Cell Surface Proteolysis Research Articles

Proteomics-based discovery of a novel, structurally unique, and developmentally regulated plasminogen receptor, Plg-RKT, a major regulator of cell surface plasminogen activation

AbstractActivation of plasminogen, the zymogen of the primary thrombolytic enzyme, plasmin, is markedly promoted when plasminogen is bound to cell surfaces, arming cells with the broad spectrum proteolytic activity of plasmin. In addition to its role in thrombolysis, cell surface plasmin facilitates a wide array of physiologic and pathologic processes. Carboxypeptidase B-sensitive plasminogen binding sites promote plasminogen activation on eukaryotic cells. However, no integral membrane plasminogen receptors exposing carboxyl terminal basic residues on cell surfaces have been identified. Here we use the exquisite sensitivity of multidimensional protein identification technology and an inducible progenitor cell line to identify a novel differentiation-induced integral membrane plasminogen receptor that exposes a C-terminal lysine on the cell surface, Plg-RKT (C9orf46 homolog). Plg-RKT was highly colocalized on the cell surface with the urokinase receptor, uPAR. Our data suggest that Plg-RKT also interacts directly with tissue plasminogen activator. Furthermore, Plg-RKT markedly promoted cell surface plasminogen activation. Database searching revealed that Plg-RKT mRNA is broadly expressed by migratory cell types, including leukocytes, and breast cancer, leukemic, and neuronal cells. This structurally unique plasminogen receptor represents a novel control point for regulating cell surface proteolysis.

Cell-surface Processing of the Metalloprotease Pro-ADAMTS9 Is Influenced by the Chaperone GRP94/gp96

A disintegrin-like and metalloprotease domain with thrombospondin type 1 motifs 9 (ADAMTS9) is a highly conserved metalloprotease that has been identified as a tumor suppressor gene and is required for normal mouse development. The secreted ADAMTS9 zymogen undergoes proteolytic excision of its N-terminal propeptide by the proprotein convertase furin. However, in contrast to other metalloproteases, propeptide excision occurs at the cell surface and leads to decreased activity of the zymogen. Here, we investigated the potential cellular mechanisms regulating ADAMTS9 biosynthesis and cell-surface processing by analysis of molecular complexes formed by a construct containing the propeptide and catalytic domain of pro-ADAMTS9 (Pro-Cat) in HEK293F cells. Cross-linking of cellular proteins bound to Pro-Cat followed by mass spectrometric analysis identified UDP-glucose:glycoprotein glucosyltransferase I, heat shock protein gp96 (GRP94), BiP (GRP78), and ERdj3 (Hsp40 homolog) as associated proteins. gp96 and BiP were present at the cell surface in an immunoprecipitable complex with pro-ADAMTS9 and furin. Treatment with geldanamycin, an inhibitor of the HSP90alpha family (including gp96), led to decreased furin processing of pro-ADAMTS9 and accumulation of the unprocessed pro-ADAMTS9 at the cell surface. gp96 siRNA down-regulated the levels of cell-surface pro-ADAMTS9 and furin, whereas the levels of cell-surface pro-ADAMTS9, but not of cell-surface furin, were decreased upon treatment with BiP siRNA. These data identify for the first time the cellular chaperones associated with secretion of an ADAMTS protease and suggest a role for gp96 in modulating pro-ADAMTS9 processing.

A Mediator of Rho-dependent Invasion Moonlights as a Methionine Salvage Enzyme

RhoA controls changes in cell morphology and invasion associated with cancer phenotypes. Cell lines derived from melanoma tumors at varying stages revealed that RhoA is selectively activated in cells of metastatic origin. We describe a functional proteomics strategy to identify proteins regulated by RhoA and report a previously uncharacterized human protein, named “mediator of RhoA-dependent invasion (MRDI),” that is induced in metastatic cells by constitutive RhoA activation and promotes cell invasion. In human melanomas, MRDI localization correlated with stage, showing nuclear localization in nevi and early stage tumors and cytoplasmic localization with plasma membrane accentuation in late stage tumors. Consistent with its role in promoting cell invasion, MRDI localized to cell protrusions and leading edge membranes in cultured cells and was required for cell motility, tyrosine phosphorylation of focal adhesion kinase, and modulation of actin stress fibers. Unexpectedly MRDI had enzymatic function as an isomerase that converts the S-adenosylmethionine catabolite 5-methylribose 1-phosphate into 5-methylribulose 1-phosphate. The enzymatic function of MRDI was required for methionine salvage from S-adenosylmethionine but distinct from its function in cell invasion. Thus, mechanisms used by signal transduction pathways to control cell movement have evolved from proteins with ancient function in amino acid metabolism.

Regulation of cell surface protease matriptase by HAI2 is essential for placental development, neural tube closure and embryonic survival in mice

Hypomorphic mutations in the human SPINT2 gene cause a broad spectrum of abnormalities in organogenesis, including organ and digit duplications, atresia, fistulas, hypertelorism, cleft palate and hamartoma. SPINT2 encodes the transmembrane serine protease inhibitor HAI2 (placental bikunin), and the severe developmental effects of decreased HAI2 activity can be hypothesized to be a consequence of excess pericellular proteolytic activity. Indeed, we show here that HAI2 is a potent regulator of protease-guided cellular responses, including motogenic activity and transepithelial resistance of epithelial monolayers. Furthermore, we show that inhibition of the transmembrane serine protease matriptase (encoded by St14) is an essential function of HAI2 during tissue morphogenesis. Genetic inactivation of the mouse Spint2 gene led to defects in neural tube closure, abnormal placental labyrinth development associated with loss of epithelial cell polarity, and embryonic demise. Developmental defects observed in HAI2-deficient mice were caused by unregulated matriptase activity, as both placental development and embryonic survival in HAI2-deficient embryos were completely restored by the simultaneous genetic inactivation of matriptase. However, neural tube defects were detected in HAI2-deficient mice even in the absence of matriptase, although at lower frequency, indicating that the inhibition of additional serine protease(s) by HAI2 is required to complete neural development. Finally, by genetic complementation analysis, we uncovered a unique and complex functional interaction between HAI2 and the related HAI1 in the regulation of matriptase activity during development. This study indicates that unregulated matriptase-dependent cell surface proteolysis can cause a diverse array of abnormalities in mammalian development.

The Urokinase Plasminogen Activator Receptor Promotes Efferocytosis of Apoptotic Cells

The urokinase receptor (uPAR), expressed on the surface of many cell types, coordinates plasmin-mediated cell surface proteolysis for matrix remodeling and promotes cell adhesion by acting as a binding protein for vitronectin. There is great clinical interest in uPAR in the cancer field as numerous reports have demonstrated that up-regulation of the uPA system is correlated with malignancy of various carcinomas. Using both stable cell lines overexpressing uPAR and transient gene transfer, here we provide evidence for a non-reported role of uPAR in the phagocytosis of apoptotic cells, a process that has recently been termed efferocytosis. When uPAR was expressed in human embryonic kidney cells, hamster melanoma cells, or breast cancer cells (BCCs), there was a robust enhancement in the efferocytosis of apoptotic cells. uPAR-expressing cells failed to stimulate engulfment of viable cells, suggesting that uPAR enhances recognition of one or more determinant on the surface of the apoptotic cell. uPAR-mediated engulfment was not inhibited by expression of mutant beta5 integrin, nor was alphavbeta5 integrin-mediated engulfment modulated by cleavage of uPAR by phosphatidylinositol-specific phospholipase C. Further, we found that the more aggressive BCCs had a higher phagocytic capacity that correlated with uPAR expression and cleavage of membrane-associated uPAR in MDA-MB231 BCCs significantly impaired phagocytic activity. Because efferocytosis is critical for the resolution of inflammation and production of anti-inflammatory cytokines, overexpression of uPAR in tumor cells may promote a tolerogenic microenvironment that favors tumor progression.

QS284. Secondary Bile Acids Activate the Epidermal Growth Factor Receptor (EGFR) Through a Src Kinase-Mediated Release of EGFR Ligands in Colon Cancer Cells

Introduction: Western diets high in fats and low in fiber are associated with increases in secondary bile acids, such as deoxycholic acid (DCA) which are known to promote colon cancer. We have shown that DCA-induced signaling is mediated by TNF-α converting enzyme (TACE)-dependent cleavage of the EGFR ligands amphiregulin (AR) and transforming growth factor-α (TGF-α) and subsequent activation of the EGFR. Communication between Src kinases and EGFR signaling has been shown to involve cell surface proteolysis of EGFR ligand precursors.

Mechanisms controlling human endothelial lumen formation and tube assembly in three‐dimensional extracellular matrices

Recent data have revealed new mechanisms that underlie endothelial cell (EC) lumen formation during vascular morphogenic events in development, wound repair, and other disease states. It is apparent that EC interactions with extracellular matrices (ECMs) establish signaling cascades downstream of integrin ligation leading to activation of the Rho GTPases, Cdc42 and Rac1, which are required for lumen formation. In large part, this process is driven by intracellular vacuole formation and coalescence, which rapidly leads to the creation of fluid-filled matrix-free spaces that are then interconnected via EC-EC interactions to create multicellular tube structures. EC vacuoles markedly accumulate in a polarized fashion directly adjacent to the centrosome in a region that strongly accumulates Cdc42 protein as indicated by green fluorescent protein (GFP)-Cdc42 during the lumen formation process. Downstream of Cdc42-mediated signaling, key molecules that have been identified to be required for EC lumen formation include Pak2, Pak4, Par3, Par6, and the protein kinase C (PKC) isoforms zeta and epsilon. Together, these molecules coordinately regulate the critical EC lumen formation process in three-dimensional (3D) collagen matrices. These events also require cell surface proteolysis mediated through membrane type 1 matrix metalloproteinase (MT1-MMP), which is necessary to create vascular guidance tunnels within the 3D matrix environment. These tunnels represent physical spaces within the ECM that are necessary to regulate vascular morphogenic events, including the establishment of interconnected vascular tube networks as well as the recruitment of pericytes to initiate vascular tube maturation (via basement membrane matrix assembly) and stabilization. Current research continues to analyze how specific molecules integrate signaling information in concert to catalyze EC lumen formation, pericyte recruitment, and stabilization processes to control vascular morphogenesis in 3D extracellular matrices.

Clinical relevance of hepsin and hepatocyte growth factor activator inhibitor type 2 expression in renal cell carcinoma

Cell surface proteolysis is important for the generation of bioactive proteins mediating tumor progression. Recent studies suggest that the membrane-anchored cell surface proteinases matriptase and hepsin have significant roles in tumors. We analyzed the expression and clinical relevance of matriptase and hepsin, and their inhibitors hepatocyte growth factor activator inhibitor type 1 (HAI-1) and type 2 (HAI-2) in 66 cases of conventional renal cell carcinomas (RCC). The mRNA level was evaluated in paired samples from tumor and non-tumorous renal tissues by real-time reverse transcription-polymerase chain reaction. As matriptase and hepsin potently activate the proform of hepatocyte growth factor (HGF), the expression of HGF and its receptor, c-Met, was also analyzed. Although upregulation of matriptase was observed occasionally in RCC, the expression level was not associated with prognostic parameters. Hepsin was downregulated in RCC, particularly in early stage disease, but upregulated in advanced stages. There was a trend of higher hepsin expression in RCC with distant metastasis, and Kaplan-Meier survival curves showed that high hepsin expression was associated with reduced overall survival (P<0.01, log-rank test). Moreover, multivariate analysis indicated that hepsin was an independent prognostic factor. Overexpression of HGF or c-Met also showed reduced overall survival. We also observed a tendency of low HAI-2 expression with reduced overall survival and a statistical association between high hepsin and low HAI-2 level. No associations were observed between matriptase and HAI-1 and HAI-2. Our findings suggest that the balance between hepsin and its inhibitor, HAI-2, may have prognostic value in RCC.

Matriptase-dependent cell surface proteolysis in epithelial development and pathogenesis

Matriptase is an epithelial type II transmembrane serine protease with a complex modular structure and sophisticated activation mechanism. Reduced matriptase activity in mice or humans is associated with incomplete terminal differentiation of epidermis, epidermal appendages, oral epithelium, and, likely, other epithelial structures. Preliminary evidence indicates that matriptase is part of a serine protease zymogen activation cascade that regulates epithelial cell proliferation and fate. Matriptase activity must be tightly controlled in epithelial tissues by transcriptional and posttranslational mechanisms, as matriptase dysregulation can cause embryonic demise as well as malignant transformation.

Secondary bile acids activate the epidermal growth factor receptor (EGFR) through a SRC kinase-mediated release of EGFR ligands in polarizing colon cancer cells

Purpose: Western diets high in fats and low in fiber are associated with increases in secondary bile acids, such as deoxycholic acid (DCA) which are known to promote colon cancer. Increasing evidence suggests that DCA-induced signaling is mediated by activation of the EGFR. Communication between Src kinases and EGFR signaling has been shown to involve cell surface proteolysis of EGFR ligand precursors. We have previously shown that ligand-dependent activation of the EGFR induces nuclear targeting of cyclooxygenase-2, basolateral release of prostaglandins (PGs), and mitogenesis in a human polarizing colon cancer cell line, HCA-7 (PNAS 94: 567-662, 1997).

Plasminogen receptors: the sine qua non of cell surface plasminogen activation.

Localization of plasminogen and plasminogen activators on cell surfaces promotes plasminogen activation and serves to arm cells with the broad spectrum proteolytic activity of plasmin. Cell surface proteolysis by plasmin is an essential feature of physiological and pathological processes requiring extracellular matrix degradation for cell migration including macrophage recruitment during the inflammatory response, tissue remodeling, wound healing, tumor cell invasion and metastasis and skeletal myogenesis. Cell associated plasmin on platelets and endothelial cells is optimally localized for promotion of clot lysis. In more recently recognized functions that are likely to be independent of matrix degradation, cell surface-bound plasmin participates in prohormone processing as well as stimulation of intracellular signaling. This issue of Frontiers in Bioscience on Plasminogen Receptors encompasses chapters focusing on the kinetics of cell surface plasminogen activation and the regulation of plasminogen receptor activity as well as the contribution of plasminogen receptors to the physiological and pathophysiological processes of myogenesis, muscle regeneration and cancer. The molecular identity of plasminogen receptors is cell-type specific, with distinct molecular entities providing plasminogen receptor function on different cells. This issue includes chapters on the well studied plasminogen receptor functions.

TSH receptor signaling via cyclic AMP inhibits cell surface degradation and internalization of E-cadherin in pig thyroid epithelium.

Incorporation of E-cadherin into the adherens junction is a highly regulated process required to establish firm cell-cell adhesion in most epithelia. Less is known about the mechanisms that govern the clearance of E-cadherin from the cell surface in both normal and pathological states. In this study, we found that the steady-state removal of E-cadherin in primary cultured pig thyroid cell monolayers is slow and involves intracellular degradation. Experimental abrogation of adhesion by a Ca2+ switch induces rapid cell surface proteolysis of E-cadherin. At the same time, endocytosed intact E-cadherin and newly synthesized E-cadherin accumulate in intracellular compartments that largely escape further degradation. Acute stimulation with thyroid-stimulating hormone (TSH) or forskolin prevents all signs of accelerated E-cadherin turnover. The findings indicate that TSH receptor signaling via cyclic AMP stabilizes the assembly and retention of E-cadherin at the cell surface. This suggests a new mechanism by which TSH supports maintenance of thyroid follicular integrity.

Regulated Cell Surface Pro-EGF Ectodomain Shedding Is a Zinc Metalloprotease-dependent Process

Epidermal growth factor receptor (EGFR) ligands are synthesized as type I membrane protein precursors exposed at the cell surface. Shedding of the ectodomain of these proteins is the way cells regulate the equilibrium between cell-associated and diffusible forms of these growth factors. Whereas the regulated shedding of transforming growth factor-alpha, HB-EGF, and amphiregulin precursors have been clearly established, regulation of full-length pro-EGF shedding has not been clearly demonstrated. Here, using both wild-type and M2 mutant CHO-K1 as well as HeLa cell lines transiently transfected with epitope-tagged rat pro-EGF expression plasmid, we demonstrate that these cells synthesize EGF as a high molecular weight membrane-associated precursor glycoprotein expressed at the cell surface. All cell lines are able to release the entire ectodomain of pro-EGF in the extracellular medium following juxtamembrane cleavage of the precursor once it is present at the cell surface. More significantly we clearly established that CHO-M2 and HeLa cells only constitutively release low levels of pro-EGF. This shedding is a regulated phenomenon in wild-type CHO cells where it can be induced by different agents such as phorbol 12-myristate 13-acetate (PMA), pervanadate, and serum but not by calcium ionophores. Using specific inhibitors as well as protein kinase C (PKC) depletion, PMA stimulation was shown to be completely dependent on PKC activation whereas pervanadate and serum stimulation were not. Regulated ectodomain shedding involves the activity of a zinc metalloprotease as determined by inhibition with phenantrolin and TAPI-2 and by the results obtained with the CHO-M2 shedding defective mutant cell line. Comparison of the ability of CHO and HeLa cell lines to shed pro-EGF and pro-TNF-alpha upon stimulation greatly suggests that TACE (ADAM 17) may not be the ectoprotease involved in the secretion of pro-EGF ectodomain and that this protease, which remains to be identified, shows a restricted cellular expression pattern.

TACE cleavage of proamphiregulin regulates GPCR-induced proliferation and motility of cancer cells.

Communication between G protein-coupled receptor (GPCR) and epidermal growth factor receptor (EGFR) signalling systems involves cell surface proteolysis of EGF-like precursors. The underlying mechanisms of EGFR signal transactivation pathways, however, are largely unknown. We demonstrate that in squamous cell carcinoma cells, stimulation with the GPCR agonists LPA or carbachol specifically results in metalloprotease cleavage and release of amphiregulin (AR). Moreover, AR gene silencing by siRNA or inhibition of AR biological activity by neutralizing antibodies and heparin prevents GPCR-induced EGFR tyrosine phosphorylation, downstream mitogenic signalling events, cell proliferation, migration and activation of the survival mediator Akt/PKB. Therefore, despite some functional redundancy among EGF family ligands, the present study reveals a distinct and essential role for AR in GPCR-triggered cellular responses. Furthermore, we present evidence that blockade of the metalloprotease-disintegrin tumour necrosis factor-alpha-converting enzyme (TACE) by the tissue inhibitor of metalloprotease-3, a dominant-negative TACE mutant or RNA interference suppresses GPCR-stimulated AR release, EGFR activation and downstream events. Thus, TACE can function as an effector of GPCR-mediated signalling and represents a key element of the cellular receptor cross-talk network.

Isolation and Isotope Labeling of Cysteine- and Methionine-containing Tryptic Peptides: Application to the Study of Cell Surface Proteolysis

Inexpensive methods were developed for isolating and isotopically labeling tryptic peptides that contain either cysteine or methionine. After covalently capturing cysteine-containing peptides with pyridyl disulfide reactive groups on agarose beads, extensive wash steps were applied, and the attached peptides were released using a reducing agent. This approach results in less nonspecifically bound peptides and eliminates the possibility of generating avidin peptide background ions that can arise when using methods based on biotin and avidin (e.g. isotope-coded affinity tag). The thiols were alkylated using either N-ethyl- or N-D5-ethyl-iodoacetamide, both of which can be synthesized in a single step using inexpensive reagents. This isotopic labeling does not greatly increase the peptide mass, nor does it affect the peptide ion charge state in electrospray ionization. In addition, methionine-containing peptides were captured using commercially available methionine-reactive beads, and relative quantitation of peptides was achieved by isotopic labeling of amino groups using activated esters of either nicotinic acid or D4-nicotinic acid. These methods were used to study the metalloprotease-mediated shedding of cell surface proteins from a mouse monocyte cell line that had been treated with a phorbol ester and lipopolysaccharide. In addition to the identification of proteins previously determined to be inducibly shed, three new shed proteins were identified: CD18, ICOS ligand, and tumor endothelial marker 7-related protein.

The evolving roles of cell surface proteases in health and disease: Implications for developmental, adaptive, inflammatory, and neoplastic processes

Publisher Summary Cell surface proteolysis is an important process, crucial in a growing number of biological functions including development, normal homeostasis and adaptive responses, degenerative diseases, inflammation and repair, and several aspects of the neoplastic process. In addition to a growing appreciation of the general importance of cell surface proteolysis there has also been a growing appreciation of the diversity of enzymes and enzyme systems which are involved in this process. This chapter illustrates cell-surface proteolysis in each of these areas. Protease activities are crucial during development. In addition to extracellular matrix remodeling, cell surface proteases are essential in modulating cell proliferation, migration, and differentiation as well as in signaling cell attraction and repulsion. Maintenance of homeostasis entails a continuous, dynamic response to the environment that sometimes may include controlled proteolytic responses crucial for modulation of local growth factor levels, optimal functioning of the clotting and complement systems, and remodeling of our connective tissues and vascular system.

Acquisition of regulators of complement activation by Streptococcus pyogenes serotype M1.

Opsonization of bacteria by complement proteins is an important component of the immune response. The pathogenic bacterium Streptococcus pyogenes has evolved multiple mechanisms for the evasion of complement-mediated opsonization. One mechanism involves the binding of human regulators of complement activation such as factor H (FH) and FH-like protein 1 (FHL-1). Acquisition of these regulatory proteins can limit deposition of the opsonin C3b on bacteria, thus decreasing the pathogen's susceptibility to phagocytosis. Binding of complement regulatory proteins by S. pyogenes has previously been attributed to the streptococcal M and M-like proteins. Here, we report that the S. pyogenes cell surface protein Fba can mediate binding of FH and FHL-1. We constructed mutant derivatives of S. pyogenes that lack Fba, M1 protein, or both proteins and assayed the strains for FH binding, susceptibility to phagocytosis, and C3 deposition. Fba expression was found to be sufficient for binding of purified FH as well as for binding of FH and FHL-1 from human plasma. Plasma adsorption experiments also revealed that M1(+) Fba(+) streptococci preferentially bind FHL-1, whereas M1(-) Fba(+) streptococci have similar affinities for FH and FHL-1. Fba was found to contribute to the survival of streptococci incubated with human blood and to inhibit C3 deposition on bacterial cells. Streptococci harvested from log-phase cultures readily bound FH, but binding was greatly reduced for bacteria obtained from stationary-phase cultures. Bacteria cultured in the presence of the protease inhibitor E64 maintained FH binding activity in stationary phase, suggesting that Fba is removed from the cell surface via proteolysis. Western analyses confirmed that E64 stabilizes cell surface expression of Fba. These data indicate that Fba is an antiopsonic, antiphagocytic protein that may be regulated by cell surface proteolysis.

Protease-activated receptors: the role of cell-surface proteolysis in signalling.

Certain extracellular proteases, derived from the circulation and inflammatory cells, can specifically cleave and trigger protease-activated receptors (PARs), a small, but important, sub-group of the G-protein-coupled receptor super-family. Four PARs have been cloned and they all share the same basic mechanism of activation: proteases cleave at a specific site within the extracellular N-terminus to expose a new N-terminal tethered ligand domain, which binds to and thereby activates the cleaved receptor. Thrombin activates PAR1, PAR3 and PAR4, trypsin activates PAR2 and PAR4, and mast cell tryptase activates PAR2 in this manner. Activated PARs couple to signalling cascades that affect cell shape, secretion, integrin activation, metabolic responses, transcriptional responses and cell motility. PARs are 'single-use' receptors: proteolytic activation is irreversible and the cleaved receptors are degraded in lysosomes. Thus, PARs play important roles in 'emergency situations', such as trauma and inflammation. The availability of selective agonists and antagonists of protease inhibitors and of genetic models has generated evidence to suggests that proteases and their receptors play important roles in coagulation, inflammation, pain, healing and protection. Therefore, selective antagonists or agonists of these receptors may be useful therapeutic agents for the treatment of human diseases.

Proteolytic Cleavage of the CD44 Adhesion Molecule in Multiple Human Tumors

Cell surface adhesion molecules are crucial for the development and/or pathogenesis of various diseases including cancer. CD44 has received much interest as a major adhesion molecule that is involved in tumor progression. We have previously demonstrated that the ectodomain of CD44 undergoes proteolytic cleavage by membrane-associated metalloproteases in various tumor cell lines. The remaining membrane-bound CD44 cleavage product can be detected using antibodies against the cytoplasmic domain of CD44 (anti-CD44cyto antibody). However, the cleavage of CD44 in primary human tumors has not been investigated. Using Western blots with anti-CD44cyto antibody to assay human tumor tissues, we show that the CD44 cleavage product can be detected in 58% (42 of 72) of gliomas but not in normal brain. Enhanced CD44 cleavage was also found in 67% (28 of 42) of breast carcinomas, 45% (5 of 11) of non-small cell lung carcinomas, 90% (9 of 10) of colon carcinomas, and 25% (3 of 12) of ovarian carcinomas. Tumors expressing a CD44 splice variant showed a significantly higher incidence of enhanced CD44 cleavage. The wide prevalence of CD44 cleavage suggests that it plays an important role in the pathogenesis of human tumors.

Shedding light on sheddases: role in growth and development

The extracellular domains of several integral membrane proteins are released from the cell surface by a group of enzymes known as "sheddases" through a process called "ectodomain shedding". Because many transmembrane growth and differentiation factors, including members of the epidermal growth factor (EGF) family that play a crucial role in development, require ectodomain shedding for proper action in vivo, proteolysis is now viewed as a regulatory mechanism in the developing embryos. Two recent reports by Zhao et al. provide evidence for the role of cell surface proteolysis by an ADAM (a disintegrin and metalloprotease) in the development of murine lung. Inhibition of tumor necrosis factor-alpha converting enzyme (TACE, ADAM17) by the hydroxamic acid-based metalloprotease inhibitor (TAPI), or a targeted mutation in Zn(2+)-binding domain of TACE, disrupts two essential epithelial functions in lung development: branching morphogenesis and cytodifferentiation. Evidence for the role of ADAMs as sheddases in development and growth factor signaling is discussed.