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Balwant S. Tuana - One of the best experts on this subject based on the ideXlab platform.
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Abstract 235: Sarcolemmal Membrane Associated Protein Isoform 1: a Unique Regulator of Glucose Uptake and Metabolism in the Myocardium
Circulation Research, 2015Co-Authors: Aaraf Dewan, Maysoon Salih, Chris R. Triggle, Hong Ding, Balwant S. TuanaAbstract:As one of the leading causes of heart disease, diabetes is a problem which needs a solution. Regulation of glucose uptake and metabolism within skeletal and cardiac muscle has proven capable of altering systemic glucose levels and impacting metabolism to potentially improve patient outcomes. Unfortunately, to date, very few muscle specific metabolic regulators are known which can allow us to achieve blood glucose uptake and metabolism. Sarcolemmal Membrane Associated Protein Isoform 1 (SLMAP1) is a novel protein expressed predominantly within muscle tissue. It has been linked to diabetes through animal models, although its role in metabolism remains to be defined. Here we describe a novel role for SLMAP1 in glucose metabolism within the myocardium. We engineered a transgenic (TG) mouse with cardiac specific expression of SLMAP1. Using neonatal cardiomyocytes (NCMs) collected from these mice we performed glucose uptake assays with 2-deoxy-glucose (2DG), measured glycolytic rate using an Extracellular Flux ...
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abstract 400 regulated levels of Sarcolemmal Membrane protein isoform 3 impact cardiac function in mice
Circulation Research, 2015Co-Authors: Jana Mlynarova, Maysoon Salih, Mayra Trentinsonoda, Fernanda Gaisler Da Silva, Jennifer L Major, Marcela Sorelli Carneiroramos, Balwant S. TuanaAbstract:The Sarcolemmal Membrane-associated proteins (SLMAPs) are a family of tail-anchored Membrane proteins generated by alternative splicing of the SLMAP gene. A ubiquitously expressed SLMAP isoform 3 encompasses an N-terminal FHA domain with extended coiled-coil structure and has been implicated in cell cycle control. Heart function in transgenic mice with cardiac-specific overexpression of SLMAP3 cDNA driven by α myosin heavy chain promoter was evaluated by echocardiography. qPCR and western blot were used to analyze gene and protein expression respectively. Structure and fibrosis was analyzed by H&E and Masson’s Trichrome staining. Function analysis showed a 15% (p<0.05) decrease in ejection fraction and 19% (p<0.05) decrease in fractional shortening in transgenic mice as early as 5 weeks and persisted into old age at 44 weeks. Transgenic mice presented a mild systolic dysfunction and a trend towards dilated cardiomyopathy without any premature death. Natriuretic peptide ANP and BNP levels were not changed and there was no difference in left ventricular mass or activation of the hypertrophic factor Akt1 in SLMAP3 expressing myocardium. However, significant changes in calcium handling proteins with a significant decrease in phosphorylation of phospholamban ser16 (p<0.05) along with a down-regulation of sarco-endoplasmic reticulum Ca2+ ATPase protein and increased ryanodine receptor 2 phosphorylation ser2808 (p<0.05) were noted at 5 weeks of age in transgenic hearts. These data indicate that increased SLMAP3 levels did not influence cardiac remodeling or hypertrophic growth but did impact Membrane biology of calcium transport systems in myocardium leading to depressed contractility. Thus regulated levels of SLMAP3 are important to support normal heart function.
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abstract 2281 the drosophila Sarcolemmal Membrane associated protein dslmap is a novel regulator of cardiac development and interacts with the slit robo pathway
Circulation, 2009Co-Authors: Danya Alhyari, Momena Dawood, Margaret Sonnenfeld, Balwant S. TuanaAbstract:The Drosophila heart is composed of myocardial and pericardial cells. Together, these cells form a linear heart tube that resembles the early vertebrate heart. Many genes involved in heart development are conserved from fruit flies to mammals. For example, the Drosophila homeobox gene Tinman and its vertebrate homologue Nkx2.5 are both critical for heart development. The extracellular protein Slit and its receptor Roundabout (Robo) contribute to heart tube formation by guiding myocardial cell alignment and adhesion. The Slit/Robo complex appears to serve conserved roles across species and in different systems by influencing cell guidance and migration. Previous studies have shown that Sarcolemmal Membrane associated protein (SLMAP) is expressed early during mammalian cardiac development and that SLMAP homolog in Drosophila interacts with Slit/Robo pathway in the developing nervous system. Here, we show that DSLMAP is expressed in both myocardial and pericardial cells at early stages of Drosophila heart development. Alterations in DSLMAP expression in myocardial cells using a UAS- DSLMAP -RNAi transgene causes a delay in heart tube development and misalignment of the cardiac cells. This phenotype is similar to that observed in robo down-regulation in myocardial cells. Cell migration defects are enhanced by myocardial expression of UAS- DSLMAP RNAi. The up-regulation of DSLMAP in myocardial cells had phenotypes similar to those seen with Slit over-expression as characterized by misalignment and gaps between myocardial cells. Slit over-expression suppresses UAS- DSLMAP RNAi associated heart defect. Our data suggests that DSLMAP is expressed early in cardiac morphogenesis during Drosophila heart development and interacts with the Slit and Robo pathway to regulate cardiac cell migration and alignment
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regulated expression and temporal induction of the tail anchored Sarcolemmal Membrane associated protein is critical for myoblast fusion
Biochemical Journal, 2004Co-Authors: Rosa M Guzzo, Maysoon Salih, Jeffery Wigle, Edwin D W Moore, Balwant S. TuanaAbstract:Sarcolemmal-Membrane-associated proteins (SLMAPs) define a new class of coiled-coil tail-anchored Membrane proteins generated by alternative splicing mechanisms. An in vivo expression analysis indicated that SLMAPs are present in somites (11 days post-coitum) as well as in fusing myotubes and reside at the level of the sarcoplasmic reticulum and transverse tubules in adult skeletal muscles. Skeletal-muscle myoblasts were found to express a single 5.9 kb transcript, which encodes the full-length ∼91 kDa SLMAP3 isoform. Myoblast differentiation was accompanied by the stable expression of the ∼91 kDa SLMAP protein as well as the appearance of an ∼80 kDa isoform. Deregulation of SLMAPs by ectopic expression in myoblasts resulted in a potent inhibition of fusion without affecting the expression of muscle-specific genes. Membrane targeting of the de-regulated SLMAPs was not critical for the inhibition of myotube development. Protein–protein interaction assays indicated that SLMAPs are capable of self-assembling, and the de-regulated expression of mutants that were not capable of forming SLMAP homodimers also inhibited myotube formation. These results imply that regulated levels and the temporal induction of SLMAP isoforms are important for normal muscle development.
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a novel isoform of Sarcolemmal Membrane associated protein slmap is a component of the microtubule organizing centre
Journal of Cell Science, 2004Co-Authors: Rosa M Guzzo, Maysoon Salih, Serdal Sevinc, Balwant S. TuanaAbstract:The microtubule organizing centre (MTOC) or the centrosome serves a crucial role in the establishment of cellular polarity, organization of interphase microtubules and the formation of the bipolar mitotic spindle. We have elucidated the genomic structure of a gene encoding the Sarcolemmal Membrane-associated protein (SLMAP), which encodes a 91 kDa polypeptide with a previously uncharacterized N-terminal sequence encompassing a forkhead-associated (FHA) domain that resides at the centrosome. Anti-peptide antibodies directed against SLMAP N-terminal sequences showed colocalization with γ-tubulin at the centrosomes at all phases of the cell cycle. Agents that specifically disrupt microtubules did not affect SLMAP association with centrosomes. Furthermore, SLMAP sequences directed a reporter green fluorescent protein (GFP) to the centrosome, and deletions of the newly identified N-terminal sequence from SLMAP prevented the centrosomal targeting. Deletion-mutant analysis concluded that overall, structural determinants in SLMAP were responsible for centrosomal targeting. Elevated levels of centrosomal SLMAP were found to be lethal, whereas mutants that lacked centrosomal targeting inhibited cell growth accompanied by an accumulation of cells at the G2/M phase of the cell cycle.
Shuanghui Wang - One of the best experts on this subject based on the ideXlab platform.
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pinellia pedatisecta agglutinin targets drug resistant k562 adr leukemia cells through binding with Sarcolemmal Membrane associated protein and enhancing macrophage phagocytosis
PLOS ONE, 2013Co-Authors: Kan Chen, Xinyan Yang, Shuanghui WangAbstract:Pinellia pedatisecta agglutinin (PPA) has previously been used in labeling fractions of myeloid leukemia cells in our laboratory. We report here that a bacterial expressed recombinant PPA domain b tagged with soluble coxsackie and adenovirus receptor (sCAR-PPAb) preferentially recognized drug resistant cancer cells K562/ADR and H460/5Fu, as compared to their parental cell lines. Pretreatment of K562/ADR cells with sCAR-PPAb significantly enhanced phagocytosis of K562/ADR by macrophages in vivo. Meanwhile, in a K562/ADR xenograft model, intratumoral injection of sCAR-PPAb induced macrophage infiltration and phagocytosis. Furthermore, immunoprecipitation, mass spectrometry and Western blot identified the Membrane target of PPA on K562/ADR as Sarcolemmal Membrane associated protein (SLMAP). An antibody against SLMAP significantly promoted the phagocytosis of K562/ADR by macrophages in vitro. These findings suggest that PPA not only could be developed into a novel agent that can detect drug resistant cancer cells and predict chemotherapy outcome, but also it has potential value in immunotherapy against drug resistant cancer cells through inducing the tumoricidal activity of macrophages.
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Pinellia pedatisecta agglutinin targets drug resistant K562/ADR leukemia cells through binding with Sarcolemmal Membrane associated protein and enhancing macrophage phagocytosis
2013Co-Authors: Kan Chen, Xinyan Yang, Shuanghui WangAbstract:Pinellia pedatisecta agglutinin (PPA) has previously been used in labeling fractions of myeloid leukemia cells in our laboratory. We report here that a bacterial expressed recombinant PPA domain b tagged with soluble coxsackie and adenovirus receptor (sCAR-PPAb) preferentially recognized drug resistant cancer cells K562/ADR and H460/5Fu, as compared to their parental cell lines. Pretreatment of K562/ADR cells with sCAR-PPAb significantly enhanced phagocytosis of K562/ADR by macrophages in vivo. Meanwhile, in a K562/ADR xenograft model, intratumoral injection of sCAR-PPAb induced macrophage infiltration and phagocytosis. Furthermore, immunoprecipitation, mass spectrometry and Western blot identified the Membrane target of PPA on K562/ADR as Sarcolemmal Membrane associated protein (SLMAP). An antibody against SLMAP significantly promoted the phagocytosis of K562/ADR by macrophages in vitro. These findings suggest that PPA not only could be developed into a novel agent that can detect drug resistant cancer cells and predict chemotherapy outcome, but also it has potential value in immunotherapy against drug resistant cancer cells through inducing the tumoricidal activity of macrophages
Kan Chen - One of the best experts on this subject based on the ideXlab platform.
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pinellia pedatisecta agglutinin targets drug resistant k562 adr leukemia cells through binding with Sarcolemmal Membrane associated protein and enhancing macrophage phagocytosis
PLOS ONE, 2013Co-Authors: Kan Chen, Xinyan Yang, Shuanghui WangAbstract:Pinellia pedatisecta agglutinin (PPA) has previously been used in labeling fractions of myeloid leukemia cells in our laboratory. We report here that a bacterial expressed recombinant PPA domain b tagged with soluble coxsackie and adenovirus receptor (sCAR-PPAb) preferentially recognized drug resistant cancer cells K562/ADR and H460/5Fu, as compared to their parental cell lines. Pretreatment of K562/ADR cells with sCAR-PPAb significantly enhanced phagocytosis of K562/ADR by macrophages in vivo. Meanwhile, in a K562/ADR xenograft model, intratumoral injection of sCAR-PPAb induced macrophage infiltration and phagocytosis. Furthermore, immunoprecipitation, mass spectrometry and Western blot identified the Membrane target of PPA on K562/ADR as Sarcolemmal Membrane associated protein (SLMAP). An antibody against SLMAP significantly promoted the phagocytosis of K562/ADR by macrophages in vitro. These findings suggest that PPA not only could be developed into a novel agent that can detect drug resistant cancer cells and predict chemotherapy outcome, but also it has potential value in immunotherapy against drug resistant cancer cells through inducing the tumoricidal activity of macrophages.
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Pinellia pedatisecta agglutinin targets drug resistant K562/ADR leukemia cells through binding with Sarcolemmal Membrane associated protein and enhancing macrophage phagocytosis
2013Co-Authors: Kan Chen, Xinyan Yang, Shuanghui WangAbstract:Pinellia pedatisecta agglutinin (PPA) has previously been used in labeling fractions of myeloid leukemia cells in our laboratory. We report here that a bacterial expressed recombinant PPA domain b tagged with soluble coxsackie and adenovirus receptor (sCAR-PPAb) preferentially recognized drug resistant cancer cells K562/ADR and H460/5Fu, as compared to their parental cell lines. Pretreatment of K562/ADR cells with sCAR-PPAb significantly enhanced phagocytosis of K562/ADR by macrophages in vivo. Meanwhile, in a K562/ADR xenograft model, intratumoral injection of sCAR-PPAb induced macrophage infiltration and phagocytosis. Furthermore, immunoprecipitation, mass spectrometry and Western blot identified the Membrane target of PPA on K562/ADR as Sarcolemmal Membrane associated protein (SLMAP). An antibody against SLMAP significantly promoted the phagocytosis of K562/ADR by macrophages in vitro. These findings suggest that PPA not only could be developed into a novel agent that can detect drug resistant cancer cells and predict chemotherapy outcome, but also it has potential value in immunotherapy against drug resistant cancer cells through inducing the tumoricidal activity of macrophages
Noah Weisleder - One of the best experts on this subject based on the ideXlab platform.
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Abstract 218: The Role of Novel Tripartite Motif Proteins in Sarcolemmal Membrane Repair
Circulation Research, 2014Co-Authors: Liubov V. Gushchina, Eric X Beck, Jenna Alloush, Sayak Bhattacharya, Noah WeislederAbstract:Tripartite motif (TRIM) proteins are a superfamily of coiled-coil-containing RING E3 ligases that function in many cellular processes, particularly in Membrane repair pathways. Mitsugumin 53 (MG53) also known as TRIM72, is primary expressed in skeletal muscle and heart. Our experimental data confirm that during Membrane damage, MG53 translocates to the injury site and acts as a molecular glue to reseal the damage area. The role of MG53 in Membrane repair has been demonstrated in both in vitro studies using molecular approaches and in vivo using rodent wild type and knockout models. Thus, our data indicate that recombinant human MG53 protein can be directly applied as a therapeutic agent to increase the Membrane repair capacity of many cell types, including cardiomyocytes during acute injury or in chronic disease progression. However, the precise mechanism and potential partners by which MG53 executes its Membrane repair function are not completely understood. On the basis of the global TRIM family protein alignment, we hypothesize that there are other TRIM proteins that, alone or together with MG53, may facilitate repair by targeting the site of an injury. Moreover, data from our lab demonstrated that MG53 and these TRIM proteins can form homo- and hetero-oligomeric assemblies due to the presence of the coiled-coil region in these proteins and, further, that this may be necessary for the active Membrane resealing process. Using E. coli protein expression methodology we can generate and isolate new TRIM recombinant proteins and test if these protein complexes are effective when applied externally to cardiac and non-cardiac cells. These novel proteins will also be tested for their pharmacokinetic properties to determine their efficacy in both acute and chronic applications. Our studies should increase our knowledge of the mechanisms controlling cardiac Membrane repair and also provide novel therapeutic targets.
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expression levels of Sarcolemmal Membrane repair proteins following prolonged exercise training in mice
Indian Journal of Biochemistry & Biophysics, 2013Co-Authors: Jenna Alloush, Steve R Roof, Eric X Beck, Mark T Ziolo, Noah WeislederAbstract:Membrane repair is a conserved cellular process, where intracellular vesicles translocate to sites of plasma Membrane injury to actively reseal Membrane disruptions. Such Membrane disruptions commonly occur in the course of normal physiology, particularly in skeletal muscles due to repeated contraction producing small tears in the Sarcolemmal Membrane. Here, we investigated whether prolonged exercise could produce adaptive changes in expression levels of proteins associated with the Membrane repair process, including mitsugumin 53/tripartite motif-containing protein 72 (MG53/TRIM72), dysferlin and caveolin-3 (cav3). Mice were exercised using a treadmill running protocol and protein levels were measured by immunoblotting. The specificity of the antibodies used was established by immunoblot testing of various tissue lysates from both mice and rats. We found that MG53/TRIM72 immunostaining on isolated mouse skeletal muscle fibers showed protein localization at sites of Membrane disruption created by the isolation of these muscle fibers. However, no significant changes in the expression levels of the tested Membrane repair proteins were observed following prolonged treadmill running for eight weeks (30 to 80 min/day). These findings suggest that any compensation occurring in the Membrane repair process in skeletal muscle following prolonged exercise does not affect the expression levels of these three key Membrane repair proteins.
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enhancing muscle Membrane repair by gene delivery of mg53 ameliorates muscular dystrophy and heart failure in δ sarcoglycan deficient hamsters
Molecular Therapy, 2012Co-Authors: Ruhang Tang, Noah Weisleder, Bin Xiao, Zhenhua Yuan, Chuanxi Cai, Hua Zhu, Peihui Lin, Chunping Qiao, Christina Mayer, Xiao XiaoAbstract:Muscular dystrophies (MDs) are caused by genetic mutations in over 30 different genes, many of which encode for proteins essential for the integrity of muscle cell structure and Membrane. Their deficiencies cause the muscle vulnerable to mechanical and biochemical damages, leading to Membrane leakage, dystrophic pathology, and eventual loss of muscle cells. Recent studies report that MG53, a muscle-specific TRIM-family protein, plays an essential role in Sarcolemmal Membrane repair. Here, we show that systemic delivery and muscle-specific overexpression of human MG53 gene by recombinant adeno-associated virus (AAV) vectors enhanced Membrane repair, ameliorated pathology, and improved muscle and heart functions in δ-sarcoglycan (δ-SG)-deficient TO-2 hamsters, an animal model of MD and congestive heart failure. In addition, MG53 overexpression increased dysferlin level and facilitated its trafficking to muscle Membrane through participation of caveolin-3. MG53 also protected muscle cells by activating cell survival kinases, such as Akt, extracellular signal-regulated kinases (ERK1/2), and glycogen synthase kinase-3β (GSK-3β) and inhibiting proapoptotic protein Bax. Our results suggest that enhancing the muscle Membrane repair machinery could be a novel therapeutic approach for MD and cardiomyopathy, as demonstrated here in the limb girdle MD (LGMD) 2F model.
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detection of calcium sparks in intact and permeabilized skeletal muscle fibers
Methods of Molecular Biology, 2012Co-Authors: Noah Weisleder, Jingsong ZhouAbstract:Ca(2+) sparks are the elementary units of Ca(2+) signaling in striated muscle fibers that appear as highly localized Ca(2+) release events through ryanodine receptor (RyR) Ca(2+) release channels in the sarcoplasmic reticulum (SR). While these events are commonly observed in resting cardiac myocytes, they are rarely seen in resting skeletal muscle fibers. Since Ca(2+) spark analysis can provide extensive data on the Ca(2+) handling characteritsics of normal and diseased striated muscle, there has been interest in developing methods for observing Ca(2+) sparks in skeletal muscle. Previously, we discovered that stress generated by osmotic pressure changes induces a robust Ca(2+) spark response confined in close spatial proximity to the Sarcolemmal Membrane in wild-type intact mammalian muscles. Our studies showed these peripheral Ca(2+) sparks (PCS) were altered in dystrophic or aged skeletal muscles. Other methods to induce Ca(2+) sparks include permeabilization of the Sarcolemmal Membrane with detergents, such as saponin. In this chapter, we will discuss the methods for isolation of muscle fibers, the techniques for inducing Ca(2+) sparks in these isolated fibers, and provide guidance on the analysis of data from these experiments.
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mg53 nucleates assembly of cell Membrane repair machinery
Nature Cell Biology, 2009Co-Authors: Haruko Masumiya, Noah Weisleder, Noriyuki Matsuda, Miyuki Nishi, Moonsun Hwang, Jaekyun Ko, Angela Thornton, Xiaoli Zhao, Shinji Komazaki, Marco BrottoAbstract:Dynamic Membrane repair and remodelling is an elemental process that maintains cell integrity and mediates efficient cellular function. Here we report that MG53, a muscle-specific tripartite motif family protein (TRIM72), is a component of the Sarcolemmal Membrane-repair machinery. MG53 interacts with phosphatidylserine to associate with intracellular vesicles that traffic to and fuse with Sarcolemmal Membranes. Mice null for MG53 show progressive myopathy and reduced exercise capability, associated with defective Membrane-repair capacity. Injury of the Sarcolemmal Membrane leads to entry of the extracellular oxidative environment and MG53 oligomerization, resulting in recruitment of MG53-containing vesicles to the injury site. After vesicle translocation, entry of extracellular Ca(2+) facilitates vesicle fusion to reseal the Membrane. Our data indicate that intracellular vesicle translocation and Ca(2+)-dependent Membrane fusion are distinct steps involved in the repair of Membrane damage and that MG53 may initiate the assembly of the Membrane repair machinery in an oxidation-dependent manner.
Akinori Sato - One of the best experts on this subject based on the ideXlab platform.
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a novel disease gene for brugada syndrome Sarcolemmal Membrane associated protein gene mutations impair intracellular trafficking of hnav1 5
Circulation-arrhythmia and Electrophysiology, 2012Co-Authors: Taisuke Ishikawa, Akinori Sato, Cherisse A Marcou, David J Tester, Michael J Ackerman, Lia Crotti, Peter J Schwartz, Young Keun On, Jeong Euy ParkAbstract:Background —Mutations in genes including SCN5A encoding the α subunit of the cardiac sodium channel (hNav1.5) cause Brugada syndrome (BrS) via altered function of cardiac ion channels, but over two-thirds of BrS remains pathogenetically elusive. T-tubules and sarcoplasmic reticulum (SR) are essential in excitation of cardiomyocytes and Sarcolemmal Membrane-associated protein (SLMAP) is a protein of unknown function localizing at T-tubules and SR. Methods and Results —We analyzed 190 unrelated BrS patients for mutations in SLMAP . Two missense mutations, Val269Ile and Glu710Ala, were found in heterozygous state in two patients, which were not found in healthy individuals. Membrane surface expression of hNav1.5 in the transfected cells was affected by the mutations, and silencing of mutant SLMAP by small interfering RNA rescued the surface expression of hNav1.5. Whole-cell patch clamp recordings of hNav1.5 expressing cells transfected with mutant SLMAP confirmed the reduced hNav1.5 current. Conclusions —The mutations in SLMAP may cause BrS via modulating the intracellular trafficking of hNav1.5 channel.
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a novel disease gene for brugada syndrome Sarcolemmal Membrane associated protein gene mutations impair intracellular trafficking of hnav1 5
Circulation-arrhythmia and Electrophysiology, 2012Co-Authors: Taisuke Ishikawa, Akinori Sato, Cherisse A Marcou, David J Tester, Michael J Ackerman, Lia Crotti, Peter J Schwartz, Jeong Euy Park, Kazufumi NakamuraAbstract:Background— Mutations in genes including SCN5A encoding the α-subunit of the cardiac sodium channel (hNav1.5) cause Brugada syndrome via altered function of cardiac ion channels, but more than two-thirds of Brugada syndrome remains pathogenetically elusive. T-tubules and sarcoplasmic reticulum are essential in excitation of cardiomyocytes, and Sarcolemmal Membrane-associated protein (SLMAP) is a protein of unknown function localizing at T-tubules and sarcoplasmic reticulum. Methods and Results— We analyzed 190 unrelated Brugada syndrome patients for mutations in SLMAP . Two missense mutations, Val269Ile and Glu710Ala, were found in heterozygous state in 2 patients but were not found in healthy individuals. Membrane surface expression of hNav1.5 in the transfected cells was affected by the mutations, and silencing of mutant SLMAP by small interfering RNA rescued the surface expression of hNav1.5. Whole-cell patch-clamp recordings of hNav1.5-expressing cells transfected with mutant SLMAP confirmed the reduced hNav1.5 current. Conclusions— The mutations in SLMAP may cause Brugada syndrome via modulating the intracellular trafficking of hNav1.5 channel.
