Mutational analysis of the membrane-proximal cleavage site of L- selectin: relaxed sequence specificity surrounding the cleavage site
| Autor: | Julius Kahn, Grace I. Migaki, Takashi Kei Kishimoto |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 1995 |
| Předmět: |
Cleavage factor
Proteolysis Immunology Molecular Sequence Data Receptors Lymphocyte Homing Down-Regulation Cleavage and polyadenylation specificity factor Biology Cleavage (embryo) Transfection Mice Structure-Activity Relationship Endopeptidases medicine Immunology and Allergy Animals Humans Amino Acid Sequence L-Selectin Peptide sequence Cleavage stimulation factor Membrane Glycoproteins medicine.diagnostic_test Sequence Homology Amino Acid Articles Molecular biology Transmembrane protein Rats Transmembrane domain Mutagenesis Site-Directed Cell Adhesion Molecules Sequence Alignment |
| Zdroj: | The Journal of Experimental Medicine |
| ISSN: | 1540-9538 0022-1007 |
| Popis: | L-selectin expression is regulated in part by membrane-proximal cleavage from the cell surface of leukocytes and L-selectin-transfected cells. The downregulation of L-selectin from the surface of neutrophils is speculated to be a process involved in the adhesion cascade leading to neutrophil recruitment to sites of inflammation. We previously reported that L-selectin is cleaved between Lys321 and Ser322 in a region that links the second short consensus repeat (SCR) and the transmembrane domain. We demonstrate that replacing this cleavage domain of L-selectin with the corresponding region of E-selectin prevents L-selectin shedding, as judged by inhibiting the generation of the 68-kD soluble and 6-kD transmembrane cleavage products of L-selectin. Unexpectedly, we found that point mutations of the cleavage site, as well as mutations of multiple conserved amino acids within the cleavage domain, do not significantly affect L-selectin shedding. However, short deletions of four or five amino acids in the L-selectin cleavage domain inhibit L-selectin downregulation. Mutations that appeared to inhibit L-selectin shedding resulted in higher levels of cell surface expression, consistent with a lack of apparent proteolysis from the cell membrane. One deletion mutant, I327 delta N332, retains the native cleavage site yet inhibits L-selectin proteolysis as well. Restoring the amino acids deleted between I327 and N332 with five alanine residues restores L-selectin proteolysis. Thus, the proteolytic processing of L-selectin appears to have a relaxed sequence specificity at the cleavage site, and it may depend on the physical length or other secondary structural characteristics of the cleavage domain. |
| Databáze: | OpenAIRE |
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