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The pancreatic islets are composed of discrete hormone-producing cells that orchestrate systemic glucose homeostasis. Here we identify subsets of beta cells using a single-cell transcriptomic approach. One subset of beta cells marked by high CD63 expression is enriched for the expression of mitochondrial metabolism genes and exhibits higher mitochondrial respiration compared with CD63lo beta cells. Human and murine pseudo-islets derived from CD63hi beta cells demonstrate enhanced glucose-stimulated insulin secretion compared with pseudo-islets from CD63lo beta cells. We show that CD63hi beta cells are diminished in mouse models of and in humans with type 2 diabetes. Finally, transplantation of pseudo-islets generated from CD63hi but not CD63lo beta cells into diabetic mice restores glucose homeostasis. These findings suggest that loss of a specific subset of beta cells may lead to diabetes. Strategies to reconstitute or maintain CD63hi beta cells may represent a potential anti-diabetic therapy.

The voltage-gated sodium channel is critical for cardiomyocyte function and consists of a protein complex comprising a pore-forming α subunit and two associated β subunits. It has been shown previously that the associated β2 subunits promote cell surface expression of the α subunit. The major α isoform in the adult human heart is NaV1.5, and germline mutations in the NaV1.5-encoding gene, sodium voltage-gated channel α subunit 5 (SCN5A), often cause inherited arrhythmias. Here, we investigated the mechanisms that regulate β2 trafficking and how they may determine proper NaV1.5 cell surface localization. Using heterologous expression in polarized Madin-Darby canine kidney cells, we show that β2 is N-glycosylated in vivo and in vitro at residues 42, 66, and 74, becoming sialylated only at Asn-42. We found that fully nonglycosylated β2 was mostly retained in the endoplasmic reticulum, indicating that N-linked glycosylation is required for efficient β2 trafficking to the apical plasma membrane. The nonglycosylated variant reached the cell surface by bypassing the Golgi compartment at a rate of only approximately one-third of that of WT β2. YFP-tagged, nonglycosylated β2 displayed mobility kinetics in the plane of the membrane similar to that of WT β2. However, it was defective in promoting surface localization of NaV1.5. Interestingly, β2 with a single intact glycosylation site was as effective as the WT in promoting NaV1.5 surface localization. In conclusion, our results indicate that N-linked glycosylation of β2 is required for surface localization of NaV1.5, a property that is often defective in inherited cardiac arrhythmias. [ABSTRACT FROM AUTHOR]

Alterations in ubiquitin-mediated degradation of NaV1.5 can cause arrhythmia Conversely, a LQT3-associated point mutation disrupting the existing C-terminal PY motif, Y1977N, causes gain-of-function by preventing interaction with Nedd4-2, therefore restricting degradation of Na SB V sb 1.5 (Figure 1). In a clever way, these researchers also disrupted the newly generated PY motif of the BrS-associated variant, nicely restoring Na SB V sb 1.5 levels and function as a consequence of decreased ubiquitination by Nedd4-2 and degradation. [Extracted from the article]

Background Information Cardiac channelopathies arise by mutations in genes encoding ion channel subunits. One example is Brugada Syndrome (BrS), which causes arrhythmias and sudden death. BrS is often associated with mutations in SCN5A, encoding Nav1.5, the α subunit of the major cardiac voltage-gated sodium channel. This channel forms a protein complex including one or two associated β subunits as well as other proteins. Results We analysed regulation of Nav1.5 localisation and trafficking by β2, specifically, Nav1.5 arrival to the cell surface. We used polarised Madin-Darby canine kidney (MDCK) cells and mouse atria-derived HL-1 cells, which retain phenotypic features of adult cardiomyocytes. In both, Nav1.5 was found essentially intracellular, mainly in the endoplasmic reticulum, whereas β2 localised to the plasma membrane, and was restricted to the apical surface in MDCK cells. A fraction of β2 interacted with Nav1.5, despite their limited overlap. Importantly, β2 promoted Nav1.5 localisation to the cell surface. Both β2 WT and the BrS-associated mutation D211G (substitution of Asp for Gly) effectively reached the plasma membrane. Strikingly, however, β2 D211G was defective in promoting Nav1.5 surface localisation. Conclusions Our data sustain that β2 promotes surface localisation of Nav1.5, which can be affected due to β2 mutations associated with channelopathies. Significance Our findings add to the understanding of β2 role in Nav1.5 trafficking and localisation, which must influence cell excitability and electrical coupling in the heart. This study will contribute to knowledge on development of arrhythmias. [ABSTRACT FROM AUTHOR]

Together with obesity and type 2 diabetes, metabolic dysfunction-associated steatotic liver disease (MASLD) is a growing global epidemic. Activation of the complement system and infiltration of macrophages has been linked to progression of metabolic liver disease. The role of complement receptors in macrophage activation and recruitment in MASLD remains poorly understood. In human and mouse, C3AR1 in the liver is expressed primarily in Kupffer cells, but is downregulated in humans with MASLD compared to obese controls. To test the role of complement 3a receptor (C3aR1) on macrophages and liver resident macrophages in MASLD, we generated mice deficient in C3aR1 on all macrophages (C3aR1-MφKO) or specifically in liver Kupffer cells (C3aR1-KpKO) and subjected them to a model of metabolic steatotic liver disease. We show that macrophages account for the vast majority of C3ar1 expression in the liver. Overall, C3aR1-MφKO and C3aR1-KpKO mice have similar body weight gain without significant alterations in glucose homeostasis, hepatic steatosis and fibrosis, compared to controls on a MASLD-inducing diet. This study demonstrates that C3aR1 deletion in macrophages or Kupffer cells, the predominant liver cell type expressing C3aR1 , has no significant effect on liver steatosis, inflammation or fibrosis in a dietary MASLD model., Competing Interests: Declaration of Competing Interest None

The voltage-gated sodium channel is critical for cardiomyocyte function and consists of a protein complex comprising a pore-forming α subunit and two associated β subunits. It has been shown previously that the associated β2 subunits promote cell surface expression of the α subunit. The major α isoform in the adult human heart is Na V 1.5, and germline mutations in the Na V 1.5-encoding gene, sodium voltage-gated channel α subunit 5 ( SCN5A ), often cause inherited arrhythmias. Here, we investigated the mechanisms that regulate β2 trafficking and how they may determine proper Na V 1.5 cell surface localization. Using heterologous expression in polarized Madin-Darby canine kidney cells, we show that β2 is N -glycosylated in vivo and in vitro at residues 42, 66, and 74, becoming sialylated only at Asn-42. We found that fully nonglycosylated β2 was mostly retained in the endoplasmic reticulum, indicating that N -linked glycosylation is required for efficient β2 trafficking to the apical plasma membrane. The nonglycosylated variant reached the cell surface by bypassing the Golgi compartment at a rate of only approximately one-third of that of WT β2. YFP-tagged, nonglycosylated β2 displayed mobility kinetics in the plane of the membrane similar to that of WT β2. However, it was defective in promoting surface localization of Na V 1.5. Interestingly, β2 with a single intact glycosylation site was as effective as the WT in promoting Na V 1.5 surface localization. In conclusion, our results indicate that N -linked glycosylation of β2 is required for surface localization of Na V 1.5, a property that is often defective in inherited cardiac arrhythmias., (© 2019 Cortada et al.)

Background Information: Cardiac channelopathies arise by mutations in genes encoding ion channel subunits. One example is Brugada Syndrome (BrS), which causes arrhythmias and sudden death. BrS is often associated with mutations in SCN5A, encoding Na v 1.5, the α subunit of the major cardiac voltage-gated sodium channel. This channel forms a protein complex including one or two associated β subunits as well as other proteins., Results: We analysed regulation of Na v 1.5 localisation and trafficking by β2, specifically, Na v 1.5 arrival to the cell surface. We used polarised Madin-Darby canine kidney (MDCK) cells and mouse atria-derived HL-1 cells, which retain phenotypic features of adult cardiomyocytes. In both, Na v 1.5 was found essentially intracellular, mainly in the endoplasmic reticulum, whereas β2 localised to the plasma membrane, and was restricted to the apical surface in MDCK cells. A fraction of β2 interacted with Na v 1.5, despite their limited overlap. Importantly, β2 promoted Na v 1.5 localisation to the cell surface. Both β2 WT and the BrS-associated mutation D211G (substitution of Asp for Gly) effectively reached the plasma membrane. Strikingly, however, β2 D211G was defective in promoting Na v 1.5 surface localisation., Conclusions: Our data sustain that β2 promotes surface localisation of Na v 1.5, which can be affected due to β2 mutations associated with channelopathies., Significance: Our findings add to the understanding of β2 role in Na v 1.5 trafficking and localisation, which must influence cell excitability and electrical coupling in the heart. This study will contribute to knowledge on development of arrhythmias., (© 2017 Société Française des Microscopies and Société de Biologie Cellulaire de France. Published by John Wiley & Sons Ltd.)