title: Cleavage within Reelin repeat 3 regulates the duration and range of the signaling activity of Reelin protein.
abstract: Reelin is a secreted glycoprotein that plays essential roles in the brain. Reelin is specifically cleaved at two distinct sites, called N-t and C-t, with the former being the major one. N-t cleavage can occur both in the extracellular space and in the endosomes, although the physiological importance of endosomal N-t cleavage has not been investigated. In this study, we first determined the exact N-t cleavage site catalyzed by a protease secreted by cerebral cortical neurons. Cleavage occurred between Pro-1244 and Ala-1245 within Reelin repeat 3. A Reelin mutant in which Pro-1244 was replaced with aspartate (Reelin-PD) was resistant to a protease secreted by cultured cerebral cortical neurons, and its biological activity stayed active longer than that of wild-type Reelin. Interestingly, Reelin-PD remained in the intracellular compartments longer than wild-type Reelin and persistently activated downstream signaling. Therefore, N-t cleavage of Reelin is required for halting the signaling machinery in the extracellular space as well as within endosomes of target neurons. We established a monoclonal antibody specific to uncleaved Reelin protein and found that it is localized in the vicinity of Reelin-producing cells, whereas the N-terminal fragment diffuses, or is transported, to distant regions. These data demonstrate that N-t cleavage of Reelin plays critical roles in regulating the duration and range of Reelin functions both in the extracellular milieu and in the intracellular compartments.
authors.affiliation: From the Department of Biomedical Science, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya, Aichi 467-8603, Japan.
authors.forename: Mari
authors.initials: M
authors.lastname: Koie
authors.forename: Kyoko
authors.initials: K
authors.lastname: Okumura
authors.forename: Arisa
authors.initials: A
authors.lastname: Hisanaga
authors.forename: Takana
authors.initials: T
authors.lastname: Kamei
authors.forename: Kazutomo
authors.initials: K
authors.lastname: Sasaki
authors.forename: Mengyan
authors.initials: M
authors.lastname: Deng
authors.forename: Atsushi
authors.initials: A
authors.lastname: Baba
authors.forename: Takao
authors.initials: T
authors.lastname: Kohno
authors.forename: Mitsuharu
authors.initials: M
authors.lastname: Hattori
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001224:Aspartic Acid / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D015153:Blotting, Western; D015816:Cell Adhesion Molecules, Neuronal / Q000235:genetics* / Q000378:metabolism; D002478:Cells, Cultured; D011992:Endosomes / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000235:genetics* / Q000378:metabolism; D005110:Extracellular Space / Q000378:metabolism; D005260:Female; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D009419:Nerve Tissue Proteins / Q000235:genetics* / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism; D011392:Proline / Q000235:genetics* / Q000378:metabolism; D059748:Proteolysis; D000091042:Reelin Protein; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics*
chemical_list: D015816:Cell Adhesion Molecules, Neuronal; D016326:Extracellular Matrix Proteins; D009419:Nerve Tissue Proteins; D000091042:Reelin Protein; D001224:Aspartic Acid; D011392:Proline; D010447:Peptide Hydrolases; C000717127:RELN protein, human; C000717128:Reln protein, mouse; D012697:Serine Endopeptidases
keywords: ADAM ADAMTS; Brain; Dab1; Endosomes; Neurons; Protein Degradation; Reelin; Signal Transduction