The mouse ortholog of human FACE-1, Zmpste24, is a multispanning membrane protein widely distributed in mammalian tissues 1,2 and structurally related to Afc1p/ste24p, a yeast metalloproteinase involved in the maturation of fungal pheromones 3 . Disruption of the gene Zmpste24 caused severe growth retardation and premature death in homozygous-null mice. Histopathological analysis of the mutant mice revealed several abnormalities, including dilated cardiomyopathy, muscular dystrophy and lipodystrophy. These alterations are similar to those developed by mice deficient in A-type lamin 4 , a major component of the nuclear lamina 5 , and phenocopy most defects observed in humans with diverse congenital laminopathies 6-8 . In agreement with this finding, Zmpste24-null mice are defective in the proteolytic processing of prelamin A. This deficiency in prelamin A maturation leads to the generation of abnormalities in nuclear architecture that probably underlie the many phenotypes observed in both mice and humans with mutations in the lamin A gene. These results indicate that prelamin A is a specific substrate for Zmpste24 and demonstrate the usefulness of genetic approaches for identifying the in vivo substrates of proteolytic enzymes.To clarify the function of Zmpste24, and to identify in vivo substrates targeted by this proteinase, we generated Zmpste24-null mice (Fig. 1a). After heterozygote intercrossing, we obtained Zmpste24-null, heterozygous and wildtype mice in the expected mendelian ratio. We verified homozygosity with respect to the mutated allele by Southern blot (Fig. 1b), and lack of Zmpste24 Fig. 1 Generation of Zmpste24-deficient mice. a, Schematic representation of the wildtype Zmpste24 locus (WT), targeting vector and targeted allele (KO). Positions of restriction enzyme sites and probes used for Southern-blot analysis are shown. b, Southern-blot analysis of genomic DNA from three littermate progeny of Zmpste24 heterozygote crosses. Probing of EcoRI-digested DNA revealed fragments of 6 kb and 4 kb for wildtype and disrupted alleles, respectively. Probing of BamHI-digested DNA revealed fragments of 10 kb and 8 kb for wildtype and disrupted alleles, respectively. c, Extracts of kidneys from wildtype and Zmpste24 -/-mice were examined with a monoclonal antibody directed against the C terminus of Zmpste24. d, Photograph of two littermate progeny of a Zmpste24 heterozygote cross, at 3 mo. e, Radiograph of a Zmpste24 -/-mouse at 3 mo, compared with a wildtype control. f, Cumulative plot of body weight versus age. Dots represent mean values, and error bars indicate s.e.m.

Matrix metalloproteinases (MMPs) have fundamental roles in tumor progression, but most clinical trials with MMP inhibitors have not shown improvements in individuals with cancer. This may be partly because broad-range inhibitors also reduce host-protective antitumor properties of individual MMPs. We generated mice deficient in collagenase-2 (Mmp8), an MMP mainly produced by neutrophils in inflammatory reactions and detected in some malignant tumors. Loss of Mmp8 did not cause abnormalities during embryonic development or in adult mice. Contrary to previous studies with MMP-deficient mice, however, the absence of Mmp8 strongly increased the incidence of skin tumors in male Mmp8(-/-)mice. Female Mmp8(-/-)mice whose ovaries were removed or were treated with tamoxifen were also more susceptible to tumors compared with wild-type mice. Bone marrow transplantation experiments confirmed that Mmp8 supplied by neutrophils was sufficient to restore the natural protection against tumor development mediated by this protease in male mice. Histopathological analysis showed that mutant mice had abnormalities in the inflammatory response induced by carcinogens. Our study identifies a paradoxical protective role for Mmp8 in cancer and provides a genetic model to evaluate the molecular basis of gender differences in cancer susceptibility.

Proteolytic events at the cell surface are essential in the regulation of signal transduction pathways. During the past years, the family of type II transmembrane serine proteases (TTSPs) has acquired an increasing relevance because of their privileged localization at the cell surface, although our current understanding of the biologic function of most TTSPs is limited. Here we show that matriptase-2 (Tmprss6), a recently described member of the TTSP family, is an essential regulator of iron homeostasis. Thus, Tmprss6 Ϫ/Ϫ mice display an overt phenotype of alopecia and a severe iron deficiency anemia. These hematologic alterations found in Tmprss6 Ϫ/Ϫ mice are accompanied by a marked up-regulation of hepcidin, a negative regulator of iron export into plasma. Likewise, Tmprss6 Ϫ/Ϫ mice have reduced ferroportin expression in the basolateral membrane of enterocytes and accumulate iron in these cells. Iron-dextran therapy rescues both alopecia and hematologic alterations of Tmprss6 Ϫ/Ϫ mice, providing causal evidence that the anemic phenotype of these mutant mice results from the blockade of intestinal iron export into plasma after dietary absorption. On the basis of these findings, we conclude that matriptase-2 activity represents a novel and relevant step in hepcidin regulation and iron homeostasis. IntroductionPericellular proteolysis is an essential event that determines the relations between the cell and its microenvironment. This crucial process in the development and maintenance of multicellular organisms requires the remodeling of extracellular matrix components as well as the posttranslational regulation of a wide range of cell-surface receptors, regulatory proteins, and adhesion molecules. 1 The increasing relevance of proteolytic processes localized at the cell surface has attracted notable attention on membraneassociated proteolytic systems, including the family of type II transmembrane serine proteases (TTSPs). 2,3 The TTSP family is composed of more than 20 different members that share a number of structural features: a single-pass transmembrane domain located near the short cytoplasmic amino-terminal tail, a central region containing different protein-interacting domains, and a carboxyterminal catalytic region with the structural characteristics of serine proteases. The large variability of the central modular region together with the diverse expression patterns of TTSP family members suggest that these enzymes may play different physiologic and pathologic roles, although only a few of these functions have been identified so far. Thus, enteropeptidase is mainly expressed in the duodenum and plays an essential role in food digestion as activator of pancreatic trypsinogen to trypsin. 4 Hepsin, is mainly expressed in liver, but it is highly up-regulated in prostate cancer. 5,6 Matriptase/MT-SP1 is a widely studied member of the TTSP family because of its relevance in diverse processes, including cancer progression. 7,8 Mutant mice deficient in matriptase die shortly after birth because of aberrant skin ...

Matrix metalloproteinases (MMPs) have been implicated in numerous tissue-remodeling processes. The finding that mice deficient in collagenase-2 (MMP-8) are more susceptible to develop skin cancer, prompted us to investigate the role of this protease in cutaneous wound healing. We have observed a significant delay in wound closure in MMP8-/- mice and an altered inflammatory response in their wounds, with a delay of neutrophil infiltration during the first days and a persistent inflammation at later time points. These changes were accompanied by alterations in the TGF-beta1 signaling pathway and by an apoptosis defect in MMP8-/- mice. The delay in wound healing observed in MMP8-/- mice was rescued by bone marrow transplantation from wild-type mice. Analysis of other MMPs showed that MMP8-/- mice had a significant increase in the expression of MMP-9, suggesting that both proteases might act coordinately in this process. This possibility was further supported by the novel finding that MMP-8 and MMP-9 form specific complexes in vivo. Taken together, these data indicate that MMP-8 participates in wound repair by contributing to the resolution of inflammation and open the possibility to develop new strategies for treating wound healing defects.

Mutations in different genes encoding sarcomeric proteins are responsible for 50-60% of familial cases of hypertrophic cardiomyopathy (HCM); however, the genetic alterations causing the disease in one-third of patients are currently unknown. Here we describe a case with familial HCM of unknown cause. Whole-exome sequencing reveals a variant in the gene encoding the sarcomeric protein filamin C (p.A1539T) that segregates with the disease in this family. Sequencing of 92 HCM cases identifies seven additional variants segregating with the disease in eight families. Patients with FLNC mutations show marked sarcomeric abnormalities in cardiac muscle, and functional analysis reveals that expression of these FLNC variants resulted in the formation of large filamin C aggregates. Clinical studies indicate that FLNC-mutated patients have higher incidence of sudden cardiac death. On the basis of these findings, we conclude that mutations in the gene encoding the sarcomeric protein filamin C cause a new form of familial HMC.