Absence of Shb impairs insulin secretion by elevated FAK activity in pancreatic islets

The Src homology-2 domain containing protein B (SHB) has previously been shown to function as a pleiotropic adapter protein, conveying signals from receptor tyrosine kinases to intracellular signaling intermediates. The overexpression ofShbin β-cells promotes β-cell proliferation by increased insulin receptor substrate (IRS) and focal adhesion kinase (FAK) activity, whereasShbdeficiency causes moderate glucose intolerance and impaired first-peak insulin secretion. Using an array of techniques, including live-cell imaging, patch-clamping, immunoblotting, and semi-quantitative PCR, we presently investigated the causes of the abnormal insulin secretory characteristics inShb-knockout mice.Shb-knockout islets displayed an abnormal signaling signature with increased activities of FAK, IRS, and AKT. β-catenin protein expression was elevated and it showed increased nuclear localization. However, there were no major alterations in the gene expression of various proteins involved in the β-cell secretory machinery. Nor wasShbdeficiency associated with changes in glucose-induced ATP generation or cytoplasmic Ca2+handling. In contrast, the glucose-induced rise in cAMP, known to be important for the insulin secretory response, was delayed in theShb-knockout compared with WT control. Inhibition of FAK increased the submembrane cAMP concentration, implicating FAK activity in the regulation of insulin exocytosis. In conclusion,Shbdeficiency causes a chronic increase in β-cell FAK activity that perturbs the normal insulin secretory characteristics of β-cells, suggesting multi-faceted effects of FAK on insulin secretion depending on the mechanism of FAK activation.

Download Full-text

Expression of Src Homology 2 Domain-Containing Protein Tyrosine Phosphatase Substrate-1 in Pancreatic β-Cells and Its Role in Promotion of Insulin Secretion and Protection against Diabetes

Endocrinology ◽

10.1210/en.2008-0236 ◽

2008 ◽

Vol 149 (11) ◽

pp. 5662-5669 ◽

Cited By ~ 9

Author(s):

Masaki Kobayashi ◽

Hiroshi Ohnishi ◽

Hideki Okazawa ◽

Yoji Murata ◽

Yuriko Hayashi ◽

...

Keyword(s):

Insulin Secretion ◽

Protein Tyrosine Phosphatase ◽

Tyrosine Phosphatase ◽

Β Cells ◽

Pancreatic Β Cells ◽

Protein Tyrosine ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain ◽

Phosphatase Substrate

Download Full-text

α2-adrenergic signaling disrupts β cell BDNF-TrkB receptor tyrosine kinase signaling

10.1101/400010 ◽

2018 ◽

Cited By ~ 1

Author(s):

Michael A. Kalwat ◽

Zhimin Huang ◽

Derk D. Binns ◽

Kathleen McGlynn ◽

Melanie H. Cobb

Keyword(s):

Insulin Secretion ◽

Tyrosine Kinases ◽

Careful Consideration ◽

Mitogen Activated Protein Kinase ◽

Published Data ◽

High Dose ◽

Kinase Signaling ◽

Β Cells ◽

Β Cell ◽

Adrenergic Signaling

AbstractAdrenergic signaling is a well-known input into pancreatic islet function. Specifically, the insulin-secreting islet β cell expresses the Gi/o-linked α2-adrenergic receptor, which upon activation suppresses insulin secretion. The use of adrenergic agonist epinephrine at micromolar doses may have supraphysiological effects. We found that pretreating β cells with micromolar concentrations of epinephrine differentially inhibited activation of receptor tyrosine kinases. We chose TrkB as an example because of its relative sensitivity to the effects of epinephrine and due to its potential regulatory role in the β cell. Our characterization of brain-derived neurotrophic factor (BDNF)-TrkB signaling in MIN6 β cells showed that TrkB is activated by BDNF as expected, leading to canonical TrkB autophosphorylation and subsequent downstream signaling, as well as chronic effects on β cell growth. Micromolar, but not nanomolar, concentrations of epinephrine blocked BDNF-induced TrkB autophosphorylation and downstream mitogen-activated protein kinase pathway activation, suggesting an inhibitory phenomenon at the receptor level. We determined epinephrine-mediated inhibition of TrkB activation to be Gi/o-dependent using pertussis toxin, arguing against an off-target effect of high dose epinephrine. Published data suggested that inhibition of potassium channels or phosphoinositide-3-kinase signaling may abrogate the negative effects of epinephrine, however these did not rescue TrkB signaling in our experiments. Taken together, these results show that 1) TrkB kinase signaling occurs in β cells and 2) use of epinephrine in studies of insulin secretion requires careful consideration of concentration-dependent effects. BDNF-TrkB signaling in β cells may underlie pro-survival or growth signaling and warrants further study.

Download Full-text

p56lck interacts via its src homology 2 domain with the ZAP-70 kinase.

Journal of Experimental Medicine ◽

10.1084/jem.179.4.1163 ◽

1994 ◽

Vol 179 (4) ◽

pp. 1163-1172 ◽

Cited By ~ 105

Author(s):

P Duplay ◽

M Thome ◽

F Hervé ◽

O Acuto

Keyword(s):

Tyrosine Kinases ◽

Intracellular Signaling ◽

Specific Binding ◽

Sh2 Domain ◽

Jurkat Cells ◽

Vitro Assay ◽

Phosphorylated Proteins ◽

Src Homology 2 ◽

Src Homology

p56lck, a member of the src family of protein tyrosine kinases, is an essential component in T cell receptor (TCR) signal transduction. p56lck contains a src homology 2 (SH2) domain found in a number of proteins involved in intracellular signaling. SH2 domains have been implicated in protein-protein interactions by binding to sequences in target proteins containing phosphorylated tyrosine. Using an in vitro assay, we have studied specific binding of tyrosine-phosphorylated proteins to a recombinant p56lck SH2 domain. In nonactivated Jurkat cells, two tyrosine-phosphorylated proteins were detected. Stimulation with anti-CD3 monoclonal antibodies induced the binding of seven additional tyrosine-phosphorylated proteins to the SH2 domain of p56lck. We have identified the zeta-associated tyrosine kinase, ZAP-70, as one of these proteins. Evidence suggests that binding of ZAP-70 to p56lck SH2 is direct and not mediated by zeta. The significance of this interaction was further investigated in vivo. p56lck could be coprecipitated with the zeta/ZAP-70 complex and conversely, ZAP-70 was detected in p56lck immunoprecipitates of activated Jurkat cells. The physical association of p56lck and ZAP-70 during activation supports the recently proposed functional cooperation of these two tyrosine kinases in TCR signaling.

Download Full-text

GPVI and α2β1 play independent critical roles during platelet adhesion and aggregate formation to collagen under flow

Blood ◽

10.1182/blood-2004-11-4434 ◽

2005 ◽

Vol 106 (4) ◽

pp. 1268-1277 ◽

Cited By ~ 109

Author(s):

Kendra L. Sarratt ◽

Hong Chen ◽

Mary M. Zutter ◽

Samuel A. Santoro ◽

Daniel A. Hammer ◽

...

Keyword(s):

Intracellular Signaling ◽

Platelet Adhesion ◽

Glycoprotein Vi ◽

Collagen Receptors ◽

Src Homology 2 ◽

Approaches To Study ◽

Src Homology ◽

Src Homology 2 Domain ◽

Density Results

AbstractThe roles of the 2 major platelet-collagen receptors, glycoprotein VI (GPVI) and integrin α2β1, have been intensely investigated using a variety of methods over the past decade. In the present study, we have used pharmacologic and genetic approaches to study human and mouse platelet adhesion to collagen under flow conditions. Our studies demonstrate that both GPVI and integrin α2β1 play significant roles for platelet adhesion to collagen under flow and that the loss of both receptors completely ablates this response. Intracellular signaling mediated by the cytoplasmic adaptor Src homology 2 domain-containing leukocyte protein of 76 kDa (SLP-76) but not by the transmembrane adaptor linker for activation of T cells (LAT) is critical for platelet adhesion to collagen under flow. In addition, reduced GPVI receptor density results in severe defects in platelet adhesion to collagen under flow. Defective adhesion to collagen under flow is associated with prolonged tail-bleeding times in mice lacking one or both collagen receptors. These studies establish platelet-collagen responses under physiologic flow as the consequence of a close partnership between 2 structurally distinct receptors and suggest that both receptors play significant hemostatic roles in vivo.

Download Full-text

Tyrosine Phosphorylation of Tub and Its Association with Src Homology 2 Domain-containing Proteins Implicate Tub in Intracellular Signaling by Insulin

Journal of Biological Chemistry ◽

10.1074/jbc.274.35.24980 ◽

1999 ◽

Vol 274 (35) ◽

pp. 24980-24986 ◽

Cited By ~ 68

Author(s):

Rosana Kapeller ◽

Ann Moriarty ◽

Ann Strauss ◽

Hilde Stubdal ◽

Kelly Theriault ◽

...

Keyword(s):

Tyrosine Phosphorylation ◽

Intracellular Signaling ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain

Download Full-text

Functional specificity of cytoplasmic and transmembrane tyrosine kinases: identification of 130- and 75-kilodalton substrates of c-fps/fes tyrosine kinase in macrophages

Molecular and Cellular Biology ◽

10.1128/mcb.14.7.4606-4615.1994 ◽

1994 ◽

Vol 14 (7) ◽

pp. 4606-4615

Author(s):

L B Areces ◽

P Dello Sbarba ◽

M Jücker ◽

E R Stanley ◽

R A Feldman

Keyword(s):

Tyrosine Kinase ◽

Tyrosine Phosphorylation ◽

Tyrosine Kinases ◽

Protein Tyrosine Kinase ◽

Phosphatidylinositol 3 Kinase ◽

Macrophage Cell ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain ◽

Stimulating Factor

c-fps/fes encodes a 92-kDa protein-tyrosine kinase (NCP92) that is expressed at the highest levels in macrophages. To determine if c-fps/fes can mediate the action of the colony-stimulating factor 1 (CSF-1) receptor (CSF-1R) and to identify potential targets of c-fps/fes in macrophages, we have overexpressed c-fps/fes in a CSF-1-dependent macrophage cell line. A 30- to 50-fold overexpression of c-fps/fes partially released these cells from their factor dependence by a nonautocrine mechanism, and this correlated with the tyrosine phosphorylation of two proteins of 130 and 75 kDa (P130 and P75). c-fps/fes did not cause tyrosine phosphorylation or activation of CSF-1 dependent targets, including CSF-1R, Shc, and phosphatidylinositol 3-kinase, and conversely, CSF-1 did not induce tyrosine phosphorylation of P130 and P75. P75 appears to be a novel phosphotyrosyl protein, whereas P130 cross-reacts with a known substrate of v-src. P130 and P75 may be direct substrates of c-fps/fes: P130 was tightly associated with NCP92, and the src homology 2 domain of NCP92 specifically bound phosphorylated P130 and P75 but not the CSF-1-induced phosphotyrosyl proteins, consistent with the possibility that P130 and P75 are physiological targets of c-fps/fes. We conclude that although c-fps/fes can functionally substitute for CSF-1R to a certain extent, these tyrosine kinases act largely independently of each other and that P130 and P75 are novel targets whose mechanisms of action may be unrelated to the signalling pathways utilized by receptor tyrosine kinases.

Download Full-text

Functional specificity of cytoplasmic and transmembrane tyrosine kinases: identification of 130- and 75-kilodalton substrates of c-fps/fes tyrosine kinase in macrophages.

Molecular and Cellular Biology ◽

10.1128/mcb.14.7.4606 ◽

1994 ◽

Vol 14 (7) ◽

pp. 4606-4615 ◽

Cited By ~ 21

Author(s):

L B Areces ◽

P Dello Sbarba ◽

M Jücker ◽

E R Stanley ◽

R A Feldman

Keyword(s):

Tyrosine Kinase ◽

Tyrosine Phosphorylation ◽

Tyrosine Kinases ◽

Protein Tyrosine Kinase ◽

Phosphatidylinositol 3 Kinase ◽

Macrophage Cell ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain ◽

Stimulating Factor

Download Full-text

Commonin VitroSubstrate Specificity and Differential Src Homology 2 Domain Accessibility Displayed by Two Members of the Src Family of Protein-tyrosine Kinases, c-Src and Hck

Journal of Biological Chemistry ◽

10.1074/jbc.273.27.16756 ◽

1998 ◽

Vol 273 (27) ◽

pp. 16756-16763 ◽

Cited By ~ 12

Author(s):

Robert J. Sicilia ◽

Margaret L. Hibbs ◽

Paul A. Bello ◽

Jeffrey D. Bjorge ◽

Donald J. Fujita ◽

...

Keyword(s):

Tyrosine Kinases ◽

Protein Tyrosine Kinases ◽

Protein Tyrosine ◽

Src Homology 2 ◽

Src Homology ◽

Src Homology 2 Domain

Download Full-text

Mitochondrial damages and the regulation of insulin secretion

Biochemical Society Transactions ◽

10.1042/bst0340824 ◽

2006 ◽

Vol 34 (5) ◽

pp. 824-827 ◽

Cited By ~ 21

Author(s):

P. Maechler ◽

P.B.M. de Andrade

Keyword(s):

Insulin Secretion ◽

Mitochondrial Function ◽

Metabolic Stress ◽

Β Cells ◽

Β Cell ◽

Mitochondrial Dna Mutation ◽

Cellular Models ◽

Cell Functions ◽

Direct Demonstration ◽

Insulin Exocytosis

Pancreatic β-cells are able to respond to nutrients, principally glucose, as the primary stimulus for insulin exocytosis. This unique feature requires translation of metabolic substrates into intracellular messengers recognized by the exocytotic machinery. Central to this signal transduction mechanism, mitochondria integrate and generate metabolic signals, thereby coupling glucose recognition with insulin secretion. In response to a glucose rise, nucleotides and metabolites are generated by mitochondria and participate, together with cytosolic Ca2+, in the stimulation of insulin exocytosis. Mitochondrial defects, such as mutations and ROS (reactive oxygen species) production, might be associated with β-cell failure in the course of diabetes. mtDNA (mitochondrial DNA) mutation A3243G is associated with MIDD (mitochondrial inherited diabetes and deafness). A common hypothesis to explain the link between the genotype and the phenotype is that the mutation might impair mitochondrial metabolism expressly required for β-cell functions, although this assumption lacks direct demonstration. mtDNA-deficient cellular models are glucose-unresponsive and are defective in mitochondrial function. Recently, we used clonal cytosolic hybrid cells (namely cybrids) harbouring mitochondria derived from MIDD patients. Compared with control mtDNA from the same patient, the A3243G mutation markedly modified metabolic pathways. Moreover, cybrid cells carrying patient-derived mutant mtDNA exhibited deranged cell Ca2+ handling and elevated ROS under metabolic stress. In animal models, transgenic mice lacking expression of the mitochondrial genome specifically in β-cells are diabetic and their islets are incable of releasing insulin in response to glucose. These various models demonstrate the fragility of nutrient-stimulated insulin secretion, caused primarily by defective mitochondrial function.

Download Full-text

Ontology of the apelinergic system in mouse pancreas during pregnancy and relationship with β-cell mass

Scientific Reports ◽

10.1038/s41598-021-94725-0 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Brenda Strutt ◽

Sandra Szlapinski ◽

Thineesha Gnaneswaran ◽

Sarah Donegan ◽

Jessica Hill ◽

...

Keyword(s):

Cell Proliferation ◽

Insulin Secretion ◽

Glucose Transporter ◽

Cell Mass ◽

Elevated Serum ◽

Pancreatic Β Cell ◽

Β Cells ◽

Β Cell ◽

Glucose Transporter 2 ◽

Glucose Stimulated Insulin Secretion

AbstractThe apelin receptor (Aplnr) and its ligands, Apelin and Apela, contribute to metabolic control. The insulin resistance associated with pregnancy is accommodated by an expansion of pancreatic β-cell mass (BCM) and increased insulin secretion, involving the proliferation of insulin-expressing, glucose transporter 2-low (Ins+Glut2LO) progenitor cells. We examined changes in the apelinergic system during normal mouse pregnancy and in pregnancies complicated by glucose intolerance with reduced BCM. Expression of Aplnr, Apelin and Apela was quantified in Ins+Glut2LO cells isolated from mouse pancreata and found to be significantly higher than in mature β-cells by DNA microarray and qPCR. Apelin was localized to most β-cells by immunohistochemistry although Aplnr was predominantly associated with Ins+Glut2LO cells. Aplnr-staining cells increased three- to four-fold during pregnancy being maximal at gestational days (GD) 9–12 but were significantly reduced in glucose intolerant mice. Apelin-13 increased β-cell proliferation in isolated mouse islets and INS1E cells, but not glucose-stimulated insulin secretion. Glucose intolerant pregnant mice had significantly elevated serum Apelin levels at GD 9 associated with an increased presence of placental IL-6. Placental expression of the apelinergic axis remained unaltered, however. Results show that the apelinergic system is highly expressed in pancreatic β-cell progenitors and may contribute to β-cell proliferation in pregnancy.

Download Full-text