Biomedical subjects

Karin List

Publications and source records attributed to Karin List.

10 recordsLinked to original sources

PubMed2006

Evidence for a matriptase-prostasin proteolytic cascade regulating terminal epidermal differentiation.

Recent gene ablation studies in mice have shown that matriptase, a type II transmembrane serine protease, and prostasin, a glycosylphosphatidylinositol-anchored membrane serine protease, are both required for processing of the epidermis-specific polyprotein, profilaggrin, stratum corneum formation, and acquisition of epidermal barrier function. Here we present evidence that matriptase acts upstream of prostasin in a zymogen activation cascade that regulates terminal epidermal differentiation and is required for prostasin zymogen activation. Enzymatic gene trapping of matriptase combined with prostasin immunohistochemistry revealed that matriptase was co-localized with prostasin in transitional layer cells of the epidermis and that the developmental onset of expression of the two membrane proteases was coordinated and correlated with acquisition of epidermal barrier function. Purified soluble matriptase efficiently converted soluble prostasin zymogen to an active two-chain form that formed SDS-stable complexes with the serpin protease nexin-1. Whereas two forms of prostasin with molecular weights corresponding to the prostasin zymogen and active prostasin were present in wild type epidermis, prostasin was exclusively found in the zymogen form in matriptase-deficient epidermis. These data suggest that matriptase, an autoactivating protease, acts upstream from prostasin to initiate a zymogen cascade that is essential for epidermal differentiation.

Animals↗

PubMed2006

Delineation of matriptase protein expression by enzymatic gene trapping suggests diverging roles in barrier function, hair formation, and squamous cell carcinogenesis.

The membrane serine protease matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma.

Animals↗

PubMed2005

Deregulated matriptase causes ras-independent multistage carcinogenesis and promotes ras-mediated malignant transformation.

Overexpression of the type II transmembrane serine protease matriptase is a highly consistent feature of human epithelial tumors. Here we show that matriptase possesses a strong oncogenic potential when unopposed by its endogenous inhibitor, HAI-1. Modest orthotopic overexpression of matriptase in the skin of transgenic mice caused spontaneous squamous cell carcinoma and dramatically potentiated carcinogen-induced tumor formation. Matriptase-induced malignant conversion was preceded by progressive interfollicular hyperplasia, dysplasia, follicular transdifferentiation, fibrosis, and dermal inflammation. Furthermore, matriptase induced activation of the pro-tumorigenic PI3K-Akt signaling pathway. This activation was frequently accompanied by H-ras or K-ras mutations in carcinogen-induced tumors, whereas matriptase-induced spontaneous carcinoma formation occurred independently of ras activation. Increasing epidermal HAI-1 expression completely negated the oncogenic effects of matriptase. The data implicate dysregulated matriptase expression in malignant epithelial transformation.

9,10-Dimethyl-1,2-benzanthracene↗

PubMed2004

Specific immunoassays for detection of intact and cleaved forms of the urokinase receptor.

BACKGROUND: The cell surface receptor (uPAR) for urokinase plasminogen activator (uPA) is a strong prognostic marker in several types of cancer. uPA cleaves the three-domain protein uPAR(I-III) into two fragments: uPAR(I), which contains domain I; and uPAR(II-III), which contains domains II and III. Established immunoassays measure a combination of uPAR forms. Our aim was to design immunoassays for specific quantification of the individual forms of uPAR. METHODS: Using appropriate combinations of epitope-mapped monoclonal antibodies (Mabs) for capture and europium-labeled detection Mabs, we designed two-site sandwich time-resolved fluorescence immunoassays (TR-FIAs): TR-FIA 1 to measure uPAR(I-III) alone; TR-FIA 2 to measure both uPAR(I-III) and uPAR(II-III); and TR-FIA 3 to measure uPAR(I). To avoid detection of uPAR(I-III) in TR-FIA 3, we used a combination of the peptide uPAR antagonist AE120 and a domain I antibody, R3. AE120 blocks the binding of R3 to uPAR(I-III). In contrast, AE120 does not interact with liberated domain I and therefore does not interfere with the binding of R3 to uPAR(I). RESULTS: The limits of quantification (CV <20%) determined by adding the proteins to uPAR-depleted plasma were <3 pmol/L in all three assays. The interassay CVs in plasma with added analytes were <11%, and recoveries were between 93% and 105%. Cross-reactivities of purified proteins in the three TR-FIAs were no more than 4%. Studies on chymotrypsin cleavage of uPAR and size-exclusion chromatography of plasma with and without added protein further supported the specificity of the assays. CONCLUSIONS: The three novel TR-FIAs accurately quantify uPAR(I-III) alone, uPAR(I-III) together with uPAR(II-III), and uPAR(I), respectively, in biological samples, including plasma, and thus are well suited for studies of the diagnostic and prognostic value of individual uPAR forms in cancer patients.

Biomarkers, Tumor↗

PubMed2003

Loss of proteolytically processed filaggrin caused by epidermal deletion of Matriptase/MT-SP1.

Profilaggrin is a large epidermal polyprotein that is proteolytically processed during keratinocyte differentiation to release multiple filaggrin monomer units as well as a calcium-binding regulatory NH2-terminal filaggrin S-100 protein. We show that epidermal deficiency of the transmembrane serine protease Matriptase/MT-SP1 perturbs lipid matrix formation, cornified envelope morphogenesis, and stratum corneum desquamation. Surprisingly, proteomic analysis of Matriptase/MT-SP1-deficient epidermis revealed the selective loss of both proteolytically processed filaggrin monomer units and the NH2-terminal filaggrin S-100 regulatory protein. This was associated with a profound accumulation of profilaggrin and aberrant profilaggrin-processing products in the stratum corneum. The data identify keratinocyte Matriptase/MT-SP1 as an essential component of the profilaggrin-processing pathway and a key regulator of terminal epidermal differentiation.

Animals↗

PubMed2003

The low density lipoprotein receptor-related protein modulates protease activity in the brain by mediating the cellular internalization of both neuroserpin and neuroserpin-tissue-type plasminogen activator complexes.

Proteases contribute to a variety of processes in the brain; consequently, their activity is carefully regulated by protease inhibitors, such as neuroserpin. This inhibitor is thought to be secreted by axons at synaptic regions where it controls tissue-type plasminogen activator (tPA) activity. Mechanisms regulating neuroserpin are not known, and the current studies were undertaken to define the cellular pathways involved in neuroserpin catabolism. We found that both active neuroserpin and neuroserpin.tPA complexes were internalized by mouse cortical cultures and embryonic fibroblasts in a process mediated by the low density lipoprotein receptor-related protein (LRP). Surprisingly, despite the fact that active neuroserpin is internalized by LRP, this form of the molecule does not directly bind to LRP on its own, indicating the requirement of a cofactor for neuroserpin internalization. Our studies ruled out the possibility that endogenously produced plasminogen activators (i.e. tPA and urokinase-type plasminogen activator) are responsible for the LRP-mediated internalization of active neuroserpin, but could not rule out the possibility that another cell-associated proteases capable of binding active neuroserpin functions in this capacity. In summary, neuroserpin levels appear to be carefully regulated by LRP and an unidentified cofactor, and this pathway may be critical for maintaining the balance between proteases and inhibitors.

Animals↗

PubMed2003

The pro-urokinase plasminogen-activation system in the presence of serpin-type inhibitors and the urokinase receptor: rescue of activity through reciprocal pro-enzyme activation.

The reciprocal pro-enzyme activation system of plasmin, urokinase-type plasminogen activator (uPA) and their respective zymogens is a potent mechanism in the generation of extracellular proteolytic activity. Plasminogen activator inhibitor type 1 (PAI-1) acts as a negative regulator. This system is complicated by a poorly understood intrinsic reactivity of the uPA pro-enzyme (pro-uPA) before proteolytic activation, directed against both plasminogen and PAI-1. We have studied the integrated activation mechanism under the repression of PAI-1 in a purified system. A covalent reaction between pro-uPA and PAI-1 was positively demonstrated but the reaction of PAI-1 with two-chain uPA was found to be at least 1000-fold faster. However, in spite of this very fast inhibition, two-chain uPA still became the dominant plasminogen activator when plasminogen was incubated with pro-uPA and PAI-1. The activity pattern observed under these conditions revealed an initial lag phase, followed by a continuous generation of minute amounts of active two-chain uPA, this uPA having a short lifetime before inhibition but still succeeding to generate new plasmin activity, thus preventing a complete inactivation of the feedback system. This property of the activation system was retained even in the simultaneous presence of PAI-1 and alpha(2)-antiplasmin. Addition of soluble uPA receptor to the system did not change the role of pro-uPA and the same pattern was observed when pro-uPA was bound to the uPA receptor on U937 cells. The present mechanism maintains the system at standby level and may be triggered to increased activity without the need for an external initiating event.

Enzyme Activation↗

PubMed2003

uPARAP/Endo180 is essential for cellular uptake of collagen and promotes fibroblast collagen adhesion.

The uptake and lysosomal degradation of collagen by fibroblasts constitute a major pathway in the turnover of connective tissue. However, the molecular mechanisms governing this pathway are poorly understood. Here, we show that the urokinase plasminogen activator receptor-associated protein (uPARAP)/Endo180, a novel mesenchymally expressed member of the macrophage mannose receptor family of endocytic receptors, is a key player in this process. Fibroblasts from mice with a targeted deletion in the uPARAP/Endo180 gene displayed a near to complete abrogation of collagen endocytosis. Furthermore, these cells had diminished initial adhesion to a range of different collagens, as well as impaired migration on fibrillar collagen. These studies identify a central function of uPARAP/Endo180 in cellular collagen interactions.

Animals↗

PubMed2002

Matriptase/MT-SP1 is required for postnatal survival, epidermal barrier function, hair follicle development, and thymic homeostasis.

Matriptase/MT-SP1 is a novel tumor-associated type II transmembrane serine protease that is highly expressed in the epidermis, thymic stroma, and other epithelia. A null mutation was introduced into the Matriptase/MT-SP1 gene of mice to determine the role of Matriptase/MT-SP1 in epidermal development and neoplasia. Matriptase/MT-SP1-deficient mice developed to term but uniformly died within 48 h of birth. All epidermal surfaces of newborn mice were grossly abnormal with a dry, red, shiny, and wrinkled appearance. Matriptase/MT-SP1-deficiency caused striking malformations of the stratum corneum, characterized by dysmorphic and pleomorphic corneocytes and the absence of vesicular bodies in transitional layer cells. This aberrant skin development seriously compromised both inward and outward epidermal barrier function, leading to the rapid and fatal dehydration of Matriptase/MT-SP1-deficient pups. Loss of Matriptase/MT-SP1 also seriously affected hair follicle development resulting in generalized follicular hypoplasia, absence of erupted vibrissae, lack of vibrissal hair canal formation, ingrown vibrissae, and wholesale abortion of vibrissal follicles. Furthermore, Matriptase/MT-SP1-deficiency resulted in dramatically increased thymocyte apoptosis, and depletion of thymocytes. This study demonstrates that Matriptase/MT-SP1 has pleiotropic functions in the development of the epidermis, hair follicles, and cellular immune system.

Amino Acid Sequence↗

PubMedDate not supplied

Matriptase: potent proteolysis on the cell surface.

Matriptase is a type II transmembrane serine protease expressed in most human epithelia, where it is coexpressed with its cognate transmembrane inhibitor, hepatocyte growth factor activator inhibitor (HAI)-1. Activation of the matriptase zymogen requires sequential N-terminal cleavage, activation site autocleavage, and transient association with HAI-1. Matriptase has an essential physiological role in profilaggrin processing, corneocyte maturation, and lipid matrix formation associated with terminal differentiation of the oral epithelium and the epidermis, and is also critical for hair follicle growth. Matriptase and HAI expression are frequently dysregulated in human cancer, and matriptase expression that is unopposed by HAI-1 potently promotes carcinogenesis and metastatic dissemination in animal models.

Animals↗