Strain-specific and tissue-specific expression of mouse mast cell secretory granule proteases.

Previous studies have revealed that activin A and transforming growth factor-␤ 1 (TGF-␤ 1 ) induced migration and morphological changes toward differentiation in bone marrow-derived cultured mast cell progenitors (BMCMCs). Here we show up-regulation of mouse mast cell protease-7 (mMCP-7), which is expressed in differentiated mast cells, by activin A and TGF-␤ 1 in BMCMCs, and the molecular mechanism of the gene induction of mmcp-7. Smad3, a signal mediator of the activin/TGF-␤ pathway, transcriptionally activated mmcp-7. Microphthalmia-associated transcription factor (MITF), a tissue-specific transcription factor predominantly expressed in mast cells, melanocytes, and heart and skeletal muscle, inhibited Smad3-mediated mmcp-7 transcription. MITF associated with Smad3, and the C terminus of MITF and the MH1 and linker region of Smad3 were required for this association. Complex formation between Smad3 and MITF was neither necessary nor sufficient for the inhibition of Smad3 signaling by MITF. MITF inhibited the transcriptional activation induced by the MH2 domain of Smad3. In addition, MITF-truncated N-terminal amino acids could associate with Smad3 but did not inhibit Smad3-mediated transcription. The level of Smad3 was decreased by co-expression of MITF but not of dominantnegative MITF, which resulted from proteasomal protein degradation. The changes in the level of Smad3 protein were paralleled by those in Smad3-mediated signaling activity. These findings suggest that MITF negatively regulates Smad-dependent activin/TGF-␤ signaling in a tissue-specific manner.

Section: Discussionmentioning

confidence: 99%

Transcriptional Activation of Mouse Mast Cell Protease-7 by Activin and Transforming Growth Factor-β Is Inhibited by Microphthalmia-associated Transcription Factor

Funaba¹,

Ikeda

Murakami

et al. 2003

“…Preliminary studies confirmed that MC-deficient W/W v mice possess a greatly diminished ability to kill KP1415 bacteria relative to MC-sufficient WBB6F1 ϩ/ϩ mice or C57BL/6 mice (data not shown). It has been shown previously that the MCs in WBB6F1 ϩ/ϩ and C57BL/6 mice differ in their expression of mMCP-7, but not of mMCP-6 (47)(48)(49). The finding that WBB6F1 ϩ/ϩ and C57BL/6 mice do not differ significantly in their ability to combat a KP1415 bacterial infection now indicates that the primary immunologic defect in MC-deficient W/W v mice is not a consequence of their failure to express mMCP-7.…”

Section: Isolated From a Phage Display Peptide Librarymentioning

confidence: 99%

Evaluation of the Substrate Specificity of Human Mast Cell Tryptase βI and Demonstration of Its Importance in Bacterial Infections of the Lung

Huang¹,

Sanctis²,

O’Brien³

et al. 2001

139

132

Human pulmonary mast cells (MCs) express tryptases ␣ and ␤I, and both granule serine proteases are exocytosed during inflammatory events. Recombinant forms of these tryptases were generated for the first time to evaluate their substrate specificities at the biochemical level and then to address their physiologic roles in pulmonary inflammation. Analysis of a tryptase-specific, phage display peptide library revealed that tryptase ␤I prefers to cleave peptides with 1 or more Pro residues flanked by 2 positively charged residues. Although recombinant tryptase ␤I was unable to activate cultured cells that express different types of protease-activated receptors, the numbers of neutrophils increased >100-fold when enzymatically active tryptase ␤I was instilled into the lungs of mice. In contrast, the numbers of lymphocytes and eosinophils in the airspaces did not change significantly. More important, the tryptase ␤I-treated mice exhibited normal airway responsiveness. Neutrophils did not extravasate into the lungs of tryptase ␣-treated mice. Thus, this is the first study to demonstrate that the two nearly identical human MC tryptases are functionally distinct in vivo. When MCdeficient W/W v mice were given enzymatically active tryptase ␤I or its inactive zymogen before pulmonary infection with Klebsiella pneumoniae, tryptase ␤I-treated W/W v mice had fewer viable bacteria in their lungs relative to zymogen-treated W/W v mice. Because neutrophils are required to combat bacterial infections, human tryptase ␤I plays a critical role in the antibacterial host defenses of the lung by recruiting neutrophils in a manner that does not alter airway reactivity.

“…These three MC tryptases also can be differentially expressed within specific tissues of the same mouse strain. For example, the safranin ϩ MCs in the skin of BALB/cJ mice contain abundant amounts of mMCP-6 and mMCP-7, whereas the safranin ϩ MCs in the peritoneal cavity of this mouse strain express mMCP-6 but not mMCP-7 (29). The intestines of C57BL/6J mice also contain more Prss31 than mMCP-6 or mMCP-7 (15).…”

Section: Expression Of Tryptase Genes In Mcsmentioning

confidence: 99%

Mast Cell-restricted Tryptases: Structure and Function in Inflammation and Pathogen Defense

McNeil

Adachi

Stevens

2007

Mast cells (MCs) are highly specialized immune cells present in mammals and in lower organisms that predate the development of adaptive immunity. The strong evolutionary pressure to retain MCs for >500 million years suggests critical roles for these cells in our survival. In support of this conclusion, no human has been identified to date that lacks MCs, despite the adverse roles of MCs in systemic anaphylaxis and varied inflammatory disorders. MCs express numerous lineage-restricted neutral proteases, and four members of the chromosome 17A3.3 family of tryptases are preferentially expressed in mouse MCs. The anatomical location of MCs at host-environment interfaces has raised the possibility that some of these enzymes are evolutionally conserved because they are needed for combating infectious organisms. Here we review recent insights into the structure and function of MC tryptases in inflammation and host defense against bacteria and other infectious organisms. MC-restricted Granule Proteases and ProteoglycansA characteristic morphologic feature of mammalian mast cells (MCs) 3 is their electron-dense secretory granules, which contain serglycin proteoglycans (1, 2) bearing heparin (for review see Ref. 3) and chondroitin sulfates diB and E (4, 5). Heparin is the most negatively charged molecule in the body, and each rat and mouse peritoneal MC contains ϳ25 pg of this glycosaminoglycan, thereby explaining the avidity of MCs for cationic dyes (6). The repeating Ser-Gly sequence in serglycin where its glycosaminoglycans are attached cannot be cleaved by any known protease. This protease resistance feature is biologically relevant because the primary function of serglycin proteoglycans in MCs is to provide a scaffold for formation of macromolecular complexes with the cell's varied granule proteases. When properly folded, a positively charged face forms on the surface of each granule protease that allows it to bind tightly to the proteoglycan's negatively charged glycosaminoglycans (7-9).The proteases packaged in the secretory granules of mouse MCs include carboxypeptidase A3 (CPA3), granzyme B, cathepsin G, neuropsin, transmembrane tryptase/tryptase ␥/protease serine member S (Prss) 31, mouse MC protease (mMCP) 1-10, and mMCP-11/Prss34. Definitive evidence of the importance of heparin-containing serglycin proteoglycans in the packaging of specific cassettes of proteases in the MC's granules came with the characterization of N-deacetylase/Nsulfotransferase-2 (NDST-2)-null mice (10, 11). NDST-2 is essential for heparin biosynthesis and is preferentially expressed in MCs. Because of the loss of fully sulfated heparin, the MCs in the skin and peritoneal cavities of NDST-2-null mice store reduced amounts of mMCP-6 and almost no CPA3, mMCP-4, and mMCP-5, even though all four MC-restricted protease genes are transcribed at normal rates. The accumulated data led to the current view that serglycin proteoglycans are essential for the post-translational processing and granule accumulation of most, if not all, proteases stor...