Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.4.21.64 (proteinase K)
 4,071 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Embryonic data and ultrastructural analyses suggest that the primitive endothelium signals undifferentiated mesenchymal cells to migrate to the forming blood vessel and subsequently regulates mural cell growth and behavior. Upon maturation of the blood vessel, chemotactic and mitogenic signals are apparently diminished and differentiated smooth muscle cells normally remain quiescent. This homeostasis is seemingly upset in conditions which lead to pathologies characterized by smooth muscle cell hyperplasia such as atherosclerosis. By culturing endothelial cells at different densities, we attempted to re-create the various stages of vascular development. Whereas media conditioned by sparse endothelial cells stimulate smooth muscle cells, media conditioned by dense endothelial cell cultures are inhibitory. Culture of sparse smooth muscle cells in media conditioned for 3 days by postconfluent endothelial cell cultures leads to dose-dependent and reversible smooth muscle cell inhibition. Furthermore, in the presence of the endothelial cell-derived inhibitor, smooth muscle cells are rendered refractory to mitogens such as fibroblast growth factor and platelet-derived growth factor. The inhibitory activity is not attributable to the well-characterized inhibitors of smooth muscle cell growth, transforming growth factor type-beta, prostaglandin I2, or heparan sulfate proteoglycan. Partial characterization of the inhibitory conditioned media suggests that the active molecule is smaller than 1,000 da, and stable to boiling as well as proteinase K and heparinase digestion. These findings support the concept that there is intercellular communication between endothelial cells and smooth muscle cells and provide evidence for a novel endothelial cell-derived smooth muscle cell growth inhibitor.
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PMID:Density-dependent endothelial cell production of an inhibitor of smooth muscle cell growth. 822 80

Recombinant adeno-associated viruses (AAVs) are promising vectors in the field of gene therapy. Different AAV serotypes display distinct tissue tropism, believed to be related to the distribution of their receptors on target cells. Of the 11 well-characterized AAV serotypes, heparan sulfate proteoglycan and sialic acid have been suggested to be the attachment receptors for AAV type 2 and types 4 and 5, respectively. In this report, we identify the receptor for the two closely related serotypes, AAV1 and AAV6. First, we demonstrate using coinfection experiments and luciferase reporter analysis that AAV1 and AAV6 compete for similar receptors. Unlike heparin sulfate, enzymatic or genetic removal of sialic acid markedly reduced AAV1 and AAV6 binding and transduction. Further analysis using lectin staining and lectin competition assays identified that AAV1 and AAV6 use either alpha2,3-linked or alpha2,6-linked sialic acid when transducing numerous cell types (HepG2, Pro-5, and Cos-7). Treatment of cells with proteinase K but not glycolipid inhibitor reduced AAV1 and AAV6 infection, supporting the hypothesis that the sialic acid that facilitates infection is associated with glycoproteins rather than glycolipids. In addition, we determined by inhibitor (N-benzyl GalNAc)- and cell line-specific (Lec-1) studies that AAV1 and AAV6 require N-linked and not O-linked sialic acid. Furthermore, a resialylation experiment on a deficient Lec-2 cell line confirmed a 2,3 and 2,6 N-linked sialic acid requirement, while studies of mucin with O-linked sialic acid showed no inhibition effect for AAV1 and AAV6 transduction on Cos-7 cells. Finally, using a glycan array binding assay we determined that AAV1 efficiently binds to NeuAcalpha2-3GalNAcbeta1-4GlcNAc, as well as two glycoproteins with alpha2,3 and alpha2,6 N-linked sialic acids. Taken together, competition, genetic, inhibitor, enzymatic reconstitution, and glycan array experiments support alpha2,3 and alpha2,6 sialic acids that are present on N-linked glycoproteins as primary receptors for efficient AAV1 and AAV6 viral infection.
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PMID:Alpha2,3 and alpha2,6 N-linked sialic acids facilitate efficient binding and transduction by adeno-associated virus types 1 and 6. 1694 May 21