 significantly reduces C2C12 myoblast proliferation but has minimal effect on differentiation. Ano6 deficiency significantly reduces ERK/AKT phosphorylation, which has been shown to be involved in regulation of cancer cell proliferation by another Anoctamin member. Taken together, our data demonstrate for the first time that Ano6 plays an essential role in C2C12 myoblast proliferation, likely via regulating the ERK/AKT signaling pathway.