Most secretory proteins travel through a well-documented conventional secretion pathway involving the endoplasmic reticulum (ER) and the Golgi complex. However, recently, it has been shown that a ...significant number of proteins reach the plasma membrane or extracellular space via unconventional routes. Unconventional protein secretion (UPS) can be divided into two types: (i) the extracellular secretion of cytosolic proteins that do not bear a signal peptide (i.e. leaderless proteins) and (ii) the cell-surface trafficking of signal-peptide-containing transmembrane proteins via a route that bypasses the Golgi. Understanding the UPS pathways is not only important for elucidating the mechanisms of intracellular trafficking pathways but also has important ramifications for human health, because many of the proteins that are unconventionally secreted by mammalian cells and microorganisms are associated with human diseases, ranging from common inflammatory diseases to the lethal genetic disease of cystic fibrosis. Therefore, it is timely and appropriate to summarize and analyze the mechanisms of UPS involvement in disease pathogenesis, as they may be of use for the development of new therapeutic approaches. In this Review, we discuss the intracellular trafficking pathways of UPS cargos, particularly those related to human diseases. We also outline the disease mechanisms and the therapeutic potentials of new strategies for treating UPS-associated diseases.
Sodium-glucose cotransporter 2 (SGLT2) inhibitors reduce cardiovascular events in humans with type 2 diabetes (T2D); however, the underlying mechanism remains unclear. Activation of the NLR family, ...pyrin domain-containing 3 (NLRP3) inflammasome and subsequent interleukin (IL)-1β release induces atherosclerosis and heart failure. Here we show the effect of SGLT2 inhibitor empagliflozin on NLRP3 inflammasome activity. Patients with T2D and high cardiovascular risk receive SGLT2 inhibitor or sulfonylurea for 30 days, with NLRP3 inflammasome activation analyzed in macrophages. While the SGLT2 inhibitor's glucose-lowering capacity is similar to sulfonylurea, it shows a greater reduction in IL-1β secretion compared to sulfonylurea accompanied by increased serum β-hydroxybutyrate (BHB) and decreased serum insulin. Ex vivo experiments with macrophages verify the inhibitory effects of high BHB and low insulin levels on NLRP3 inflammasome activation. In conclusion, SGLT2 inhibitor attenuates NLRP3 inflammasome activation, which might help to explain its cardioprotective effects.
Lactobacillus is a probiotic with therapeutic potential for several diseases, including liver disease. However, the therapeutic effect of L. plantarum against nonalcoholic steatohepatitis (NASH) and ...its underlying mechanisms remain unelucidated. Therefore, we delineated the L. plantarum-mediated NASH regulation in a mouse model to understand its therapeutic effect. We used a choline-deficient high-fat diet (CD-HFD)-induced murine model that recapitulated the critical features of human metabolic syndrome and investigated the effect of L. plantarum on NASH pathogenesis using transcriptomic, metagenomic, and immunohistochemistry analyses. Validation experiments were performed using liver organoids and a murine model fed a methionine-choline-deficient (MCD) diet. L. plantarum treatment in mice significantly decreased liver inflammation and improved metabolic phenotypes, such as insulin tolerance and the hepatic lipid content, compared with those in the vehicle group. RNA-sequencing analysis revealed that L. plantarum treatment significantly downregulated inflammation-related pathways. Shotgun metagenomic analysis revealed that L-arginine biosynthesis-related microbial genes were significantly upregulated in the L. plantarum group. We also confirmed the elevated arginine levels in the serum of the L. plantarum group. We further used liver organoids and mice fed an MCD diet to demonstrate that L-arginine alone was sufficient to alleviate liver inflammation. Our data revealed a novel and counterintuitive therapeutic effect of L. plantarum on alleviating NASH-related liver inflammation by increasing circulating L-arginine.
Although hundreds of cytosolic or transmembrane molecules form the primary cilium, few secreted molecules are known to contribute to ciliogenesis. Here, homologous secreted metalloproteases ADAMTS9 ...and ADAMTS20 are identified as ciliogenesis regulators that act intracellularly. Secreted and furin-processed ADAMTS9 bound heparan sulfate and was internalized by LRP1, LRP2 and clathrin-mediated endocytosis to be gathered in Rab11 vesicles with a unique periciliary localization defined by super-resolution microscopy. CRISPR-Cas9 inactivation of ADAMTS9 impaired ciliogenesis in RPE-1 cells, which was restored by catalytically active ADAMTS9 or ADAMTS20 acting in trans, but not by their proteolytically inactive mutants. Their mutagenesis in mice impaired neural and yolk sac ciliogenesis, leading to morphogenetic anomalies resulting from impaired hedgehog signaling, which is transduced by primary cilia. In addition to their cognate extracellular proteolytic activity, ADAMTS9 and ADAMTS20 thus have an additional proteolytic role intracellularly, revealing an unexpected regulatory dimension in ciliogenesis.
The most prevalent disease-causing mutation of CFTR is the deletion of Phe508 (ΔF508), which leads to defects in conventional Golgi-mediated exocytosis and cell surface expression. We report that ...ΔF508-CFTR surface expression can be rescued in vitro and in vivo by directing it to an unconventional GRASP-dependent secretion pathway. An integrated molecular and physiological analysis indicates that mechanisms associated with ER stress induce cell surface trafficking of the ER core-glycosylated wild-type and ΔF508-CFTR via the GRASP-dependent pathway. Phosphorylation of a specific site of GRASP and the PDZ-based interaction between GRASP and CFTR are critical for this unconventional surface trafficking. Remarkably, transgenic expression of GRASP in ΔF508-CFTR mice restores CFTR function and rescues mouse survival without apparent toxicity. These findings provide insight into how unconventional protein secretion is activated, and offer a potential therapeutic strategy for the treatment of cystic fibrosis and perhaps diseases stemming from other misfolded proteins.
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► Mechanisms associated with ER stress activate unconventional protein secretion ► The GRASP-dependent pathway traffics core-glycosylated CFTR to the cell surface ► GRASP specifically associates with CFTR by a PDZ-based mechanism ► GRASP rescues the defects of ΔF508-CFTR in HEK293 cells and mice
The common cystic fibrosis mutant of CFTR becomes functional and can reverse disease symptoms in mice when transported to the cell surface through a secretion pathway that bypasses the Golgi.
Autism spectrum disorder (ASD) is a group of conditions characterized by impaired social interaction and communication, and restricted and repetitive behaviours. ASD is a highly heritable disorder ...involving various genetic determinants. Shank2 (also known as ProSAP1) is a multi-domain scaffolding protein and signalling adaptor enriched at excitatory neuronal synapses, and mutations in the human SHANK2 gene have recently been associated with ASD and intellectual disability. Although ASD-associated genes are being increasingly identified and studied using various approaches, including mouse genetics, further efforts are required to delineate important causal mechanisms with the potential for therapeutic application. Here we show that Shank2-mutant (Shank2(-/-)) mice carrying a mutation identical to the ASD-associated microdeletion in the human SHANK2 gene exhibit ASD-like behaviours including reduced social interaction, reduced social communication by ultrasonic vocalizations, and repetitive jumping. These mice show a marked decrease in NMDA (N-methyl-D-aspartate) glutamate receptor (NMDAR) function. Direct stimulation of NMDARs with D-cycloserine, a partial agonist of NMDARs, normalizes NMDAR function and improves social interaction in Shank2(-/-) mice. Furthermore, treatment of Shank2(-/-) mice with a positive allosteric modulator of metabotropic glutamate receptor 5 (mGluR5), which enhances NMDAR function via mGluR5 activation, also normalizes NMDAR function and markedly enhances social interaction. These results suggest that reduced NMDAR function may contribute to the development of ASD-like phenotypes in Shank2(-/-) mice, and mGluR modulation of NMDARs offers a potential strategy to treat ASD.
Celotno besedilo
Dostopno za:
DOBA, IJS, IZUM, KILJ, KISLJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Abstract
Podocyte, the gatekeeper of the glomerular filtration barrier, is a primary target for growth factor and Ca
2+
signaling whose perturbation leads to proteinuria. However, the effects of ...insulin action on store-operated Ca
2+
entry (SOCE) in podocytes remain unknown. Here, we demonstrated that insulin stimulates SOCE by VAMP2-dependent Orai1 trafficking to the plasma membrane. Insulin-activated SOCE triggers actin remodeling and transepithelial albumin leakage via the Ca
2+
-calcineurin pathway in podocytes. Transgenic
Orai1
overexpression in mice causes podocyte fusion and impaired glomerular filtration barrier. Conversely, podocyte-specific
Orai1
deletion prevents insulin-stimulated SOCE, synaptopodin depletion, and proteinuria. Podocyte injury and albuminuria coincide with Orai1 upregulation at the hyperinsulinemic stage in diabetic (
db/db
) mice, which can be ameliorated by the suppression of Orai1-calcineurin signaling. Our results suggest that tightly balanced insulin action targeting podocyte Orai1 is critical for maintaining filter integrity, which provides novel perspectives on therapeutic strategies for proteinuric diseases, including diabetic nephropathy.
Nephrotic syndrome (NS), the association of gross proteinuria, hypoalbuminaemia, edema, and hyperlipidemia, can be clinically divided into steroid-sensitive (SSNS) and steroid-resistant (SRNS) forms. ...SRNS regularly progresses to end-stage renal failure. By homozygosity mapping and whole exome sequencing, we here identify recessive mutations in Crumbs homolog 2 (CRB2) in four different families affected by SRNS. Previously, we established a requirement for zebrafish crb2b, a conserved regulator of epithelial polarity, in podocyte morphogenesis. By characterization of a loss-of-function mutation in zebrafish crb2b, we now show that zebrafish crb2b is required for podocyte foot process arborization, slit diaphragm formation, and proper nephrin trafficking. Furthermore, by complementation experiments in zebrafish, we demonstrate that CRB2 mutations result in loss of function and therefore constitute causative mutations leading to NS in humans. These results implicate defects in podocyte apico-basal polarity in the pathogenesis of NS.
Discriminating between inherited and non-inherited sporadic hearing loss is challenging. Here, we attempted to delineate genetic inheritance in simplex cases of severe-to-profound congenital hearing ...loss in Korean children. Variations in SLC26A4 and GJB2 in 28 children with bilateral severe-to-profound non-syndromic hearing loss (NSHL) without familial history were analyzed using Sanger sequencing. Genetic analysis of individuals without mutations in SLC26A4 and GJB2 was performed by whole exome sequencing (WES). Bi-allelic mutations in SLC26A4 and GJB2 were identified in 12 and 3 subjects, respectively. Of the 13 individuals without mutations in SLC26A4 and GJB2, 2 and 1 carried compound heterozygous mutations in MYO15A and CDH23, respectively. Thus, 64.3% (18/28) of individuals with NSHL were determined to be genetically predisposed. Individuals with sporadic severe-to-profound NSHL were found to mostly exhibit an autosomal recessive inheritance pattern. Novel causative candidate genes for NSHL were identified by analysis of WES data of 10 families without mutations in known causative genes. Bi-allelic mutations predisposing to NSHL were identified in 64.3% of subjects with sporadic severe-to-profound NSHL. Given that several causative genes for NSHL are still unidentified, genetic inheritance of sporadic congenital hearing loss could be more common than that indicated by our results.
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