The ectodomains of numerous proteins are released from cells by proteolysis to yield soluble intercellular regulators. The responsible protease, tumor necrosis factor-alpha converting enzyme (TACE), has been identified only in the case when tumor necrosis factor-alpha (TNFalpha) is released. Analyses of cells lacking this metalloproteinase-disintegrin revealed an expanded role for TACE in the processing of other cell surface proteins, including a TNF receptor, the L-selectin adhesion molecule, and transforming growth factor-alpha (TGFalpha). The phenotype of mice lacking TACE suggests an essential role for soluble TGFalpha in normal development and emphasizes the importance of protein ectodomain shedding in vivo.

Heparin‐binding epidermal growth factor (HB‐EGF) and betacellulin (BTC) are activating ligands for EGF receptor (EGFR/ErbB1) and ErbB4. To identify their physiological functions, we disrupted mouse HB‐EGF and BTC alleles by homologous recombination. Most HB‐EGF−/− mice died before weaning, and survivors had enlarged, dysfunctional hearts and reduced lifespans. Although BTC−/− mice were viable and fertile and displayed no overt defects, the lifespan of double null HB‐EGF−/−/BTC−/− mice was further reduced, apparently due to accelerated heart failure. HB‐EGF−/− newborns had enlarged and malformed semilunar and atrioventricular heart valves, and hypoplastic, poorly differentiated lungs. Defective cardiac valvulogenesis was the result of abnormal mesenchymal cell proliferation during remodeling, and was associated with dramatic increases in activated Smad1/5/8. Consistent with the phenotype, HB‐EGF transcripts were localized to endocardial cells lining the margins of wild‐type valves. Similarly defective valvulogenesis was observed in newborn mice lacking EGFR and tumor necrosis factor‐α converting enzyme (TACE). These results suggest that cardiac valvulogenesis is dependent on EGFR activation by TACE‐derived soluble HB‐EGF, and that EGFR signaling is required to regulate bone morphogenetic protein signaling in this context.

We previously implicated tumor necrosis factor-␣ converting enzyme (TACE/ADAM17) in the processing of the integral membrane precursor to soluble transforming growth factor-␣ (TGF-␣), pro-TGF-␣. Here we examined TGF-␣ processing in a physiologically relevant cell model, primary keratinocytes, showing that cells lacking TACE activity shed dramatically less TGF-␣ as compared with wild-type cultures and that TGF-␣ cleavage was partially restored by infection of TACE-deficient cells with TACE-encoding adenovirus. Moreover, cotransfection of TACE-deficient fibroblasts with pro-TGF-␣ and TACE cDNAs increased shedding of mature TGF-␣ with concomitant conversion of cell-associated pro-TGF-␣ to a processed form. Purified TACE accurately cleaved pro-TGF-␣ in vitro at the N-terminal site and also cleaved a soluble form of pro-TGF-␣ containing only the ectodomain at the C-terminal site. In vitro, TACE accurately cleaved peptides corresponding to cleavage sites of several epidermal growth factor (EGF) family members, and transfection of TACE into TACEdeficient cells increased the shedding of amphiregulin and heparin-binding EGF (HB-EGF) proteins. Consistent with the hypothesis that TACE regulates EGF receptor (EGFR) ligand availability in vivo, mice heterozygous for Tace and homozygous for an impaired EGFR allele (wa-2) were born with open eyes significantly more often than Tace ؉/؉ Egfr wa-2/wa-2 counterparts. Collectively, these data support a broad role for TACE in the regulated shedding of EGFR ligands.

Mechanisms of progression of chronic renal diseases, a major healthcare burden, are poorly understood. Angiotensin II (AngII), the major renin-angiotensin system effector, is known to be involved in renal deterioration, but the molecular pathways are still unknown. Here, we show that mice overexpressing a dominant negative isoform of epidermal growth factor receptor (EGFR) were protected from renal lesions during chronic AngII infusion. Transforming growth factor-alpha (TGF-alpha) and its sheddase, TACE (also known as ADAM17), were induced by AngII treatment, TACE was redistributed to apical membranes and EGFR was phosphorylated. AngII-induced lesions were substantially reduced in mice lacking TGF-alpha or in mice given a specific TACE inhibitor. Pharmacologic inhibition of AngII prevented TGF-alpha and TACE accumulation as well as renal lesions after nephron reduction. These findings indicate a crucial role for AngII-dependent EGFR transactivation in renal deterioration and identify in TACE inhibitors a new therapeutic strategy for preventing progression of chronic renal diseases.

Development 132, 3923-3933.On p. 3924 of this article in the section 'Mammary organoid culture', the concentrations of three constituents in the basal medium are incorrect. The correct concentrations are 10 g/ml insulin, 5.5 g/ml transferrin and 5 ng/ml sodium selenite.In addition, the authors also wish to acknowledge Jimmie E. Fata for his suggestions regarding this assay.The authors apologise to readers for these mistakes.

Rapidly progressive glomerulonephritis (RPGN) is a clinical a morphological expression of severe glomerular injury. Glomerular injury manifests as a proliferative histological pattern (“crescents”) with accumulation of T cells and macrophages, and proliferation of intrinsic glomerular cells. We show de novo induction of heparin-binding epidermal growth factor-like growth factor (HB-EGF) in intrinsic glomerular epithelial cells (podocytes) from both mice and humans with RPGN. HB-EGF induction increases phosphorylation of the EGFR/ErbB1 receptor in mice with RPGN. In HB-EGF-deficient mice, EGFR activation in glomeruli is absent and the course of RPGN is improved. Autocrine HB-EGF induces a phenotypic switch in podocytes in vitro. Conditional deletion of the Egfr gene from podocytes of mice alleviates the severity of RPGN. Pharmacological blockade of EGFR also improves the course of RPGN, even when started 4 days after the induction of experimental RPGN. This suggests that targeting the HB-EGF/EGFR pathway could also be beneficial for treatment of human RPGN.

Collagen XVII, a type II transmembrane protein and epithelial adhesion molecule, can be proteolytically shed from the cell surface to generate a soluble collagen. Here we investigated the release of the ectodomain and identi®ed the enzymes involved. After surface biotinylation of keratinocytes, the ectodomain was detectable in the medium within minutes and remained stable for >48 h. Shedding was enhanced by phorbol esters and inhibited by metalloprotease inhibitors, including hydroxamates and TIMP-3, but not by inhibitors of other protease classes or by TIMP-2. This pro®le implicated MMPs or ADAMs as candidate sheddases. MMP-2, MMP-9 and MT1-MMP were excluded, but TACE, ADAM-10 and ADAM-9 were shown to be expressed in keratinocytes and to be actively involved. Transfection with cDNAs for the three ADAMs resulted in increased shedding and, vice versa, in TACE-de®cient cells shedding was signi®cantly reduced, indicating that transmembrane collagen XVII represents a novel class of substrates for ADAMs. Functionally, release of the ectodomain of collagen XVII from the cell surface was associated with altered keratinocyte motility in vitro.

Activation of inflammatory pathways may contribute to the beginning and the progression of both atherosclerosis and type 2 diabetes. Here we report a novel interaction between insulin action and control of inflammation, resulting in glucose intolerance and vascular inflammation and amenable to therapeutic modulation. In insulin receptor heterozygous (Insr +/-) mice, we identified the deficiency of tissue inhibitor of metalloproteinase 3 (Timp3, an inhibitor of both TNF-α-converting enzyme [TACE] and MMPs) as a common bond between glucose intolerance and vascular inflammation. Among Insr +/-mice, those that develop diabetes have reduced Timp3 and increased TACE activity. Unchecked TACE activity causes an increase in levels of soluble TNF-α, which subsequently promotes diabetes and vascular inflammation. Double heterozygous Insr +/-Timp3 +/-mice develop mild hyperglycemia and hyperinsulinemia at 3 months and overt glucose intolerance and hyperinsulinemia at 6 months. A therapeutic role for Timp3/TACE modulation is supported by the observation that pharmacological inhibition of TACE led to marked reduction of hyperglycemia and vascular inflammation in Insr +/-diabetic mice, as well as by the observation of increased insulin sensitivity in Tace +/-mice compared with WT mice. Our results suggest that an interplay between reduced insulin action and unchecked TACE activity promotes diabetes and vascular inflammation.