title: Structural requirements regulate endoproteolytic release of the L-selectin (CD62L) adhesion receptor from the cell surface of leukocytes.
abstract: L-selectin mediates leukocyte rolling on vascular endothelium at sites of inflammation and lymphocyte migration to peripheral lymph nodes. L-selectin is rapidly shed from the cell surface after leukocyte activation by a proteolytic mechanism that cleaves the receptor in a membrane proximal extracellular region. This process may allow rapid leukocyte detachment from the endothelial surface before entry into tissues. In this study, the structural requirements for regulation of human L-selectin endoproteolytic release were examined through analysis of chimeric selectin molecules and mutant L-selectin receptors. The use of chimeric selectins and a cytoplasmic tail truncation mutant demonstrated that the extracellular membrane-proximal 15-amino acid region of L-selectin is required for endoproteolytic release. The introduction of alanine-scanning mutations within this membrane-proximal region did not prevent endoproteolytic release, indicating that a specific amino acid motif was not an absolute requirement for cleavage. Furthermore, alterations within the putative primary cleavage site (K283-S284) resulted in either constitutive endoproteolytic release of the receptor or inhibition of cell activation-induced shedding to variable extents. The length of the membrane-proximal region was also critical since truncations of this region completely abolished endoproteolytic release. Thus, release of L-selectin is likely to be regulated by the generation of an appropriate tertiary conformation within the membrane-proximal region of the receptor which allows recognition by a membrane-bound endoprotease with relaxed sequence specificity that cleaves the receptor at a specific distance from the plasma membrane. These observations suggest a generalized protein-processing pathway involved in the endoproteolytic release of specific transmembrane proteins which harbor widely differing primary sequences at or neighboring their cleavage sites.
authors.affiliation: Department of Immunology, Duke University Medical Center, Durham, North Carolina 27710, USA.
authors.forename: A
authors.initials: A
authors.lastname: Chen
authors.forename: P
authors.initials: P
authors.lastname: Engel
authors.forename: T F
authors.initials: TF
authors.lastname: Tedder
mesh_terms: D000595:Amino Acid Sequence; D015815:Cell Adhesion Molecules / Q000737:chemistry / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D010450:Endopeptidases / Q000378:metabolism*; D019041:L-Selectin; D008562:Membrane Glycoproteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D016201:Receptors, Lymphocyte Homing / Q000737:chemistry / Q000378:metabolism*; D011993:Recombinant Fusion Proteins; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D012995:Solubility; D013329:Structure-Activity Relationship; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
chemical_list: D015815:Cell Adhesion Molecules; D008562:Membrane Glycoproteins; D016201:Receptors, Lymphocyte Homing; D011993:Recombinant Fusion Proteins; D019041:L-Selectin; D010450:Endopeptidases; D013755:Tetradecanoylphorbol Acetate