On the role of dysferlin in striated muscle: membrane repair, t-tubules and Ca2+ handling.

Dysferlin is a 237 kDa membrane-associated protein characterised by multiple C2 domains with a diverse role in skeletal and cardiac muscle physiology. Mutations in DYSF are known to cause various types of human muscular dystrophies, known collectively as dysferlinopathies, with some patients developing cardiomyopathy. A myriad of in vitro membrane repair studies suggest that dysferlin plays an integral role in the membrane repair complex in skeletal muscle. In comparison, less is known about dysferlin in the heart, but mounting evidence suggests that dysferlin's role is similar in both muscle types. Recent findings have shown that dysferlin regulates Ca2+ handling in striated muscle via multiple mechanisms and that this becomes more important in conditions of stress. Maintenance of the transverse (t)-tubule network and the tight coordination of excitation-contraction coupling are essential for muscle contractility. Dysferlin regulates the maintenance and repair of t-tubules, and it is suspected that dysferlin regulates t-tubules and sarcolemmal repair through a similar mechanism. This review focuses on the emerging complexity of dysferlin's activity in striated muscle. Such insights will progress our understanding of the proteins and pathways that regulate basic heart and skeletal muscle function and help guide research into striated muscle pathology, especially that which arises due to dysferlin dysfunction.

Role of mitochondrial dysfunction in kidney disease: Insights from the cGAS-STING signaling pathway.

ABSTRACT: Over the past decade, mitochondrial dysfunction has been investigated as a key contributor to acute and chronic kidney disease. However, the precise molecular mechanisms linking mitochondrial damage to kidney disease remain elusive. The recent insights into the cyclic guanosine monophosphate-adenosine monophosphate (GMP-AMP) synthetase (cGAS)-stimulator of interferon gene (STING) signaling pathway have revealed its involvement in many renal diseases. One of these findings is that mitochondrial DNA (mtDNA) induces inflammatory responses via the cGAS-STING pathway. Herein, we provide an overview of the mechanisms underlying mtDNA release following mitochondrial damage, focusing specifically on the association between mtDNA release-activated cGAS-STING signaling and the development of kidney diseases. Furthermore, we summarize the latest findings of cGAS-STING signaling pathway in cell, with a particular emphasis on its downstream signaling related to kidney diseases. This review intends to enhance our understanding of the intricate relationship among the cGAS-STING pathway, kidney diseases, and mitochondrial dysfunction.

Touch sensation requires the mechanically gated ion channel ELKIN1.

Touch perception is enabled by mechanically activated ion channels, the opening of which excites cutaneous sensory endings to initiate sensation. In this study, we identify ELKIN1 as an ion channel likely gated by mechanical force, necessary for normal touch sensitivity in mice. Touch insensitivity in Elkin1-/- mice was caused by a loss of mechanically activated currents (MA currents) in around half of all sensory neurons activated by light touch (low-threshold mechanoreceptors). Reintroduction of Elkin1 into sensory neurons from Elkin1-/- mice restored MA currents. Additionally, small interfering RNA-mediated knockdown of ELKIN1 from induced human sensory neurons substantially reduced indentation-induced MA currents, supporting a conserved role for ELKIN1 in human touch. Our data identify ELKIN1 as a core component of touch transduction in mice and potentially in humans.

Structure, Function, and Regulation of the Junctophilin Family.

In both excitable and nonexcitable cells, diverse physiological processes are linked to different calcium microdomains within nanoscale junctions that form between the plasma membrane and endo-sarcoplasmic reticula. It is now appreciated that the junctophilin protein family is responsible for establishing, maintaining, and modulating the structure and function of these junctions. We review foundational findings from more than two decades of research that have uncovered how junctophilin-organized ultrastructural domains regulate evolutionarily conserved biological processes. We discuss what is known about the junctophilin family of proteins. Our goal is to summarize the current knowledge of junctophilin domain structure, function, and regulation and to highlight emerging avenues of research that help our understanding of the transcriptional, translational, and post-translational regulation of this gene family and its roles in health and during disease.

SCRIB controls apical contractility during epithelial differentiation.

Although mutations in the SCRIB gene lead to multiple morphological organ defects in vertebrates, the molecular pathway linking SCRIB to organ shape anomalies remains elusive. Here, we study the impact of SCRIB-targeted gene mutations during the formation of the gut epithelium in an organ-on-chip model. We show that SCRIB KO gut-like epithelia are flatter with reduced exposed surface area. Cell differentiation on filters further shows that SCRIB plays a critical role in the control of apical cell shape, as well as in the basoapical polarization of myosin light chain localization and activity. Finally, we show that SCRIB serves as a molecular scaffold for SHROOM2/4 and ROCK1 and identify an evolutionary conserved SHROOM binding site in the SCRIB carboxy-terminal that is required for SCRIB function in the control of apical cell shape. Our results demonstrate that SCRIB plays a key role in epithelial morphogenesis by controlling the epithelial apical contractility during cell differentiation.

A MEC-2/stomatin condensate liquid-to-solid phase transition controls neuronal mechanotransduction during touch sensing.

A growing body of work suggests that the material properties of biomolecular condensates ensuing from liquid-liquid phase separation change with time. How this aging process is controlled and whether the condensates with distinct material properties can have different biological functions is currently unknown. Using Caenorhabditis elegans as a model, we show that MEC-2/stomatin undergoes a rigidity phase transition from fluid-like to solid-like condensates that facilitate transport and mechanotransduction, respectively. This switch is triggered by the interaction between the SH3 domain of UNC-89 (titin/obscurin) and MEC-2. We suggest that this rigidity phase transition has a physiological role in frequency-dependent force transmission in mechanosensitive neurons during body wall touch. Our data demonstrate a function for the liquid and solid phases of MEC-2/stomatin condensates in facilitating transport or mechanotransduction, and a previously unidentified role for titin homologues in neurons.

Cis-inhibition suppresses basal Notch signaling during sensory organ precursor selection.

The emergence of the sensory organ precursor (SOP) from an equivalence group in Drosophila is a paradigm for studying single-cell fate specification through Notch-mediated lateral inhibition. Yet, it remains unclear how only a single SOP is selected from a relatively large group of cells. We show here that a critical aspect of SOP selection is controlled by cis-inhibition (CI), whereby the Notch ligands, Delta (Dl), cis-inhibit Notch receptors in the same cell. Based on the observation that the mammalian ligand Dl-like 1 cannot cis-inhibit Notch in Drosophila, we probe the role of CI in vivo. We develop a mathematical model for SOP selection where Dl activity is independently regulated by the ubiquitin ligases Neuralized and Mindbomb1. We show theoretically and experimentally that Mindbomb1 induces basal Notch activity, which is suppressed by CI. Our results highlight the trade-off between basal Notch activity and CI as a mechanism for singling out a SOP from a large equivalence group.

Revisiting the Role of CD63 as Pro-Tumorigenic or Anti-Tumorigenic Tetraspanin in Cancers and its Theragnostic Implications.

Cluster of differentiation antigen 63 (CD63) belongs to a superfamily of proteins, usually defined as tetraspanins which are known to transverse the bilayer membranes four times. The expression of CD63 has been shown to get altered in several cancers, where it has been demonstrated to act as both a tumor promoter and tumor suppressor. The present review describes the mechanism of how CD63 promotes tumor formation in certain cancer types while inhibiting in some other specific cancers. Glycosylation, a post-translational process plays a significant role in regulating the expression and function of these membrane proteins. Being a crucial exosomal flag protein, CD63 has been found to get involved in endosomal cargo sorting as well as the production of extracellular vesicles. Increased expression of exosomal CD63 derived from advanced tumors has demonstrated its role in promoting metastasis. CD63 also regulates the characteristic and function of stem cells on which they get expressed. This particular tetraspanin has been discovered to participate in gene fusion to perform distinctive roles in certain specific cancer types like breast cancer and pigmented epithelioid melanocytoma. Furthermore, this review mentions twelve different microRNAs obtained from miRDB that might target CD63. A few theragnostic uses of this membrane protein are also discussed. Thereby, the review indicates that further studies on CD63 might prove it to be an effective therapeutic target in different cancers in the coming future.

Recruitment of the SNX17-Retriever recycling pathway regulates synaptic function and plasticity.

Trafficking of cell-surface proteins from endosomes to the plasma membrane is a key mechanism to regulate synaptic function. In non-neuronal cells, proteins recycle to the plasma membrane either via the SNX27-Retromer-WASH pathway or via the recently discovered SNX17-Retriever-CCC-WASH pathway. While SNX27 is responsible for the recycling of key neuronal receptors, the roles of SNX17 in neurons are less understood. Here, using cultured hippocampal neurons, we demonstrate that the SNX17 pathway regulates synaptic function and plasticity. Disruption of this pathway results in a loss of excitatory synapses and prevents structural plasticity during chemical long-term potentiation (cLTP). cLTP drives SNX17 recruitment to synapses, where its roles are in part mediated by regulating the surface expression of ß1-integrin. SNX17 recruitment relies on NMDAR activation, CaMKII signaling, and requires binding to the Retriever and PI(3)P. Together, these findings provide molecular insights into the regulation of SNX17 at synapses and define key roles for SNX17 in synaptic maintenance and in regulating enduring forms of synaptic plasticity.

Cell surface protein aggregation triggers endocytosis to maintain plasma membrane proteostasis.

The ability of cells to manage consequences of exogenous proteotoxicity is key to cellular homeostasis. While a plethora of well-characterised machinery aids intracellular proteostasis, mechanisms involved in the response to denaturation of extracellular proteins remain elusive. Here we show that aggregation of protein ectodomains triggers their endocytosis via a macroendocytic route, and subsequent lysosomal degradation. Using ERBB2/HER2-specific antibodies we reveal that their cross-linking ability triggers specific and fast endocytosis of the receptor, independent of clathrin and dynamin. Upon aggregation, canonical clathrin-dependent cargoes are redirected into the aggregation-dependent endocytosis (ADE) pathway. ADE is an actin-driven process, which morphologically resembles macropinocytosis. Physical and chemical stress-induced aggregation of surface proteins also triggers ADE, facilitating their degradation in the lysosome. This study pinpoints aggregation of extracellular domains as a trigger for rapid uptake and lysosomal clearance which besides its proteostatic function has potential implications for the uptake of pathological protein aggregates and antibody-based therapies.

Linked Chemotype Cellular Uptake Is Facilitated by IFITMs.

Interferon-induced transmembrane proteins (IFITM) modulate cell permeability of diverse linked chemotypes.

Endoplasmic Reticulum-Plasma Membrane Junctions as Sites of Depolarization-Induced Ca2+ Signaling in Excitable Cells.

Membrane contact sites between endoplasmic reticulum (ER) and plasma membrane (PM), or ER-PM junctions, are found in all eukaryotic cells. In excitable cells they play unique roles in organizing diverse forms of Ca2+ signaling as triggered by membrane depolarization. ER-PM junctions underlie crucial physiological processes such as excitation-contraction coupling, smooth muscle contraction and relaxation, and various forms of activity-dependent signaling and plasticity in neurons. In many cases the structure and molecular composition of ER-PM junctions in excitable cells comprise important regulatory feedback loops linking depolarization-induced Ca2+ signaling at these sites to the regulation of membrane potential. Here, we describe recent findings on physiological roles and molecular composition of native ER-PM junctions in excitable cells. We focus on recent studies that provide new insights into canonical forms of depolarization-induced Ca2+ signaling occurring at junctional triads and dyads of striated muscle, as well as the diversity of ER-PM junctions in these cells and in smooth muscle and neurons.

Reconstitution of the S. aureus agr quorum sensing pathway reveals a direct role for the integral membrane protease MroQ in pheromone biosynthesis.

In Staphylococcus aureus, virulence is under the control of a quorum sensing (QS) circuit encoded in the accessory gene regulator (agr) genomic locus. Key to this pathogenic behavior is the production and signaling activity of a secreted pheromone, the autoinducing peptide (AIP), generated following the ribosomal synthesis and posttranslational modification of a precursor polypeptide, AgrD, through two discrete cleavage steps. The integral membrane protease AgrB is known to catalyze the first processing event, generating the AIP biosynthetic intermediate, AgrD (1-32) thiolactone. However, the identity of the second protease in this biosynthetic pathway, which removes an N-terminal leader sequence, has remained ambiguous. Here, we show that membrane protease regulator of agr QS (MroQ), an integral membrane protease recently implicated in the agr response, is directly involved in AIP production. Genetic complementation and biochemical experiments reveal that MroQ proteolytic activity is required for AIP biosynthesis in agr specificity group I and group II, but not group III. Notably, as part of this effort, the biosynthesis and AIP-sensing arms of the QS circuit were reconstituted together in vitro. Our experiments also reveal the molecular features guiding MroQ cleavage activity, a critical factor in defining agr specificity group identity. Collectively, our study adds to the molecular understanding of the agr response and Staphylococcus aureus virulence.

The fliR gene contributes to the virulence of S. marcescens in a Drosophila intestinal infection model.

Serratia marcescens is an opportunistic bacterium that infects a wide range of hosts including humans. It is a potent pathogen in a septic injury model of Drosophila melanogaster since a few bacteria directly injected in the body cavity kill the insect within a day. In contrast, flies do not succumb to ingested bacteria for days even though some bacteria cross the intestinal barrier into the hemolymph within hours. The mechanisms by which S. marcescens attacks enterocytes and damages the intestinal epithelium remain uncharacterized. To better understand intestinal infections, we performed a genetic screen for loss of virulence of ingested S. marcescens and identified FliR, a structural component of the flagellum, as a virulence factor. Next, we compared the virulence of two flagellum mutants fliR and flhD in two distinct S. marcescens strains. Both genes are required for S. marcescens to escape the gut lumen into the hemocoel, indicating that the flagellum plays an important role for the passage of bacteria through the intestinal barrier. Unexpectedly, fliR but not flhD is involved in S. marcescens-mediated damages of the intestinal epithelium that ultimately contribute to the demise of the host. Our results therefore suggest a flagellum-independent role for fliR in bacterial virulence.

A Variety of Nucleic Acid Species Are Sensed by cGAS, Implications for Its Diverse Functions.

Cyclic GMP-AMP synthase (cGAS) recognizes double-stranded DNA (dsDNA) derived from invading pathogens and induces an interferon response via activation of the key downstream adaptor protein stimulator of interferon genes (STING). This is the most classic biological function of the cGAS-STING signaling pathway and is critical for preventing pathogenic microorganism invasion. In addition, cGAS can interact with various types of nucleic acids, including cDNA, DNA : RNA hybrids, and circular RNA, to contribute to a diverse set of biological functions. An increasing number of studies have revealed an important relationship between the cGAS-STING signaling pathway and autophagy, cellular senescence, antitumor immunity, inflammation, and autoimmune diseases. This review details the mechanism of action of cGAS as it interacts with different types of nucleic acids, its rich biological functions, and the potential for targeting this pathway to treat various diseases.

Lamina-associated polypeptide 2α is required for intranuclear MRTF-A activity.

Myocardin-related transcription factor A (MRTF-A), a coactivator of serum response factor (SRF), regulates the expression of many cytoskeletal genes in response to cytoplasmic and nuclear actin dynamics. Here we describe a novel mechanism to regulate MRTF-A activity within the nucleus by showing that lamina-associated polypeptide 2α (Lap2α), the nucleoplasmic isoform of Lap2, is a direct binding partner of MRTF-A, and required for the efficient expression of MRTF-A/SRF target genes. Mechanistically, Lap2α is not required for MRTF-A nuclear localization, unlike most other MRTF-A regulators, but is required for efficient recruitment of MRTF-A to its target genes. This regulatory step takes place prior to MRTF-A chromatin binding, because Lap2α neither interacts with, nor specifically influences active histone marks on MRTF-A/SRF target genes. Phenotypically, Lap2α is required for serum-induced cell migration, and deregulated MRTF-A activity may also contribute to muscle and proliferation phenotypes associated with loss of Lap2α. Our studies therefore add another regulatory layer to the control of MRTF-A-SRF-mediated gene expression, and broaden the role of Lap2α in transcriptional regulation.

CD4 T cell-intrinsic STING signaling controls the differentiation and effector functions of TH1 and TH9 cells.

BACKGROUND: While stimulator of interferon genes (STING) activation in innate immune cells of the tumor microenvironment can result in CD8 T cell-dependent antitumor immunity, whether STING signaling affects CD4 T-cell responses remains elusive. METHODS: Here, we tested whether STING activation modulated the effector functions of CD4 T cells in vivo by analyzing tumor-infiltrating CD4 T cells and evaluating the contribution of the CD4 T cell-derived cytokines in the antitumor activity of the STING ligand 2'3'-cyclic guanosine monophosphate-adenosine monophosphate (cGAMP) in two mouse tumor models. We performed ex vivo experiments to assess the impact of STING activation on CD4 T-cell differentiation and investigate the underlying molecular mechanisms. Finally, we tested whether STING activation enhances TH9 cell antitumor activity against mouse melanoma upon adoptive transfer. RESULTS: We found that activation of STING signaling cell-intrinsically enhances the differentiation and antitumor functions of TH1 and TH9 cells by increasing their respective production of interferon gamma (IFN-γ) and interleukin-9. IRF3 and type I interferon receptors (IFNARs) are required for the STING-driven enhancement of TH1 cell differentiation. However, STING activation favors TH9 cell differentiation independently of the IFNARs/IRF3 pathway but through mammalian target of rapamycin (mTOR) signaling, underscoring that STING activation differentially affects the fate of distinct CD4 T-cell subsets. The therapeutic effect of STING activation relies on TH1 and TH9-derived cytokines, and STING activation enhances the antitumor activity of TH9 cells upon adoptive transfer. CONCLUSION: Our results reveal the STING signaling pathway as a therapeutic target to boost CD4 T-cell effector functions and antitumor immunity.

Depression-like symptoms due to Dcf1 deficiency are alleviated by intestinal transplantation of Lactobacillus murine and Lactobacillus reuteri.

Depression, characterized by low mood, is a complex mental disorder that is a serious threat to human health. Depression is thought to be caused by a combination of genetic, environmental and psychological factors. However, the pathophysiology of depression remains unclear. In the present study, we found that Dcf1 knockout (KO) mice had depression-like symptoms and disruptive changes in gamma-aminobutyric acid (GABA) concentration and GABA receptor expression were found in the hippocampus of Dcf1 KO and WT mice. Furthermore, the gut microbiota composition of Dcf1 KO mice was significantly different from that of wildtype (WT) mice and Dcf1 KO mice showed lower Firmicutes and Lactobacillus content compared to WT mice. In addition, the depression-like behavior of Dcf1 KO mice was alleviated by the administration of microbiota. More surprisingly, after treatment with Lactobacillus murine and Lactobacillus reuteri, two Lactobacillus species with proportionally greater differences in content between the WT and KO groups, KO mice showed similar GABA content, as well as restored GABA-related receptor expression, as the WT group. Our data elucidated a possible mechanism of depression induction by gut microbiota in Dcf1 KO mice and provide a new avenue to explore the treatment of depression by gut microbiota.

Proline-rich transmembrane protein 2 specifically binds to GluA1 but has no effect on AMPA receptor-mediated synaptic transmission.

BACKGROUND: Proline-rich transmembrane protein 2 (PRRT2) is a neuron-specific protein associated with seizures, dyskinesia, and intelligence deficit. Previous studies indicate that PRRT2 regulates neurotransmitter release from presynaptic membranes. However, PRRT2 can also bind AMPA-type glutamate receptors (AMPARs), but its postsynaptic functions remain unclear. METHODS AND RESULTS: Whole-exome sequencing used to diagnose a patient with mental retardation identified a nonsense mutation in the PRRT2 gene (c.649C>T; p.R217X). To understand the pathology of the mutant, we cloned mouse Prrt2 cDNA and inserted a premature stop mutation at Arg223, the corresponding site of Arg217 in human PRRT2. In mouse hippocampal tissues, Prrt2 interacted with GluA1/A2 AMPAR heteromers but not GluA2/A3s, via binding to GluA1. Additionally, Prrt2 suppressed GluA1 expression and localization on cell membranes of HEK 293T cells. However, when Prrt2 was overexpressed in individual hippocampal neurons using in utero electroporation, AMPAR-mediated synaptic transmission was unaffected. Deletion of Prrt2 with the CRIPR/Cas9 technique did not affect AMPAR-mediated synaptic transmission. Furthermore, deletion or overexpression of Prrt2 did not affect GluA1 expression and distribution in primary neuronal culture. CONCLUSIONS: The postsynaptic functions of Prrt2 demonstrate that Prrt2 specifically interacts with the AMPAR subunit GluA1 but does not regulate AMPAR-mediated synaptic transmission. Therefore, our study experimentally excluded a postsynaptic regulatory mechanism of Prrt2. The pathology of PRRT2 variants in humans likely originates from defects in neurotransmitter release from the presynaptic membrane as suggested by recent studies.