Cleavage of Chromogranin A N-terminal Domain by Plasmin Provides a New Mechanism for Regulating Cell Adhesion

Colombo, Barbara; Longhi, Renato; Marinzi, Chiara; Magni, Fulvio; Cattaneo, Angela; Yoo, Seung Hyun; Curnis, Flavio; Corti, Angelo

doi:10.1074/jbc.m202637200

Cited by 34 publications

(23 citation statements)

References 49 publications

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“…Presence of the insulin B9-23 peptide in secretory granules of islet beta cells and direct delivery of these peptide fragments to the islet DCs has been previously reported [23]. Plasmin in the secretory granules [24] cleaves VS-1 at lysine sites [25]. Thus, the presence of a natural peptide in the region spanning amino acids 27-46 of the VS-1 molecule that overlaps with the ChgA 29-42 epitope is highly probable.…”

Section: Discussionmentioning

confidence: 99%

Vasostatin‐1 antigenic epitope mapping for induction of cellular and humoral immune responses to chromogranin A autoantigen in NOD mice

et al. 2014

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Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionChromogranin A (ChgA) is a prohormone in neuroendocrine tissues that is a target of pathogenic CD4 + T cells in NOD mice [1,2].This prohormone is an acidic secretory protein found in the granules of neuroendocrine cells, and can be cleaved to form functional proteins, including vasostatin-1 (VS-1, ChgA 1-76), vasostatin-2 (VS-2, ChgA 1-113), and WE-14 [3]. In pancreatic β-cells, VS-1 is a major fragment of ChgA that remains after proteolysis [4][5][6][7] and is the critical peptide for the formation of secretory granules that constitutively store insulin [8].Correspondence: Dr. Bhagirath Singh e-mail: bsingh@uwo.ca BDC2.5 T cells are the most extensively studied T cells of the diabetes-prone NOD mouse. Significantly important findings have been made with T-cell receptor transgenic BDC2.5 NOD mice generated using these T cells [9]. For further functional studies, it is important to evaluate the antigen specificity of these T cells. We have identified the region between residues 29 and 42 of the ChgA molecule to be the antigenic epitope for BDC2.5 T cells in NOD mice [1]. Since this peptide contains cysteine and methionine, oxidation of the −SH and −S-CH 3 side chains of these amino acids could have a significant impact on its antigenic activity [10,11]. Recently, another group [12] reported conflicting results about the antigenicity of ChgA 29-42 peptide and we suggest that it may be due to issues relating to the solubility of the peptide and/or potential oxidation of cysteine and methionine residues. The inhibitory dose of peptide (IC 50 ) for 50% reduction in binding of biotinylated HEL 10-23 to the I-A g7 is shown. Numbers in bold (1 to 9) indicate the binding register. Substitutions of amino acids in the sequence are shown in bold letters.In this study, we have done fine mapping of the VS-1 epitope ChgA 29-42 through amino acid deletion and alanine scanning of the I-A g7 binding register (KCVLEVISD). We characterized the presence of ChgA fragments in the pancreas of diabetes-prone NOD mice. Our findings also indicate the presence of elevated titers of antibody response against the VS-1 epitope ChgA 29-42 in diabetic NOD mice. ResultsChgA 34-42 is a binding register of the ChgA 29-42 epitope for I-A g7Binding of the peptide epitopes to the I-A g7 molecule occurs at P1, P4, P6, P7, and P9 MHC class II anchor positions while P2, P3, P5, and P8 positions act as T-cell receptor contact residues [13]. We generated a library of peptides for binding and T-cell stimulation studies ( Table 1). The binding capacity of the various peptides was determined as a percentage of inhibition of binding for biotinylated hen egg lysozyme (HEL) 10-23 (Bio-HEL) peptide to I-A g7 MHC class II in a competitive assay (Fig. 1B-D). The concentration of the competitor peptide required for 50% inhibition of binding of the labeled HEL peptide indicated as IC 50 (Table 1). The HEL 10-23 and IIP peptides were used as p...

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Section: Discussionmentioning

confidence: 99%

Vasostatin‐1 antigenic epitope mapping for induction of cellular and humoral immune responses to chromogranin A autoantigen in NOD mice

et al. 2014

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“…An emerging area of research has demonstrated a novel role for the serine protease plasmin as a prohormone-processing protease in the neuroendocrine system (Parmer et al, 2000;Hoover-Plow et al, 2001;Jiang et al, 2001;Colombo et al, 2002;Q. Jiang et al, 2002;Pang et al, 2004;N.…”

Section: Introductionmentioning

confidence: 99%

Cell-Surface Actin Binds Plasminogen and Modulates Neurotransmitter Release from Catecholaminergic Cells

Miles¹,

Andronicos²,

Baik³

et al. 2006

J. Neurosci.

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An emerging area of research has documented a novel role for the plasminogen activation system in the regulation of neurotransmitter release. Prohormones, secreted by cells within the sympathoadrenal system, are processed by plasmin to bioactive peptides that feed back to inhibit secretagogue-stimulated release. Catecholaminergic cells of the sympathoadrenal system are prototypic prohormone-secreting cells. Processing of prohormones by plasmin is enhanced in the presence of catecholaminergic cells, and the enhancement requires binding of plasmin(ogen) to cellular receptors. Consequently, modulation of the local cellular fibrinolytic system of catecholaminergic cells results in substantial changes in catecholamine release. However, mechanisms for enhancing prohormone processing and cellsurface molecules mediating the enhancement on catecholaminergic cells have not been investigated. Here we show that plasminogen activation was enhanced Ͼ6.5-fold on catecholaminergic cells. Carboxypeptidase B treatment decreased cell-dependent plasminogen activation by ϳ90%, suggesting that the binding of plasminogen to proteins exposing C-terminal lysines on the cell surface is required to promote plasminogen activation. We identified catecholaminergic plasminogen receptors required for enhancing plasminogen activation, using a novel strategy combining targeted specific proteolysis using carboxypeptidase B with a proteomics approach using twodimensional gel electrophoresis, radioligand blotting, and tandem mass spectrometry. Two major plasminogen-binding proteins that exposed C-terminal lysines on the cell surface contained amino acid sequences corresponding to ␤/␥-actin. An anti-actin monoclonal antibody inhibited cell-dependent plasminogen activation and also enhanced nicotine-dependent catecholamine release. Our results suggest that cell-surface-expressed forms of actin bind plasminogen, thereby promoting plasminogen activation and increased prohormone processing leading to inhibition of neurotransmitter release.

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“…b Schematic representation of the N-terminal domain of CgA showing the RGD motif and the amphipathic a-helix. Antibody epitope topography and ezrin-binding-protein-50-homology domain [15][16][17][18] are also indicated. mAb 7D1 and 5A8, but not B4E11 can block the CgA/avb6 interaction (RGE) on skin wound healing in mice.…”

Section: Discussionmentioning

confidence: 99%

“…We have previously shown that CgA can also inhibit the adhesion of fibroblasts to various extracellular-matrix proteins, whereas the CgA 1-78 fragment (VS-1) can promote cell adhesion and spreading [15][16][17][18]. CgA and VS-1 can also enhance endothelial cell-cell adhesion and protect the endothelial barrier integrity from vascular leakage induced by tumor necrosis factor alpha [19,20].…”

Section: Introductionmentioning

confidence: 99%

Chromogranin A binds to αvβ6-integrin and promotes wound healing in mice

Curnis

Gasparri

Longhi

et al. 2012

Cell. Mol. Life Sci.

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Chromogranin A (CgA), a secretory protein expressed by many neuroendocrine cells, neurons, cardiomyocytes, and keratinocytes, is the precursor of various peptides that regulate the carbohydrate/lipid metabolism and the cardiovascular system. We have found that CgA, locally administered to injured mice, can accelerate keratinocyte proliferation and wound healing. This biological activity was abolished by the Asp(45)Glu mutation. CgA and its N-terminal fragments, but not the corresponding Asp(45)Glu mutants, could selectively recognize the αvβ6-integrin on keratinocytes (a cell-adhesion receptor that is up-regulated during wound healing) and regulate keratinocyte adhesion, proliferation, and migration. No binding was observed to other integrins such as αvβ3, αvβ5, αvβ8, α5β1, α1β1, α3β1, α6β4, α6β7 and α9β1. Structure-activity studies showed that the entire CgA(39-63) region is crucial for αvβ6 recognition (K(i) = 7 nM). This region contains an RGD site (residues CgA(43-45)) followed by an amphipathic α-helix (residues CgA(47-63)), both crucial for binding affinity and selectivity. These results suggest that the interaction of the RGD/α-helix motif of CgA with αvβ6 regulates keratinocyte physiology in wound healing.

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Cleavage of Chromogranin A N-terminal Domain by Plasmin Provides a New Mechanism for Regulating Cell Adhesion

Cited by 34 publications

References 49 publications

Vasostatin‐1 antigenic epitope mapping for induction of cellular and humoral immune responses to chromogranin A autoantigen in NOD mice

Vasostatin‐1 antigenic epitope mapping for induction of cellular and humoral immune responses to chromogranin A autoantigen in NOD mice

Cell-Surface Actin Binds Plasminogen and Modulates Neurotransmitter Release from Catecholaminergic Cells

Chromogranin A binds to αvβ6-integrin and promotes wound healing in mice

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