Molecular Characterization of a Novel Basement Membrane-associated Proteoglycan, Leprecan

Abstract

A monoclonal antibody was used in early studies to identify a novel chondroitin sulfate proteoglycan, secreted by L-2 cells, the core protein of which was approximately 100 kDa. To characterize this proteoglycan core protein at the molecular level, an L-2 cell cDNA library was probed by expression screening and solution hybridization. Northern blot analysis assigned transcript size to approximately 3.1 kilobases and, after contig assembly, the coding region of the mRNA corresponded to 2.18 kilobases. Immunoassays were performed to confirm the identity of this sequence, using a polyclonal antibody raised against an expressed fusion protein encoded by sequence representing the carboxyl half of the molecule. The antibody recognized the core protein in Western blots after prior digestion of the intact proteoglycan with chondroitinase ABC. Immunostaining tissue sections with the same antibody localized the proteoglycan to basement membranes, and expression of the entire sequence in Chinese hamster ovary K-1 cells showed that the protein encoded by the sequence secreted as a chondroitin sulfate proteoglycan. The core protein not only has motifs permitting glycosylation as a proteoglycan, but also possesses the endoplasmic reticulum retrieval signal, KDEL, which suggests that, in addition to its role as a basement membrane component, it may also participate in the secretory pathway of cells.

Gros1, a potential growth suppressor on chromosome 1: its identity to basement membrane-associated proteoglycan, leprecan

“…The actual number and size of GAG chains attached to Gros1/leprecan remains to be known. The size of the secreted form was identical to that of the cellular form (Wassenhove-McCarthy and McCarthy, 1999). They have also shown that the secreted form retains C-terminal KDEL signal and present evidence for the secretion of native leprecan.…”

“…Overexpression of the long form (84-kDa) of the protein is seen to have growth suppressive activity in NIH3T3 cells, consistent with its down regulation in many of the human transformed cells and similar to some other proteoglycans. Based on the structural characterization of the protein (Wassenhove-McCarthy and McCarthy, 1999), chromosomal localization and biological activity, it may contribute to the tumor suppressor activity lost in many chromosome 1-linked malignancies. Functional mechanism for the role of this novel proteoglycan in growth suppression, signi®cance of di erentially spliced forms and Gros1/leprecan interposed cancer therapy warrants further investigations.…”

“…The gene is therefore named Gros1 for Growth Suppressor on chromosome 1. An independent study cloned rat homolog of Gros1 as a novel leucine proline-enriched basement membrane-associated proteoglycan, leprecan (Wassenhove-McCarthy and McCarthy, 1999). Taken together, Gros1/leprecan is a novel proteoglycan and tumor suppressor whose genetic alterations may be involved in chromosome 1 linked human malignancies.…”

Characterization of Recombinant Human Prolyl 3-Hydroxylase Isoenzyme 2, an Enzyme Modifying the Basement Membrane Collagen IV

“…Data base analyses of vertebrate genomes pointed to the existence of a family with three members, named P3Hs 1-3 (11). P3H1 was found to be identical to leprecan, or growth suppressor 1 (Gros1), which had previously been identified as a basement membrane-associated glycoprotein in rats and as a potential growth suppressor in mice (12,13). The leprecan-like 1 protein, which corresponds to P3H2, has been shown to be localized to the endoplasmic reticulum and Golgi network and to be expressed in many secretory tissues (14).…”

The prolyl 3-hydroxylases P3H2 and P3H3 are novel targets for epigenetic silencing in breast cancer

“…The prolyl 3-hydroxylases (P3H), P3H2 and P3H3, were originally termed Leprel1 and Leprel2 due to their 'Leprecan-like' amino-acid sequence identity to Leprecan, now termed P3H1 (Wassenhove-McCarthy and McCarthy, 1999;Jarnum et al, 2004;Vranka et al, 2004). Along with the collagen prolyl 4-hydroxylases (c-P4H) and lysyl hydroxylases (LH), the P3H belong to the 2-oxoglutarate dioxygenases (Vranka et al, 2004).…”