| الوصف: |
Macrophages are terminally differentiated cells of the mononuclear phagocyte lineage and are specialized for locomotion and phagocytosis. Colony-stimulating factor 1 (CSF-1) is the primary regulator of the survival, proliferation, and differentiation of mononuclear phagocytic cells (12, 39) as well as macrophage morphology and motility (9, 45). Cells of the BAC1.2F5 mouse macrophage cell line have retained many characteristics of primary macrophages, including dependence on CSF-1 for survival and proliferation (30) and a pleiomorphic but adherent phenotype. When starved of CSF-1, BAC1.2F5 cells round up, retract their pseudopodia (9), and eventually die. With the addition of CSF-1 to quiescent cells, the CSF-1 receptor (CSF-1R [c-fms]) is rapidly activated, leading to receptor autophosphorylation and tyrosine phosphorylation of a number of cytoskeletal proteins and cytoplasmic proteins associated with signaling (26, 47). Morphological changes are also effected rapidly upon CSF-1 stimulation, with macrophage spreading and extension of lamellipodia and formation of ruffles on the cell surface (9), followed by cell polarization and increased motility (3, 45). Tyrosine phosphorylation is a critical control mechanism for the regulation of cell survival, proliferation, morphology, and motility. The balance of cellular tyrosine phosphorylation is controlled by the coordinated actions of protein tyrosine kinases and protein tyrosine phosphatases (PTPs). A number of PTPs have been implicated in receptor tyrosine kinase (RTK) signaling and proliferative pathways, in particular the cytosolic, SH2 domain-containing PTPs SHP-1 and SHP-2 (42). Cell morphology and motility are regulated both by RTK-linked intracellular signaling pathways and by interactions between the cell and its extracellular matrix (ECM) (29). ECM ligand-mediated activation of integrin cell adhesion receptors (24) triggers tyrosine phosphorylation in a variety of cell lines (15), including the tyrosine phosphorylation of focal adhesion kinase (FAK), Pyk2, paxillin, and p130cas (6, 11, 17, 32). These phosphorylated proteins are concentrated at points of interaction between the ECM and the cytoskeleton, known as focal contacts. Paxillin is one of the more highly tyrosine phosphorylated focal adhesion proteins and requires the coordinated actions of both the FAK/Pyk2 and the Src family kinases to be fully tyrosine phosphorylated (36). As in more rigid cells such as fibroblasts, the dynamic shape and motility of macrophages is controlled by cell-ECM interactions. However, their focal contacts are much smaller and are termed focal complexes (2). Tyrosine kinases implicated in the phosphorylation of focal adhesion proteins and in focal adhesion formation include FAK, Pyk2, and the Src family kinases (15). Fewer PTPs are known to dephosphorylate focal adhesion proteins, although two cytoplasmic PTPs, PTP-PEST and PTP1B, have been shown to specifically dephosphorylate p130cas (16, 19, 27). The cytoplasmic enzymes PTP-PEST, PTP1B, and SHP-2 and the tumor suppressor PTEN appear to play important roles in fibroblast focal contact formation, spreading, and migration (4, 5, 20, 21, 27, 48), while SHP-1 deficiency results in an increase in macrophage spreading (34). Several other PTPs have been shown to be localized to focal contacts or important in cell adhesion. LAR is a membrane-spanning PTP with fibronectin type III (FNIII) repeats in its extracellular domain (ECD) (40). It is localized to focal contacts in a distribution that suggests it may be important in their disassembly (38). FNIII repeats may be involved in cell adhesion (37, 43). The PTPμ family of PTPs, which contain FNIII repeats in their ECDs, interact homophilically to mediate cell-cell aggregation (10, 14, 35). Several Drosophila PTPs with FNIII repeats in their ECDs and a mammalian PTP, PTPζ/RPTPβ, have been implicated in neuronal adhesion and motor axon guidance (42). PTPφ is a complex PTP with at least five different isoforms (33, 41). The largest isoforms each contain an identical large ECD with eight FNIII repeats and are expressed in brain and kidney. The remaining isoforms of PTPφ, which are selectively expressed in macrophages (33) and B cells (1), comprise two membrane-spanning molecules with an identical, very short ECD lacking FNIII repeats and a cytosolic enzyme. Thus, PTPφ could be involved in cell-cell adhesion of stationary cells in the brain and kidney yet may mediate different functions, such as motility and/or phagocytosis, in macrophages. In the present study, we have analyzed BAC1.2F5 macrophages overexpressing either the wild-type (WT) or a phosphatase-inactive (C325S) form of the larger membrane-spanning isoform of PTPφ in order to understand its function. |
