Glycosylation directs targeting and activation of cystatin f from intracellular and extracellular sources.
Cystatin F is a cysteine protease inhibitor that is selectively expressed in immune cells and unlike other cystatin family members is targeted to a significant extent to intracellular compartments. Initially made as an inactive glycosylated disulfide-linked dimer, cystatin F is converted to an active monomer by proteolytic cleavage following transport to the endosomal/lysosomal system. This active form of cystatin F targets cathepsin C/DPPI and probably other cathepsins in immune cells. We show that efficient targeting of cystatin F to the endocytic pathway is dependent not on its unique dimeric conformation but rather on its oligosaccharide chains. We demonstrate the unusual addition of N-linked sugars to an Asn-X-Cys motif in cystatin F and provide evidence that the mannose 6-phosphate sorting machinery is used to divert cystatin F from the secretory pathway and to mediate its uptake from extracellular pools. These studies identify a function for the oligosaccharides on cystatin F and raise the possibility that cystatin F might regulate proteases in trans by secretion in an inactive form by one cell and subsequent internalization and activation by another cell.
Colbert
Jeff D
JD
Division of Cell Biology and Immunology, College of Life Sciences, University of Dundee, Dundee, UK. j.colbert@dundee.ac.uk
Plechanovová
Anna
A
Watts
Colin
C
D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000378:metabolism; D014408:Biomarkers, Tumor / Q000737:chemistry / Q000378:metabolism*; D002460:Cell Line; D015891:Cystatins / Q000737:chemistry / Q000378:metabolism*; D019281:Dimerization; D011992:Endosomes / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D008360:Mannosephosphates / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009844:Oligosaccharides / Q000737:chemistry / Q000378:metabolism; D011487:Protein Conformation
D014408:Biomarkers, Tumor; C116891:CST7 protein, human; D015891:Cystatins; D008360:Mannosephosphates; D009844:Oligosaccharides; C027693:mannose-6-phosphate; D001216:Asparagine