Molecular Cloning, Characterization, and Genetic Mapping of the cDNA Coding for a Novel Secretory Protein of Mouse. DEMONSTRATION OF ALTERNATIVE SPLICING IN SKIN AND CARTILAGE

Abstract

A novel 85-kDa protein secreted by the mouse stromal osteogenic cell line MN7 was identified using two-dimensional polyacrylamide gel electrophoresis (Mathieu, E., Meheus, L., Raymackers, J., and Merregaert, J. (1994) J. Bone Miner. Res. 9, 903-913). Degenerate primers were used to isolate the cDNA coding for this protein. The full-length cDNA clone is 1.9 kilobases (kb) and codes for a protein of 559 amino acid residues. The DNA and deduced amino acid sequences have no counterparts in public data bases, but a structural similarity involving typical cysteine doublets can be observed to serum albumin family proteins and to Endo16 (a calcium-binding protein of sea urchin). Northern blot analysis revealed the presence of a 1.9-kb transcript in various tissues, and a shorter transcript of 1.5 kb, derived by alternative splicing in tail, front paw and skin of embryonic mice. The gene for the p85 protein, termed Ecm1 (for extracellular matrix protein 1), is a single-copy gene, which was localized to the region on mouse chromosome 3 known to contain at least one locus associated with developmental disorders of the skin, soft coat (soc). Alternative splicing may serve as a mechanism for generating functional diversity in the Ecm1 gene.

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

“…SASDL2ϩ, which contains a piece of exon 6 and the complete alternatively spliced exon 7, includes a part of the two tandem repeat domains of ECM1a (aa 151-405) (Fig. 3A) (1,8). Approximately 50% of the mutations in patients with lipoid proteinosis cluster to exons 6 and 7 of the ECM1 gene, which makes this an attractive region to look for protein interaction partners of ECM1a (35).…”

“…The ECM1 protein contains a 19-amino acid signal peptide followed by four domains: a cysteine-free N-terminal segment, two tandem repeats, and a C-terminal segment. The two tandem repeats and the C-terminal domain contain cysteines in a typical CC-(X 7-10 )C arrangement that is capable of forming protein double loops that could be involved in protein-protein interactions (1,8). More recently, a rudimentary three-dimensional model divided the ECM1a protein into four distinct domains: an NH 2 -terminal domain forming ␣-helical structures, followed by three domains, whose amino acid sequences were highly comparable with the third domain of human serum albumin: SASDL2 (serum albumin subdomain-like 2), SASDL3, and SASDL4 (9).…”

Importance of Extracellular Matrix Protein 1 (ECM1) in Maintaining the Functional Integrity of the Human Skin

“…1A) [2,3]. The protein harbours abundant cysteine residues (4.8%) with a typical arrangement distributed according to the CC-(X [7][8][9][10] )-C pattern of six cysteine doublets [2]. This cysteine arrangement, which is also found in serum albumin [14,15] and in the sea urchin Endo16 protein [16] may form doubleloop structures [14,17,18], specifying putative important biological ligand interactions [16,19].…”

“…1A) [2,3]. The protein harbours abundant cysteine residues (4.8%) with a typical arrangement distributed according to the CC-(X [7][8][9][10] )-C pattern of six cysteine doublets [2].…”

“…The mouse extracellular matrix protein 1 (Ecm1) was first identified in 1994 as a novel glycosylated 85 kDa protein secreted in the conditioned medium of the murine osteogenic stromal cell line, MN7 [1,2]. The human ECM1 gene is located on chromosome 1q21.2 [3,4] and encodes for four splice variants (Fig.…”