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R Numerof

Publications and source records attributed to R Numerof.

10 recordsLinked to original sources
PubMed2001

Co-cultured human mast cells stimulate fibroblast-mediated contraction of collagen gels.

In the current study, we asked whether mast cells might modulate remodeling of extracellular matrix by affecting fibroblast-mediated contraction of three-dimensional collagen gels. Mast cells and human lung fibroblasts were co-cultured in floating type I collagen gels. The area of the gels was measured by an image analyzer. Mast cells in co-culture augmented fibroblast contractility (P < 0.001) in a time- and concentration dependent manner. The tryptase inhibitor bis(5-amidino-2-benzimidazo-lyl)methane (BABIM) were unable to block the augmented fibroblast contractility induced by co-cultured mast cells and tryptase added alone in the culture system had no effect on contractility, suggesting that other mediators besides tryptase might be involved. The amount of collagen in dissolved gels, measured as hydroxyproline, did not change after co-culture indicating that degradation of collagen may not be a major mechanism. Our findings support the hypothesis that the activity of mast cells may drive rearrangement of extracellular matrix and this and could subsequently lead to fibrosis and tissue dysfunction.

Animals↗
PubMed1997

Interactions of mast cell tryptase with thrombin receptors and PAR-2.

Tryptase is a serine protease secreted by mast cells that is able to activate other cells. In the present studies we have tested whether these responses could be mediated by thrombin receptors or PAR-2, two G-protein-coupled receptors that are activated by proteolysis. When added to a peptide corresponding to the N terminus of PAR-2, tryptase cleaved the peptide at the activating site, but at higher concentrations it also cleaved downstream, as did trypsin, a known activator of PAR-2. Thrombin, factor Xa, plasmin, urokinase, plasma kallikrein, and tissue kallikrein had no effect. Tryptase also cleaved the analogous thrombin receptor peptide at the activating site but less efficiently. When added to COS-1 cells expressing either receptor, tryptase stimulated phosphoinositide hydrolysis. With PAR-2, this response was half-maximal at 1 nM tryptase and could be inhibited by the tryptase inhibitor, APC366, or by antibodies to tryptase and PAR-2. When added to human endothelial cells, which normally express PAR-2 and thrombin receptors, or keratinocytes, which express only PAR-2, tryptase caused an increase in cytosolic Ca2+. However, when added to platelets or CHRF-288 cells, which express thrombin receptors but not PAR-2, tryptase caused neither aggregation nor increased Ca2+. These results show that 1) tryptase has the potential to activate both PAR-2 and thrombin receptors; 2) for PAR-2, this potential is realized, although cleavage at secondary sites may limit activation, particularly at higher tryptase concentrations; and 3) in contrast, although tryptase clearly activates thrombin receptors in COS-1 cells, it does not appear to cleave endogenous thrombin receptors in platelets or CHRF-288 cells. These distinctions correlate with the observed differences in the rate of cleavage of the PAR-2 and thrombin receptor peptides by tryptase. Tryptase is the first protease other than trypsin that has been shown to activate human PAR-2. Its presence within mast cell granules places it in tissues where PAR-2 is expressed but trypsin is unlikely to reach.

Amino Acid Sequence↗
PubMed1996

Syk-dependent phosphorylation of Shc. A potential link between FcepsilonRI and the Ras/mitogen-activated protein kinase signaling pathway through SOS and Grb2.

Antigen receptors on T- and B-cells activate Ras through a signaling pathway that results in the tyrosine phosphorylation of Shc and the formation of a complex of Shc with the Grb2 adaptor protein. The high affinity receptor for immunoglobulin E (FcepsilonRI) in cultured mast (RBL-2H3) cells has been reported to function differently. Here we show to the contrary that engagement of FcepsilonRI with antigen leads to increased tyrosine phosphorylation of Shc and the association of Shc with Grb2 and other proteins (p120 and p140). Like the FcepsilonRI-mediated activation of the mitogen-activated protein kinase cascade, these responses are dependent on the tyrosine kinase Syk; they are enhanced by overexpression of Syk and are blocked by expression of dominant-negative Syk. Sos is constitutively associated with Grb2 in these cells but dissociates from Shc on stimulation with antigen. These reactions are rapid, reversible, and associated with the activation of Ras. Therefore, the Syk-dependent tyrosine phosphorylation of Shc and its association with Grb2 may provide a pathway through Sos for activation of Ras by FcepsilonRI.

Adaptor Proteins, Signal Transducing↗
PubMed1996

IgE receptor (Fc epsilon RI) and signal transduction.

This review suggests a model in which both beta- and gamma-chains synergize in the initiation of Fc epsilon RI signal transduction function. Receptor aggregation by antigens induces activation of lyn, which is already bound to the Fc epsilon RI beta-chain under resting conditions. Whilst activated, lyn would phosphorylate the tyrosine residues in the Fc epsilon RI gamma-chain. This phosphorylation would be responsible for the recruitment of syk (probably via its SH2 domains) as well as other signalling molecules. Syk kinase would then be activated by the engagement of its SH2 domains and/or its phosphorylation. Syk could then interact with and activate (through phosphorylation) downstream effector molecules.

Enzyme Precursors↗
PubMed1994

Differential control of the tyrosine kinases Lyn and Syk by the two signaling chains of the high affinity immunoglobulin E receptor.

Nonreceptor tyrosine kinases such as the newly described 70-kDa (ZAP-70/Syk) and Src-related tyrosine kinases are coupled to a variety of receptors, including the antigen receptors on B- and T-cells and the Fc receptors for IgE (Fc epsilon RI) and IgG (Fc gamma RI, Fc gamma RIII/CD16). Various subunits of these receptors contain homologous activation motifs which appear capable of autonomously triggering cell activation. Two forms of this motif are present in the Fc epsilon RI multimeric complex: one in the beta chain and one in the gamma chain. Here we show that each of the two tyrosine kinases known to be involved in Fc epsilon RI signaling is controlled by a distinct motif-containing chain. Lyn associates with the nonactivated beta chain, whereas gamma promotes the activation of Syk. We also show that neither the beta nor the gamma motif alone can account for the full signaling capacity of the entire receptor. We propose that, upon triggering of the tetrameric receptor, Lyn already bound to beta becomes activated and phosphorylates beta and gamma; the phosphorylation of gamma induces the association of Syk with gamma and also the activation of Syk, resulting in the phosphorylation and activation of phospholipase C gamma 1. Cooperative recruitment of specific kinases by the various signaling chains found in this family of antigen receptors could represent a way to achieve the full signaling capacity of the multimeric complexes.

Amino Acid Sequence↗
PubMed1994

Kinase activation through the high-affinity receptor for immunoglobulin E.

The high-affinity receptor for IgE (Fc epsilon RI) belongs to a class of multimeric receptors associated with nonreceptor tyrosine kinases. It has been assumed that Fc epsilon RI beta and gamma chains, which have extensive cytoplasmic domains, play an important, although undefined role in coupling the receptor to signal transduction mechanisms. The results reviewed here suggest a synergistic effect of these two chains in the initiation of Fc epsilon RI signaling. According to our model, receptor engagement can activate kinase(s), such as lyn, already bound to the receptor under resting conditions. The receptor phosphorylation following this activation can be responsible for recruitment and activation of other signaling molecules, such as syk, which can then activate downstream effector molecules. This model could be extended to include other multimeric receptors, such as the T- and B-cell receptors and the low-affinity receptor for IgG (Fc gamma RIII), that control the activation of cytoplasmic tyrosine kinases.

Amino Acid Sequence↗
PubMed1992

Phosphorylation/dephosphorylation of high-affinity IgE receptors: a mechanism for coupling/uncoupling a large signaling complex.

Engagement of high-affinity IgE receptors leads to activation of tyrosine and serine/threonine kinases and the immediate phosphorylation of receptor beta (serine and tyrosine) and gamma (threonine and tyrosine) chains. Receptor disengagement leads to dephosphorylation of beta and gamma chains via the action of undefined phosphatases. Here we have identified five distinct polypeptides associated with the high-affinity IgE-receptor tetrameric complex, which apparently become phosphorylated and dephosphorylated in sequence with the beta and gamma chains. Like beta chain, polypeptides pp180, pp48, pp42, and pp28 are phosphorylated on serine and tyrosine, whereas pp125 is only phosphorylated on serine. The phosphorylation of each of these receptor-associated polypeptides is antigen-dose dependent and is restricted to activated receptor complexes. Furthermore the physical association between pp125 and the receptor is quantitatively affected by receptor phosphorylation and dephosphorylation, indicating a coupling-uncoupling mechanism. Finally, in vitro kinase experiments show that activated receptor complexes are also physically associated with tyrosine and serine/threonine kinases as part of a larger complex containing the phosphorylated polypeptides.

Animals↗