Mannose 6-phosphate

Abstract: The activation of latent transforming growth factor beta (LTGF-beta) normally seen in cocultures of bovine aortic endothelial and bovine smooth muscle cells can be inhibited by coculturing the cells with either mannose 6-phosphate (Man-6-P) or antibodies directed against the cation-independent Man-6-P/insulin-like growth factor type II receptor (anti-Man-6-PR). This result was established by measuring the ability of coculture conditioned medium (formed with or without Man-6-P or anti-Man-6-PR) to suppress bovine aortic endothelial cell migration and protease production, activities previously shown to be related to transforming growth factor beta activity. The inhibition by Man-6-P is dose dependent, with maximal inhibition seen at 100 microM and is specific because mannose 1-phosphate and glucose 6-phosphate do not interfere with activation of LTGF-beta. The inhibitory effect of anti-Man-6-PR is also specific and dose dependent; maximal inhibition of activation occurs at 400 micrograms/ml. Control experiments indicate that Man-6-P and anti-Man-6-PR do not interfere with the basal level of migration of bovine aortic endothelial cells, the migration observed when exogenous transforming growth factor beta is added, the activation of transforming growth factor beta by plasmin or transient acidification, and the release of LTGF-beta. Thus, binding to the cation-independent Man-6-P/insulin-like growth factor type II receptor appears to be a requirement for activation of LTGF-beta.

Abstract: The targeting of lysosomal enzymes from their site of synthesis in the rough endoplasmic reticulum (RER) to their final destination in lysosomes is directed by a series of protein and carbohydrate recognition signals on the enzymes. Lysosomal enzymes, along with secretory and plasma membrane proteins, contain amino-terminal signal sequences that direct the vectorial discharge of the nascent proteins into the lumen of the RER. The three classes of proteins also share a common peptide signal for asparagine glycosylation. The next signal is unique to lysosomal enzymes and permits their high-affinity binding to a specific phosphotransferase that catalyzes the formation of the mannose 6-phosphate recognition marker. This carbohydrate determinant allows binding to specific receptors that translocate the lysosomal enzymes from the Golgi complex to an acidified prelysosomal compartment. There the lysosomal enzymes are discharged for final packaging into lysosomes. Two distinct mannose 6-phosphate receptors have been identified, and cDNAs encoding their entire sequences have been cloned. An analysis of the deduced amino acid sequences of the receptors shows that each is composed of four structural domains: a signal sequence, an extracytoplasmic amino-terminal domain, a hydrophobic membrane-spanning region, and a cytoplasmic domain. The entire extracytoplasmic region of the small receptor is homologous to the 15 repeating domains that constitute the extracytoplasmic portion of the large receptor.

Abstract: The targeting of lysosomal enzymes from their site of synthesis in the rough endoplasmic reticulum (RER) to their final destination in lysosomes is a multi-step process requiring a series of interactions between cellular components and protein and carbohydrate recognition signals present on the lysosomal enzymes (1-6). These proteins share a common pathway with secretory proteins and membrane proteins during the early stages of their biosynthesis. All three classes of proteins contain a hydrophobic signal sequence that allows for their synthesis on membrane-bound polysomes in the RER and translocation into t.he lumen of this organelle. During this process the lysosomal enzymes as well as many secretory and membrane proteins are co-translationally glycosylated at selected asparagine residues. Following cleavage of the signal sequence and initial proc-essing of asparagine-linked oligosaccharides, the proteins move by vesicular transport from the RER to the Golgi apparatus where they undergo a variety of post-translational modifications and are segre- gated from one another for targeting to their final destinations (7). A key step in the sorting process is the generation of phospho- mannosyl residues on the lysosomal enzymes. The phosphorylating enzyme recognizes a protein determinant shared by lysosomal en- zymes, thereby selectively marking this class of proteins for subse-quent segregation. The phosphomannosyl residues serve as high affinity ligands for binding to mannose 6-phosphate receptors (MPRs)