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The microtubule cytoskeleton of pancreatic islet β-cells regulates glucose-stimulated insulin secretion (GSIS). We have reported that the microtubule-mediated movement of insulin vesicles away from the plasma membrane limits insulin secretion. High glucose-induced remodeling of microtubule network facilitates robust GSIS. This remodeling involves disassembly of old microtubules and nucleation of new microtubules. Here, we examine the mechanisms whereby glucose stimulation decreases microtubule lifetimes in β-cells. Using real-time imaging of photoconverted microtubules, we demonstrate that high levels of glucose induce rapid microtubule disassembly preferentially in the periphery of individual β-cells, and this process is mediated by the phosphorylation of microtubule-associated protein tau. Specifically, high glucose induces tau hyper-phosphorylation via glucose-responsive kinases GSK3, PKA, PKC, and CDK5. This causes dissociation of tau from and subsequent destabilization of microtubules. Consequently, tau knockdown in mouse islet β-cells facilitates microtubule turnover, causing increased basal insulin secretion, depleting insulin vesicles from the cytoplasm, and impairing GSIS. More importantly, tau knockdown uncouples microtubule destabilization from glucose stimulation. These findings suggest that tau suppresses peripheral microtubules turning over to restrict insulin oversecretion in basal conditions and preserve the insulin pool that can be released following stimulation; high glucose promotes tau phosphorylation to enhance microtubule disassembly to acutely enhance GSIS.
© 2020 by the American Diabetes Association.
The microtubule (MT) network is an essential regulator of insulin secretion from pancreatic β cells, which is central to blood-sugar homeostasis. We find that when glucose metabolism induces insulin secretion, it also increases formation of Golgi-derived microtubules (GDMTs), notably with the same biphasic kinetics as insulin exocytosis. Furthermore, GDMT nucleation is controlled by a glucose signal-transduction pathway through cAMP and its effector EPAC2. Preventing new GDMT nucleation dramatically affects the pipeline of insulin production, storage, and release. There is an overall reduction of β-cell insulin content, and remaining insulin becomes retained within the Golgi, likely because of stalling of insulin-granule budding. While not preventing glucose-induced insulin exocytosis, the diminished granule availability substantially blunts the amount secreted. Constant dynamic maintenance of the GDMT network is therefore critical for normal β-cell physiology. Our study demonstrates that the biogenesis of post-Golgi carriers, particularly large secretory granules, requires ongoing nucleation and replenishment of the GDMT network.
Copyright © 2019 Elsevier Ltd. All rights reserved.
The classic mode of G protein-coupled receptor (GPCR)-mediated transactivation of the receptor tyrosine kinase epidermal growth factor receptor (EGFR) transactivation occurs via matrix metalloprotease (MMP)-mediated cleavage of plasma membrane-anchored EGFR ligands. Herein, we show that the Gαs-activating GPCR ligands vasoactive intestinal peptide (VIP) and prostaglandin E (PGE ) transactivate EGFR through increased cell-surface delivery of the EGFR ligand transforming growth factor-α (TGFα) in polarizing madin-darby canine kidney (MDCK) and Caco-2 cells. This is achieved by PKA-mediated phosphorylation of naked cuticle homolog 2 (NKD2), previously shown to bind TGFα and direct delivery of TGFα-containing vesicles to the basolateral surface of polarized epithelial cells. VIP and PGE rapidly activate protein kinase A (PKA) that then phosphorylates NKD2 at Ser-223, a process that is facilitated by the molecular scaffold A-kinase anchoring protein 12 (AKAP12). This phosphorylation stabilized NKD2, ensuring efficient cell-surface delivery of TGFα and increased EGFR activation. Thus, GPCR-triggered, PKA/AKAP12/NKD2-regulated targeting of TGFα to the cell surface represents a new mode of EGFR transactivation that occurs proximal to ligand cleavage by MMPs.
© 2019 The Authors. Traffic published by John Wiley & Sons Ltd.
Excessive glucocorticoid exposure has been shown to be deleterious for pancreatic β-cell function and insulin release. However, glucocorticoids at physiological levels are essential for many homeostatic processes, including glycemic control. We show that corticosterone and cortisol and their less active precursors 11-dehydrocorticosterone (11-DHC) and cortisone suppress voltage-dependent Ca channel function and Ca fluxes in rodent as well as in human β-cells. However, insulin secretion, maximal ATP/ADP responses to glucose, and β-cell identity were all unaffected. Further examination revealed the upregulation of parallel amplifying cAMP signals and an increase in the number of membrane-docked insulin secretory granules. Effects of 11-DHC could be prevented by lipotoxicity and were associated with paracrine regulation of glucocorticoid activity because global deletion of 11β-hydroxysteroid dehydrogenase type 1 normalized Ca and cAMP responses. Thus, we have identified an enzymatically amplified feedback loop whereby glucocorticoids boost cAMP to maintain insulin secretion in the face of perturbed ionic signals. Failure of this protective mechanism may contribute to diabetes in states of glucocorticoid excess, such as Cushing syndrome, which are associated with frank dyslipidemia.
© 2017 by the American Diabetes Association.
Hyperpolarization-activated cyclic nucleotide-gated cation (HCN) channels have important functions in controlling neuronal excitability and generating rhythmic oscillatory activity. The role of tetratricopeptide repeat-containing Rab8b-interacting protein (TRIP8b) in regulation of hyperpolarization-activated inward current, I , in the thalamocortical system and its functional relevance for the physiological thalamocortical oscillations were investigated. A significant decrease in I current density, in both thalamocortical relay (TC) and cortical pyramidal neurons was found in TRIP8b-deficient mice (TRIP8b). In addition basal cAMP levels in the brain were found to be decreased while the availability of the fast transient A-type K current, I , in TC neurons was increased. These changes were associated with alterations in intrinsic properties and firing patterns of TC neurons, as well as intrathalamic and thalamocortical network oscillations, revealing a significant increase in slow oscillations in the delta frequency range (0.5-4 Hz) during episodes of active-wakefulness. In addition, absence of TRIP8b suppresses the normal desynchronization response of the EEG during the switch from slow-wave sleep to wakefulness. It is concluded that TRIP8b is necessary for the modulation of physiological thalamocortical oscillations due to its direct effect on HCN channel expression in thalamus and cortex and that mechanisms related to reduced cAMP signaling may contribute to the present findings.
β-adrenergic receptors (βARs) are well established for conveying the signal from catecholamines to adipocytes. Acting through the second messenger cyclic adenosine monophosphate (cAMP) they stimulate lipolysis and also increase the activity of brown adipocytes and the 'browning' of adipocytes within white fat depots (so-called 'brite' or 'beige' adipocytes). Brown adipose tissue mitochondria are enriched with uncoupling protein 1 (UCP1), which is a regulated proton channel that allows the dissipation of chemical energy in the form of heat. The discovery of functional brown adipocytes in humans and inducible brown-like ('beige' or 'brite') adipocytes in rodents have suggested that recruitment and activation of these thermogenic adipocytes could be a promising strategy to increase energy expenditure for obesity therapy. More recently, the cardiac natriuretic peptides and their second messenger cyclic guanosine monophosphate (cGMP) have gained attention as a parallel signaling pathway in adipocytes, with some unique features. In this review, we begin with some important historical work that touches upon the regulation of brown adipocyte development and physiology. We then provide a synopsis of some recent advances in the signaling cascades from β-adrenergic agonists and natriuretic peptides to drive thermogenic gene expression in the adipocytes and how these two pathways converge at a number of unexpected points. Finally, moving from the physiologic hormonal signaling, we discuss yet another level of control downstream of these signals: the growing appreciation of the emerging roles of non-coding RNAs as important regulators of brown adipocyte formation and function. In this review, we discuss new developments in our understanding of the signaling mechanisms and factors including new secreted proteins and novel non-coding RNAs that control the function as well as the plasticity of the brown/beige adipose tissue as it responds to the energy needs and environmental conditions of the organism.
Activation of β-adrenergic receptors (βARs) enhances both the induction of long-term potentiation (LTP) in hippocampal CA1 pyramidal cells and hippocampal-dependent cognitive function. Interestingly, previous studies reveal that coincident activation of group II metabotropic glutamate (mGlu) receptors with βARs in the hippocampal astrocytes induces a large increase in cyclic-AMP (cAMP) accumulation and release of adenosine. Adenosine then acts on A adenosine receptors at neighboring excitatory Schaffer collateral terminals, which could counteract effects of activation of neuronal βARs on excitatory transmission. On the basis of this, we postulated that activation of the specific mGlu receptor subtype that mediates this response could inhibit βAR-mediated effects on hippocampal synaptic plasticity and cognitive function. Using novel mGlu receptor subtype-selective allosteric modulators along with knockout mice we now report that the effects of mGlu agonists on βAR-mediated increases in cAMP accumulation are exclusively mediated by mGlu. Furthermore, mGlu activation inhibits the ability of the βAR agonist isoproterenol to enhance hippocampal LTP, and this effect is absent in slices treated with either a glial toxin or an adenosine A receptor antagonist. Finally, systemic administration of the mGlu agonist LY379268 disrupted contextual fear memory in a manner similar to the effect of the βAR antagonist propranolol, and this effect was reversed by the mGlu-negative allosteric modulator VU0650786. Taken together, these data suggest that mGlu can influence astrocytic signaling and modulate βAR-mediated effects on hippocampal synaptic plasticity and cognitive function.
Alveolar macrophages (AMs) are multitasking cells that maintain lung homeostasis by clearing apoptotic cells (efferocytosis) and performing antimicrobial effector functions. Different PRRs have been described to be involved in the binding and capture of non-opsonized Streptococcus pneumoniae, such as TLR-2, mannose receptor (MR) and scavenger receptors (SRs). However, the mechanism by which the ingestion of apoptotic cells negatively influences the clearance of non-opsonized S. pneumoniae remains to be determined. In this study, we evaluated whether the prostaglandin E2 (PGE) produced during efferocytosis by AMs inhibits the ingestion and killing of non-opsonized S. pneumoniae. Resident AMs were pre-treated with an E prostanoid (EP) receptor antagonist, inhibitors of cyclooxygenase and protein kinase A (PKA), incubated with apoptotic Jurkat T cells, and then challenged with S. pneumoniae. Efferocytosis slightly decreased the phagocytosis of S. pneumoniae but greatly inhibited bacterial killing by AMs in a manner dependent on PGE production, activation of the EP2-EP4/cAMP/PKA pathway and inhibition of HO production. Our data suggest that the PGE produced by AMs during efferocytosis inhibits HO production and impairs the efficient clearance non-opsonized S. pneumoniae by EP2-EP4/cAMP/PKA pathway.
Cholesterol synthesis is among the oldest metabolic pathways, consisting of the Bloch and Kandutch-Russell branches. Following lanosterol, sterols of both branches are proposed to be dedicated to cholesterol. We challenge this dogma by mathematical modeling and with experimental evidence. It was not possible to explain the sterol profile of testis in cAMP responsive element modulator tau (Crem τ) knockout mice with mathematical models based on textbook pathways of cholesterol synthesis. Our model differs in the inclusion of virtual sterol metabolizing enzymes branching from the pathway. We tested the hypothesis that enzymes from the cytochrome P450 (CYP) superfamily can participate in the catalysis of non-classical reactions. We show that CYP enzymes can metabolize multiple sterols in vitro, establishing novel branching points of cholesterol synthesis. In conclusion, sterols of cholesterol synthesis can be oxidized further to metabolites not dedicated to production of cholesterol. Additionally, CYP7A1, CYP11A1, CYP27A1, and CYP46A1 are parts of a broader cholesterol synthesis network.
Although men with testosterone deficiency are at increased risk for type 2 diabetes (T2D), previous studies have ignored the role of testosterone and the androgen receptor (AR) in pancreatic β cells. We show that male mice lacking AR in β cells (βARKO) exhibit decreased glucose-stimulated insulin secretion (GSIS), leading to glucose intolerance. The AR agonist dihydrotestosterone (DHT) enhances GSIS in cultured male islets, an effect that is abolished in βARKO(-/y) islets and human islets treated with an AR antagonist. In β cells, DHT-activated AR is predominantly extranuclear and enhances GSIS by increasing islet cAMP and activating the protein kinase A. In mouse and human islets, the insulinotropic effect of DHT depends on activation of the glucagon-like peptide-1 (GLP-1) receptor, and accordingly, DHT amplifies the incretin effect of GLP-1. This study identifies AR as a novel receptor that enhances β cell function, a finding with implications for the prevention of T2D in aging men.
Copyright © 2016 Elsevier Inc. All rights reserved.