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SUMMARY Cell fusion is essential for fertilization, myotube formation, and inflammation. Macrophages fuse in various circumstances but the molecular signals involved in the distinct steps of their fusion are not fully characterized. Using null mice and derived cells, we show that the protease MT1-MMP is necessary for macrophage fusion during osteoclast and giant cell formation in vitro and in vivo. Specifically, MT1-MMP is required for lamellipodia formation and for proper cell morphology and motility of bone marrow myeloid progenitors prior to membrane fusion. These functions of MT1-MMP do not depend on MT1-MMP catalytic activity or downstream pro-MMP-2 activation. Instead, MT1-MMP-null cells show a decreased Rac1 activity and reduced membrane targeting of Rac1 and the adaptor protein p130Cas. Retroviral rescue experiments and protein binding assays delineate a signaling pathway in which MT1-MMP, via its cytosolic tail, contributes to macrophage migration and fusion by regulating Rac1 activity through an association with p130Cas.

Tumorigenesis involves not only tumor cells that become transformed but also the peritumoral stroma which reacts inducing inflammatory and angiogenic responses. Angiogenesis, the formation of new capillaries from preexisting vessels, is an absolute requirement for tumor growth and metastasis, and it can be induced and modulated by a wide variety of soluble factors. During angiogenesis, quiescent endothelial cells are activated and they initiate migration by degrading the basement membranes through the action of specific proteases, in particular of matrix metalloproteinases (MMPs). Among these, the membrane type 1-matrix metalloproteinase (MT1-MMP) has been identified as a key player during the angiogenic response. In this review, we will summarize the role of MT1-MMP in angiogenesis and the regulatory mechanisms of this protease in endothelial cells. Since our recent findings have suggested that MT1-MMP is not universally required for angiogenesis, we hypothesize that the regulation and participation of MT1-MMP in angiogenesis may depend on the nature of the angiogenic stimulus. Experiments aimed at testing this hypothesis have shown that similarly to the chemokine stromal cell-derived factor-1 (SDF-1)/CXCL12, lipopolysaccharide (LPS) seems to induce the formation of capillary tubes by human or mouse endothelial cells (ECs) in an MT1-MMP-independent manner. The implications of these findings in the potential use of MT1-MMP inhibitors in cancer therapy are discussed.

The retinoid X receptor α (RXRα) plays a central role in the regulation of many intracellular receptor signaling pathways and can mediate ligand-dependent transcription by forming homodimers or heterodimers with other nuclear receptors. Although several members of the nuclear hormone receptor superfamily have emerged as important regulators of macrophage gene expression, the existence in vivo of an RXR signaling pathway in macrophages has not been established. Here, we provide evidence that RXRα regulates the transcription of the chemokines Ccl6 and Ccl9 in macrophages independently of heterodimeric partners. Mice lacking RXRα in myeloid cells exhibit reduced levels of CCL6 and CCL9, impaired recruitment of leukocytes to sites of inflammation, and lower susceptibility to sepsis. These studies demonstrate that macrophage RXRα plays key roles in the regulation of innate immunity and represents a potential target for immunotherapy of sepsis.nuclear hormone receptors | macrophages | innate immunity | sepsis

MT1-MMP plays IntroductionMT1-MMP is an important proteinase, with both direct actions on target substrates and indirect actions through its activation of a metalloproteinase cascade involving matrix metalloproteinase 2 (MMP2). Fibroblasts and tumor cells derived from MT1-MMPdeficient mice show impaired collagenolytic activity. 1 MT1-MMP is essential for bone maturation and lung development, but MT1-MMP function in other processes seems to be compensated in knockout mice by other metalloproteinases. 2 Supporting this, the phenotype of MMP2/MT1-MMP double-knockout mice is more severe, and these mice die in the first hours after birth. 3 MT1-MMP gene expression is regulated by signaling via ␤catenin/LEF4, Egr, NFAT, and Rac. Maturation by furin cleavage of the prodomain in MT1-MMP takes place intracellularly, so that the metalloproteinase is already mature when it reaches the plasma membrane. Hence, most regulation of MT1-MMP occurs at the plasma membrane. 4,5 MT1-MMP is inhibited by tissue inhibitors of matrix proteinases 2-4 (TIMPs 2-4), although TIMP2 is also necessary for the appropriate activation of MMP2 by MT1-MMP and forms a ternary complex with the 2 proteases. Other secreted proteins such as testican have also been shown to inhibit MT1-MMP activity. The membrane glycoprotein reversion-inducing cysteine-rich protein with Kazal motifs (RECK) is an inhibitor of MT1-MMP, MMP2, and MMP9. 6 In addition, MT1-MMP activity is self-regulated by its autocatalytic processing, which removes the catalytic domain and may act as a negative regulatory mechanism. 7 Finally, MT1-MMP activity and turnover are regulated by hemopexin-dependent oligomerization. 8 Intracellular trafficking of MT1-MMP is necessary for its participation in cell migration and is thought to replenish the plasma membrane with active new MT1-MMP molecules via a Rab8-dependent exocytic pathway. 9 Thus, cells expressing MT1-MMP mutants, which fail to internalize, show enhanced MMP2 activation but a reduced invasive capability. 6 Internalization of MT1-MMP occurs both via the clathrin and the caveolae pathways, 10,11 and MT1-MMP internalization is regulated by its inclusion in lipid rafts. 12 Cell membrane tetraspanin-based microdomains are generated by lateral association of tetraspanin proteins with several other transmembrane proteins, including integrins and immunoglobulin (Ig) superfamily members. 13 Tetraspanins are key regulators of the functions and signaling activities of their associated partners in diverse cellular processes. Tetraspanin interactions have been reported for several proteases. For example, tetraspanin CD151 binds soluble pro-MMP7, facilitating its maturation. 14 However, only MT1-MMP 15 and some ADAM (A Disintegrin and Metalloproteinase) proteins 16,17 have been shown to be included in tetraspanin microdomains. Human umbilical vein endothelial cells (HUVECs) express a repertoire of tetraspanins, including CD9, CD81, and CD151, which are localized mainly at cell-cell junctions in association with ␣3␤1 integrin. 18 The tight stoi...

Hutchinson–Gilford progeria syndrome (HGPS) is a rare genetic disorder caused by progerin, a mutant lamin A variant. HGPS patients display accelerated aging and die prematurely, typically from atherosclerosis complications. Recently, we demonstrated that progerin‐driven vascular smooth muscle cell (VSMC) loss accelerates atherosclerosis leading to premature death in apolipoprotein E‐deficient mice. However, the molecular mechanism underlying this process remains unknown. Using a transcriptomic approach, we identify here endoplasmic reticulum stress (ER) and the unfolded protein responses as drivers of VSMC death in two mouse models of HGPS exhibiting ubiquitous and VSMC‐specific progerin expression. This stress pathway was also activated in HGPS patient‐derived cells. Targeting ER stress response with a chemical chaperone delayed medial VSMC loss and inhibited atherosclerosis in both progeria models, and extended lifespan in the VSMC‐specific model. Our results identify a mechanism underlying cardiovascular disease in HGPS that could be targeted in patients. Moreover, these findings may help to understand other vascular diseases associated with VSMC death, and provide insight into aging‐dependent vascular damage related to accumulation of unprocessed toxic forms of lamin A.

The vaccinia-related kinase (VRK) proteins are a new group of three Ser-Thr kinases in the human kinome. VRK proteins are upstream regulators of several transcription factors. VRK1 phosphorylates p53 in Thr-18 within the region of binding to mdm2 preventing their interaction. The tissue distribution of three genes is still largely unknown. In the present report the expression of these genes was analyzed during murine hematopoietic development. The three genes are expressed in fetal liver and peripheral blood, with higher levels between days 11.5 and 13.5, a time when there is a massive expansion of liver cells, and thereafter their expression falls signi¢cantly. VRK genes are expressed, particularly at mid-gestation, in embryo thymus and spleen, but in adult thymus and spleen their levels are very low. VRK2 is expressed at lower levels than VRK1 and VRK3 in the mouse embryo. VRK genes play a role during embryonic development of hematopoiesis. ß

Dormant or slow-cycling tumor cells can form a residual chemoresistant reservoir responsible for relapse in patients, years after curative surgery and adjuvant therapy. We have adapted the pulse-chase expression of H2BeGFP for labeling and isolating slow-cycling cancer cells (SCCCs). SCCCs showed cancer initiation potential and enhanced chemoresistance. Cells at this slow-cycling status presented a distinctive nongenetic and cell-autonomous gene expression profile shared across different tumor types. We identified TET2 epigenetic enzyme as a key factor controlling SCCC numbers, survival, and tumor recurrence. 5-Hydroxymethylcytosine (5hmC), generated by TET2 enzymatic activity, labeled the SCCC genome in carcinomas and was a predictive biomarker of relapse and survival in cancer patients. We have shown the enhanced chemoresistance of SCCCs and revealed 5hmC as a biomarker for their clinical identification and TET2 as a potential drug target for SCCC elimination that could extend patients' survival.