ABSTRACT
Japanese species of the genus Diplazon Nees, 1819 are reviewed. Nine species are identified from Japan and one of them, D. pallicoxa Manukyan, 1987 is newly recorded. The distribution records of D. orientalis (Cameron, 1905) and D. tibiatorius (Thunberg 1822) are removed from the Japanese fauna. A key to Japanese species of this genus is provided.
Subject(s)
Hymenoptera , Animals , Hymenoptera/classification , Japan , Species SpecificityABSTRACT
Reelin is a glycoprotein essential for brain development and functions. Reelin is subject to specific proteolysis at two distinct (N-t and C-t) sites, and these cleavages significantly diminish Reelin activity. The decrease of Reelin activity is detrimental for brain function, but the protease that catalyzes specific cleavage of Reelin remains elusive. Here we found that a disintegrin and metalloproteinase with thrombospondin motifs 4 (ADAMTS-4) cleaves Reelin in an isoform-specific manner. Among ADAMTS-4 isoforms, p50 cleaves the N-t site only, while p75 cleaves both sites. This is the first report identifying a protease that can specifically cleave Reelin.
Subject(s)
ADAM Proteins/metabolism , Cell Adhesion Molecules, Neuronal/metabolism , Extracellular Matrix Proteins/metabolism , Nerve Tissue Proteins/metabolism , Procollagen N-Endopeptidase/metabolism , Serine Endopeptidases/metabolism , ADAM Proteins/chemistry , ADAM Proteins/genetics , ADAMTS4 Protein , Animals , Base Sequence , Binding Sites , Brain/embryology , Brain/metabolism , Cell Adhesion Molecules, Neuronal/chemistry , Cell Adhesion Molecules, Neuronal/genetics , DNA Primers/genetics , Extracellular Matrix Proteins/chemistry , Extracellular Matrix Proteins/genetics , Female , Gene Expression Regulation, Developmental , Gene Expression Regulation, Enzymologic , In Situ Hybridization , Isoenzymes/chemistry , Isoenzymes/genetics , Isoenzymes/metabolism , Mice , Mice, Inbred ICR , Nerve Tissue Proteins/chemistry , Nerve Tissue Proteins/genetics , Peptide Fragments/chemistry , Peptide Fragments/genetics , Peptide Fragments/metabolism , Pregnancy , Procollagen N-Endopeptidase/chemistry , Procollagen N-Endopeptidase/genetics , Proteolysis , RNA, Messenger/genetics , RNA, Messenger/metabolism , Recombinant Proteins/chemistry , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Reelin Protein , Serine Endopeptidases/chemistry , Serine Endopeptidases/genetics , Substrate Specificity , Tissue DistributionABSTRACT
Reelin is a 3461-residue secreted glycoprotein that plays a critical role in brain development through its action on target neurons. Although it is known that functional reelin protein exists as multimer formed by interchain disulfide bond(s) as well as through non-covalent interactions, the chemical nature of the multimer assembly has been elusive. In the present study, we identified, among 122 cysteines present in full-length reelin, the single critical cysteine residue (Cys(2101)) responsible for the covalent multimerization. C2101A mutant reelin failed to assemble into disulfide-bonded multimers, whereas it still exhibited non-covalently associated high molecular weight oligomeric states in solution. Detailed analysis of tryptic fragments produced from the purified reelin proteins revealed that the minimum unit of the multimer is a homodimeric reelin linked via Cys(2101) present in the central region and that this cysteine does not connect to the N-terminal region of reelin, which had been postulated as the primary oligomerization domain. A surface plasmon resonance binding assay confirmed that C2101A mutant reelin retained binding capability toward two neuronal receptors apolipoprotein E receptor 2 and very low density lipoprotein receptor. However, it failed to show signaling activity in the assay using the cultured neurons. These results indicate that an intact higher order architecture of reelin multimer maintained by both Cys(2101)-mediated homodimerization and other non-covalent association present elsewhere in the reelin primary structure are essential for exerting its full biological activity.
