Antibody to the phosphomannosyl receptor inhibits recycling of receptor in fibroblasts

We cloned and sequenced the 8767-bp full-length cDNA for the chicken cation-independent mannose-6-phosphate receptor (CI-MPR), of interest because, unlike its mammalian homologs, it does not bind insulin-like growth factor II (IGF-II). The cDNA encodes a protein of 2470 aa that includes a putative signal sequence, an extracytoplasmic domain consisting of 15 homologous repeat sequences, a 23-residue transmembrane sequence, and a 161-residue cytoplasmic sequence. Overall, it shows 60% sequence identity with human and bovine CI-MPR homologs, and all but two of 122 cysteine residues are conserved. However, it shows much less homology in the N-terminal signal sequence, in repeat 11, which is proposed to contain the IGF-II-binding site in mammalian CI-MPR homologs, and in the 14-aa residue segment in the cytoplasmic sequence that has been proposed to mediate G-protein-coupled signal transduction in response to IGF-II binding by the human CI-MPR. Transient expression in COS-7 cells produced a functional CI-MPR which exhibited mannose-6-phosphate-inhibitable binding and mediated endocytosis of recombinant human beta-glucuronidase. Expression of the functional chicken CI-MPR in mice lacking the mammalian CI-MPR should clarify the controversy over the physiological role of the IGF-II-binding site in mammalian CI-MPR homologs.

show abstract

“…Previous studies (28) showed that exposure of human cells to antiserum to human CI-MPR inhibits enzyme endocytosis.…”

Section: Resultsmentioning

confidence: 97%

Cloning and expression of the cDNA of chicken cation-independent mannose-6-phosphate receptor.

Zhou

Sly

1995

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

show abstract

“…Fluorescein-conjugated transferrin (Tfn-FITC) was purchased from Molecular Probes (Eugene, OR). Goat antisera with specificity for the human CI-MPR was a kind gift from Dr W. Sly (Saint Louis University School of Medicine, St Louis, MO) 32 and was detected with an R-phycoerythrinconjugated donkey anti-goat IgG F(abЈ) 2 fragment from Jackson ImmunoResearch Laboratories (West Grove, PA). The grB-specific inhibitor was generously supplied by Dr N. A. Thornberry (Merck Research Laboratories, Rahway, NJ).…”

Section: Cell Lines and Reagentsmentioning

confidence: 99%

The granzyme B–serglycin complex from cytotoxic granules requires dynamin for endocytosis

Veugelers

Motyka

Frantz

et al. 2004

Blood

View full text Add to dashboard Cite

Cytotoxic T lymphocytes and natural killer cells destroy target cells via the directed exocytosis of lytic effector molecules such as perforin and granzymes. The mechanism by which these proteins enter targets is uncertain. There is ongoing debate over whether the most important endocytic mechanism is nonspecific or is dependent on the cation-independent mannose 6-phosphate receptor. This study tested whether granzyme B endocytosis is facilitated by dynamin, a key factor in many endocytic pathways. Uptake of and killing by the purified granzyme B molecule occurred by both dynamin-dependent and -independent mechanisms. However most importantly, serglycin-bound granzyme B in high-molecular-weight degranulate material from cytotoxic T lymphocytes predominantly followed a dynamin-dependent pathway to kill target cells. Similarly, killing by live cytotoxic T lymphocytes was attenuated by a defect in the dynamin endocytic pathway, and in particular, the pathways characteristically activated by granzyme B were affected. We therefore propose a model where degranulated serglycin-bound granzymes require dynamin for uptake. IntroductionNatural killer cells and cytotoxic T lymphocytes (CTLs) protect a whole organism against dangerous cells, such as those that are infected with virus or are tumorigenic. 1 When natural killers and CTLs recognize a target cell, the latter is killed by either of 2 major pathways: the Fas-Fas ligand (FasL) pathway, or directional exocytosis of membrane-bound cytotoxic granules present in the cytoplasm of the killer cell. The granules mediate the demise of the target cell via the enclosed cytolytic molecules, which include, among others, a family of serine proteases called granzymes, and the pore-forming molecule, perforin (pfn). [2][3][4] The granzymes, via a pfn-dependent mechanism, induce cell death following translocation across the plasma membrane of target cells. Inside the cell, each granzyme fulfills a critical nonoverlapping role to induce apoptosis by cleaving specific subsets of substrates, such as caspases 5,6 and Bid 7-10 by granzyme B (grB), and the SET complex 11 by granzyme A. But in order to achieve cleavage of substrates, clearly a critical first step is the uptake of granzymes into the target cell.The original model of granzyme internalization proposes translocation via a pfn pore in the plasma membrane. However, recent evidence suggests that granzymes are first internalized via endocytosis and then are released into the cytoplasm with the help of pfn by an unknown mechanism. This model is predominantly based on studies performed with grB, which has largely served as the granzyme prototype to date. The first evidence that granzyme uptake occurred by endocytosis was the finding that grB enters cells autonomously. 12-14 Furthermore, grB binds to the cell surface in a concentration-dependent and saturable manner, 12 suggesting a receptor-mediated endocytic mechanism. The grB endocytosis model has further developed with the identification of the cation-independent mannose 6-phosph...

show abstract

“…The IGF-IIR/MPR has been studied more extensively, and several lines of evidence suggest that it is the more important receptor for sorting acid hydrolases to lysosomes. (i) Antibody to the IGF-IIR/MPR blocked endocytosis and sorting (20)(21)(22). (ii) Mouse cells not expressing the IGF-IIR/MPR secreted most of their newly synthesized lysosomal enzymes (14,23,24).…”

mentioning

confidence: 99%

“…Sorting was inhibited only when both anti-HIGF-IIR/MPR antibody and anti-HSMPR antibody were added to the medium. This result suggested that sorting by the SMPR could only be demonstrated in the absence of the IGF-IIR/ MPR (22). We have transfected the HSMPR cDNA into mouse L cells that are defective for the IGF-IIR/MPR (23).…”

mentioning

confidence: 99%

The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of beta-glucuronidase.

Watanabe

Grubb

Sly

1990

Proc. Natl. Acad. Sci. U.S.A.

View full text Add to dashboard Cite

We studied the function of the human small (46-kDa) mannose 6-phosphate receptor (SMPR) in transfected mouse L cells that do not express the larger insulin-like growth factor II/mannose 6-phosphate receptor. Cells overexpressing human SMPR were studied for enzyme binding to cell surface receptors, for binding to intracellular receptors in permeabilized cells, and for receptor-mediated endocytosis of recombinant human ,3-glucuronidase. Specific binding to human SMPR in permeabilized cells showed a pH optimum between pH 6.0 and pH 6.5. Binding was significant in the presence of EDTA but was enhanced by added divalent cations. Up to 2.3% of the total functional receptor could be detected on the cell surface by enzyme binding. We present experiments showing that at very high levels of overexpression, and at pH 6.5, human SMPR mediated the endocytosis of P-glucuronidase. At pH 7.5, the rate of endocytosis was only 14% the rate seen at pH 6.5. Cells overexpressing human SMPR also showed reduced secretion of newly synthesized ,B-glucuronidase when compared to cells transfected with vector only, suggesting that overexpressed human SMPR can participate in sorting of newly synthesized P-glucuronidase and partially correct the sorting defect in mouse L cells that do not express the insulin-like growth factor II/mannose 6-phosphate receptor.

show abstract

Antibody to the phosphomannosyl receptor inhibits recycling of receptor in fibroblasts

Cited by 33 publications

References 37 publications

Cloning and expression of the cDNA of chicken cation-independent mannose-6-phosphate receptor.

Cloning and expression of the cDNA of chicken cation-independent mannose-6-phosphate receptor.

The granzyme B–serglycin complex from cytotoxic granules requires dynamin for endocytosis

The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of beta-glucuronidase.

Contact Info

Product

Resources

About