Phospholipid scramblase 1 is secreted by a lipid raft-dependent pathway and interacts with the extracellular matrix protein 1 in the dermal epidermal junction zone of human skin.

We examined the interaction of ECM1 (extracellular matrix protein 1) using yeast two-hybrid screening and identified the type II transmembrane protein, PLSCR1 (phospholipid scramblase 1), as a binding partner. This interaction was then confirmed by in vitro and in vivo co-immunoprecipitation experiments, and additional pull-down experiments with GST-tagged ECM1a fragments localized this interaction to occur within the tandem repeat region of ECM1a. Furthermore, immunohistochemical staining revealed a partial overlap of ECM1 and PLSCR1 in human skin at the basal epidermal cell layer. Moreover, in human skin equivalents, both proteins are expressed at the basal membrane in a dermal fibroblast-dependent manner. Next, immunogold electron microscopy of ultrathin human skin sections showed that ECM1 and PLSCR1 co-localize in the extracellular matrix, and using antibodies against ECM1 or PLSCR1 cross-linked to magnetic immunobeads, we were able to demonstrate PLSCR1-ECM1 interaction in human skin extracts. Furthermore, whereas ECM1 is secreted by the endoplasmic/Golgi-dependent pathway, PLSCR1 release from HaCaT keratinocytes occurs via a lipid raft-dependent mechanism, and is deposited in the extracellular matrix. In summary, we here demonstrate that PLSCR1 interacts with the tandem repeat region of ECM1a in the dermal epidermal junction zone of human skin and provide for the first time experimental evidence that PLSCR1 is secreted by an unconventional secretion pathway. These data suggest that PLSCR1 is a multifunctional protein that can function both inside and outside of the cell and together with ECM1 may play a regulatory role in human skin.

Epilepsy and migraine in a patient with Urbach-Wiethe disease.

We report the clinical, neuroradiological, and molecular genetic findings in a patient with lipoid proteinosis or Urbach-Wiethe disease. Interestingly, in this patient epilepsy and migraine were the symptoms leading to the diagnosis of the disease, contrary to most patients in whom skin abnormalities are the first recognized symptoms.

Interaction between cartilage oligomeric matrix protein and extracellular matrix protein 1 mediates endochondral bone growth.

In an effort to define the biological functions of COMP, a functional genetic screen was performed. This led to the identification of extracellular matrix protein 1 (ECM1) as a novel COMP-associated partner. COMP directly binds to ECM1 both in vitro and in vivo. The EGF domain of COMP and the C-terminus of ECM1 mediate the interaction between them. COMP and ECM1 colocalize in the growth plates invivo. ECM1 inhibits chondrocyte hypertrophy, matrix mineralization, and endochondral bone formation, and COMP overcomes the inhibition by ECM1. In addition, COMP-mediated neutralization of ECM1 inhibition depends on their interaction, since COMP largely fails to overcome the ECM1 inhibition in the presence of the EGF domain of COMP, which disturbs the association of COMP and ECM1. These findings provide the first evidence linking the association of COMP and ECM1 and the biological significance underlying the interaction between them in regulating endochondral bone growth.

Interaction of extracellular matrix protein 1 with extracellular matrix components: ECM1 is a basement membrane protein of the skin.

The extracellular matrix protein 1 (ECM1) is a secreted glycoprotein, which plays an important role in the structural and functional biology of the skin as demonstrated by the identification of loss-of-function mutations in ECM1 as cause of the genodermatosis lipoid proteinosis, characterized by reduplication of the skin basement membrane and hyalinization of the underlying dermis. To search for binding partner(s) of ECM1, we tested the in vitro binding activity of ECM1a, a major isoform of four ECM1 splice variants, to different skin extracellular matrix proteins (such as laminin 332, collagen type IV, and fibronectin) and polysaccharides (such as hyaluronan, heparin, and chondroitin sulfate A) with solid-phase binding assay. We demonstrated that ECM1a utilizes different regions to bind to a variety of extracellular matrix components. Ultrastructurally, ECM1 is a basement membrane protein in human skin and is part of network-like suprastructures containing perlecan, collagen type IV, and laminin 332 as constituents. Furthermore, ECM1a enhanced the binding of collagen IV to laminin 332 dose-dependently, showing its involvement in the dermal-epidermal junction and interstitial dermis and making the functional link to the pathophysiology of lipoid proteinosis. To our knowledge, this is previously unreported.