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Start Over Author "Salvatore, C." Journal journal of biological chemistry
8 results on '"Salvatore, C."'

1. BCL-2 Selectively Interacts with the BID-induced Open Conformer of BAK, Inhibiting BAK Auto-oligomerization

Author

Salvatore C. Ruffolo and Gordon C. Shore

Subjects
Protein Conformation, Molecular Sequence Data, Apoptosis, Plasma protein binding, Mitochondrion, Biochemistry, Protein structure, Amino Acid Sequence, Molecular Biology, biology, Chemistry, Cytochrome c, Membrane Proteins, Cell Biology, Cell biology, N-terminus, bcl-2 Homologous Antagonist-Killer Protein, Proto-Oncogene Proteins c-bcl-2, biology.protein, biological phenomena, cell phenomena, and immunity, Carrier Proteins, Bacterial outer membrane, Dimerization, Bcl-2 Homologous Antagonist-Killer Protein, BH3 Interacting Domain Death Agonist Protein, Protein Binding
Abstract

Caspase-8 cleaves BID to tBID, which targets mitochondria and induces oligomerization of BAX and BAK within the outer membrane, resulting in release of cytochrome c from the organelle. Here, we have initiated these steps in isolated mitochondria derived from control and BCL-2-overexpressing cells using synthetic BH3 peptides and subsequently analyzed the BCL members by chemical cross-linking. The results show that the BH3 domain of BID interacts with and induces an "open" conformation of BAK, exposing the BAK N terminus. This open (activated) conformer of BAK potently induces oligomerization of non-activated ("closed") conformers, causing a cascade of BAK auto-oligomerization. Induction of the open conformation of BAK occurs even in the presence of excess BCL-2, but BCL-2 selectively interacts with this open conformer and blocks BAK oligomerization and cytochrome c release, dependent on the ratio of BID BH3 and BCL-2. This mechanism of inhibition by BCL-2 also occurs in intact cells stimulated with Fas or expressing tBID. Although BID BH3 interacts with both BCL-2 and BAK, the results indicate that when BCL-2 is in excess it can sequester the BID BH3-induced activated conformer of BAK, effectively blocking downstream events. This model suggests that the primary mechanism for BCL-2 blockade targets activated BAK rather than sequestering tBID.

Published
2003
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2. Caspases Cleave Focal Adhesion Kinase during Apoptosis to Generate a FRNK-like Polypeptide

Author

Sophie Roy, Nancy A. Thornberry, François G. Gervais, Salvatore C. Ruffolo, and Donald W. Nicholson

Subjects
Molecular Sequence Data, PTK2, Integrin, Apoptosis, Biochemistry, Extracellular matrix, Focal adhesion, Jurkat Cells, Animals, Humans, Amino Acid Sequence, Molecular Biology, Paxillin, Caspase, Base Sequence, Sequence Homology, Amino Acid, biology, Caspase 3, Kinase, Hydrolysis, Cell Biology, Protein-Tyrosine Kinases, Molecular biology, Cell biology, Cysteine Endopeptidases, Caspases, Focal Adhesion Kinase 1, Focal Adhesion Protein-Tyrosine Kinases, Mutagenesis, Site-Directed, biology.protein, biological phenomena, cell phenomena, and immunity, Cell Adhesion Molecules, Chickens, Oligopeptides
Abstract

Focal adhesion kinase (Fak) is a non-receptor protein-tyrosine kinase that stimulates cell spreading and motility by promoting the formation of contact sites between the cell and the extracellular matrix (focal adhesions). It suppresses apoptosis by transducing survival signals that emanate from focal adhesions via the clustering of transmembrane integrins by components of the extracellular matrix. We demonstrate that Fak is cleaved by caspases at two distinct sites during apoptosis. The sites were mapped to DQTD772, which was preferentially cleaved by caspase-3, and VSWD704, which was preferentially cleaved by caspase-6 and cytotoxic T lymphocyte-derived granzyme B. The cleavage of Fak during apoptosis separates the tyrosine kinase domain from the focal adhesion targeting (FAT) domain. The carboxyl-terminal fragments that are generated suppress phosphorylation of endogenous Fak and thus resemble a natural variant of Fak, FRNK, that inhibits Fak activity by preventing the localization of Fak to focal adhesions. The cleavage of Fak by caspases may thus play an important role in the execution of the suicide program by disabling the anti-apoptotic function of Fak. Interestingly, rodent Fak lacks an optimal caspase-3 consensus cleavage site although it is cleaved in murine cells undergoing apoptosis at an upstream site. This appears to be the first example of a caspase substrate where the cleavage sites are not conserved between species.

Published
1998
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6. Basal Activation of p7056k Results in Adipose-specific Insulin Resistance in Protein-tyrosine Phosphatase 1B-/- Mice.

Author

Ruffolo, Salvatore C., Forsell, Pontus K. A., Xiling Yuan, Desmarais, Sylvie, Himms-Hagen, Jean, Cromlish, Wanda, Wong, Kenny K., and Kennedy, Brian P.

Subjects
*PHOSPHATASES, *HYPOGLYCEMIC agents, *PANCREATIC secretions, *CHEMICAL reactions, *BIOMOLECULES, *LIFE sciences, *BIOCHEMISTRY
Abstract

Although protein-tyrosine phosphatase 1B (PTP-1B) is a negative regulator of insulin action, adipose tissue from PTP-1B-/- mice does not show enhanced insulin-stimulated insulin receptor phosphorylation. Investigation of glucose uptake in isolated adipocytes revealed that the adipocytes from PTP-1B-/- mice have a significantly attenuated insulin response as compared with PTP-1B-/- adipocytes. This insulin resistance manifests in PTP-lB! animals older than 16 weeks of age and could be partially rescued by adenoviral expression of PTP-1B in null adipocytes. Examination of adipose signaling pathways found that the basal p70S6K activity was at least 50% higher in adipose from PTP-1B-/- mice compared with wild type animals. The increased basal activity of p70S6K in PTP-1B-/- adipose corre- lated with decreases in IR substrate-1 protein levels and insulin- stimulated Akt/protein kinase B activity, explaining the decrease in insulin sensitivity even as insulin receptor phosphorylation was unaffected. The insulin resistance of the of the PTP-1B-/- adipocytes could also be rescued by treatment with rapamycin, suggesting that in adipose the loss of PTP-1B results in basal activation of mTOR (mammalian target of rapamycin) complex 1 leading to a tissue-specific insulin resistance. [ABSTRACT FROM AUTHOR]

Published
2007
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7. Disruption of the A(3) adenosine receptor gene in mice and its effect on stimulated inflammatory cells.

Author

Salvatore, C A, Tilley, S L, Latour, A M, Fletcher, D S, Koller, B H, and Jacobson, M A

Abstract

The A(3) adenosine receptor (A3AR) is one of four receptor subtypes for adenosine and is expressed in a broad spectrum of tissues. In order to study the function of A3AR, a mouse line carrying a mutant A(3) allele was generated. Mice homozygous for targeted disruption of the A3AR gene, A3AR(-/-), are fertile and visually and histologically indistinguishable from wild type mice. The lack of a functional receptor in the A3AR(-/-) mice was confirmed by molecular and pharmacological analyses. The absence of A3AR protein expression in the A3AR(-/-) mice was demonstrated by lack of N(6)-(4-amino-3-[(125)I]iodobenzyl)adenosine binding to bone marrow-derived mast cell membranes that were found to express high levels of A3AR in wild type mice. In A3AR(-/-) mice, the density of A(1) and A(2A) adenosine receptor subtypes was the same as in A3AR(+/+) mice as determined by radioligand binding to brain membranes. Additionally, A(2B) receptor transcript expression was not affected by ablation of the A3AR gene. A3AR(-/-) mice have basal heart rates and arterial blood pressures indistinguishable from A3AR(+/+) mice. Functionally, in contrast to wild type mice, adenosine and the A3AR-specific agonist, 2-chloro-N(6)-(3-iodobenzyl)-adenosine-5'-N-methyl-carboxamide (2-Cl-IB-MECA), elicit no potentiation of antigen-dependent degranulation of bone marrow-derived mast cells from A3AR(-/-) mice as measured by hexosaminidase release. Also, the ability of 2Cl-IB-MECA to inhibit lipopolysaccharide-induced tumor necrosis factor-alpha production in vivo was decreased in A3AR(-/-) mice in comparison to A3AR(+/+) mice. The A(2A) adenosine receptor agonist, 2-p-(2-carboxyethyl)phenylamino)-5'-N-ethylcarboxamidoadenosine, produced inhibition of lipopolysaccharide-stimulated tumor necrosis factor-alpha production in both A3AR(-/-) and A3AR(+/+) mice. These results show that the inhibition in vivo can be mediated by multiple subtypes, specifically the A(3) and A(2A) adenosine receptors, and A3AR activation plays an important role in both pro- and anti-inflammatory responses.

Published
2000

8. Abrogation of nerve growth factor-induced terminal differentiation by ret oncogene involves perturbation of nuclear translocation of ERK.

Author

Colucci-D'Amato GL, D'Alessio A, Califano D, Cali G, Rizzo C, Nitsch L, Santelli G, and de Franciscis V

Subjects
Animals, Biological Transport, Gene Expression Regulation physiology, PC12 Cells, Promoter Regions, Genetic, Proto-Oncogene Proteins c-fos genetics, Proto-Oncogene Proteins c-ret, Rats, Signal Transduction, Transfection, Cell Differentiation physiology, Cell Nucleus metabolism, Drosophila Proteins, Mitogen-Activated Protein Kinases metabolism, Nerve Growth Factor physiology, Oncogenes, Proto-Oncogene Proteins genetics, Receptor Protein-Tyrosine Kinases genetics
Abstract

Oncogenic variants of the receptor tyrosine kinase, Ret, cause formation of tumors of neuroendocrine derivation in the multiple endocrine neoplasia type 2 and, thus, likely interfere with antiproliferative and/or differentiative extracellular signals. Here we took advantage of two rat pheochromocytoma-derived cell lines (PC12/MEN2A and PC12/MEN2B) to investigate whether Ret-induced nerve growth factor (NGF) unresponsiveness might involve impairment of ERK signaling. In fact, these cells, stably transfected with distinct forms of the active ret oncogene, fail to block proliferation, even upon NGF stimulation. In these cells we show the presence of both chronic ERKs activity and high expression levels of MKP-3, an ERK-specific phosphatase. Despite the presence of MKP-3, ERK activity can be further stimulated by NGF, but it fails to translocate into the nucleus and consequently to induce immediate-early gene transcription. Because of the presence of MKP-3, our results suggest the existence of a negative regulatory feedback acting on ERKs as a mechanism responsible for the abrogation of NGF-induced terminal differentiation. Indeed, MKP-3 seems to be implicated in the persistence of ERKs in cell cytoplasm. This interpretation is further supported by the observation that in ret-transfected cells, forced expression of an active form of MEK-1 may overcome this block; it restores transcription from the c-fos promoter, induces translocation of ERKs into the nucleus, and inhibits cell proliferation.

Published
2000
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