Mannose 6-phosphate receptors and lysosomal enzyme targeting.

CHEMISTRY THE JOURNALOF BIOLOGICAL Vol. 264 No. 21 Issue of July 25, pp. 12115-12118,1989 0 1989 by The American Swi& for Biochemistw and MolecularBiology, Inc.

Minireview

Printed in U.S.A.

2269-residue extracytoplasmic domain,a single 23-residuetransmembrane region, and a 163-residue carboxyl-terminal cytoplasmic domain (Fig. l ) . T h e extracytoplasmic domain has 19 potential glycosylation sites, anda t least two are utilized. Therefore, the size of the mature glycoprotein is likely to be between 275 and 300 kDa. The Nancy M. DahmsS, Peter Lobel$, a n d extracytoplasmic domain has a highly repetitive structure consisting Stuart KornfeldT of 15 contiguous units that have a n average length of 147 amino acids. When the repeating units are compared to each other, numerous From the $Department of Biochemistry, Medical College of sequence identities are seen with the percent of identical residues Wisconsin, Milwaukee, WI, the §Center forAdvanced ranging from 16 to 38%. In addition, there are numerous regions of Biotechnology and Medicine, Piscataway, NJ, and the conservatively substituted amino acids and a characteristic spacing BDepartrnent of Medicine, Washington University School of cysteine residues. The receptor is known to be phosphorylated (18, of Medicine, St. Louis, Missouri 63110 19), and analysis of the cytoplasmic domain reveals sequences that are potential substrates for various protein kinases including protein The targeting of lysosomal enzymes from their site of synthesis in kinase C and casein kinasesI and I1 (13). the rough endoplasmic reticulum (RER) to their final destination in The predicted structure of the bovine CD-MPR consists of a 28lysosomes is a multi-step process requiring a series of interactions residue amino-terminal signalsequence,a159-residue extracytobetween cellular components and protein and carbohydraterecogni- plasmic domain, a single 25-residue transmembrane region, and a 67tion signals present on the lysosomal enzymes (1-6). These proteins residue carboxyl-terminal cytoplasmic domain (Fig. 1). This receptor share a commonpathwaywithsecretoryproteinsandmembrane has five potential asparagine-linked glycosylation sites, four of which proteins during the early stages of their biosynthesis.All three classes are utilized (14). of proteins contain ahydrophobicsignalsequence that allows for A comparison of the sequences of the two receptors reveals that theirsynthesisonmembrane-bound polysomes intheRERand they arerelated. The entire extracytoplasmic domain of the CD-MPR translocation into t.he lumen of this organelle. During this process is similar to each of the repeating unitsof the CI-MPR, with sequence the lysosomalenzymes as well as many secretory and membrane identity ranging from 14 to 28%. Thus, the CD-MPR is almost as proteinsareco-translationally glycosylated a t selected asparagine similar to the different repeating units of the CI-MPR as the repeating residues. Following cleavage of the signal sequence and initial proc- units are to each other. This similarity suggests that thetwo receptors essing of asparagine-linked oligosaccharides, the proteins move by share a common ancestry and that the CI-MPR arose from duplicavesicular transport from the RER to theGolgi apparatus where they tion of a single ancestral sequence. In contrast to these homologies, undergo a variety of post-translational modifications and are segre- there are no significant primary sequence similarities between their gated from one anotherfor targeting to their final destinations (7). signal sequences, transmembrane regions, or their cytoplasmic doA key step in the sorting process is the generation of phospho- mains. However, the cytoplasmic domains of both receptors contain mannosyl residues on the lysosomal enzymes. The phosphorylating potential serine phosphorylation sites and clusters of acidic residues enzyme recognizes a protein determinant shared by lysosomal en- that are also found on other recycling receptors (20). zymes, thereby selectively marking this class of proteins for subseChemical cross-linking experiments indicate that the CD-MPR is quent segregation. The phosphomannosyl residuesserve as high a dimer in the membrane (21,22) and either a dimer or a tetramer in affinity ligandsfor binding tomannose6-phosphatereceptors solution (23). The quaternary structureof the GI-MPR has notbeen (MPRs) in the Golgi. In thisway the lysosomal enzymes arephysically analyzed in great detail, but hydrodynamic measurements are conseparated from proteins destined for secretion. The ligand-receptor sistent with it being a monomer (24) while chemical cross-linking complex then exits the Golgi via a coated vesicle and is delivered to experiments indicate that it maybe an oligomer (21). Ligand binding a prelysosomal acidifiedcompartment where dissociationof the ligand studies have revealed that the CD-MPR binds1 mol of the monovaoccurs. The released lysosomal enzyme is packaged into a lysosome lent ligand Man-6-P and 0.5 mol of a divalent phosphorylated oligowhile the receptor either returns to the Golgi to repeat theprocess or saccharide/monomeric subunit (9). Therefore each functional dimer moves to the plasma membrane where it functions to internalize would have two Man-6-P binding sites, both of which can be occupied exogenous lysosomal enzymes.Recent work has focused on the MPRs. by a single oligosaccharide containing two Man-6-P residues. EviTwo different MPRs have been identified, characterized, and their dence that each polypeptide monomer can fold into an independent cDNAs cloned. The routing of the receptors has been studied, and ligand bindingunithasbeenobtained by demonstratingthat a the determinants on the receptor that mediate trafficking are begin- truncated form of the bovine CD-MPR, which behaves as a soluble ning to be defined. Some of the cellular components that interact monomer in solution, is capable of binding Man-6-P (22). The CIwith the receptors as they move from one compartment to the next MPR, on the other hand, binds 2 mol of Man-6-P or 1 mol of a have been identified. This review will summarize our current under- divalent phosphorylated oligosaccharide/monomer (8).This may instanding of the MPRs and theirbiological functions. dicate that only two of the 15 repeating segments of this receptor function in the binding of Man-6-P. While the identity of the binding Receptor Structure segmentshasnotyet been established, two differentproteolytic The first MPR to be characterized was a membrane-associated fragments of the CI-MPR encompassing repeating units 1-3 and 710 have been shown to bind a phosphorylated lysosomal enzyme.' glycoprotein with an apparent M, of 215,000. This receptor binds Thus the two functional Man-6-P binding domains are probably ligand independent of divalent cations. The other MPR is alsoa membrane-associated glycoprotein, but it has an apparent M, of contained within these regions of the receptor. 46,000 and requires divalent cations for optimal ligand binding. Both Role of the Receptors in Sorting and Endocytosis receptors show similar, but not identical, bindingspecificities toward phosphorylated oligosaccharides (8, 9). Based on their divalent catTwo complementary experimental approaches indicate that the CIionrequirements, we refer tothelargerreceptor as thecationMPR functions both in the sorting of newly synthesized lysosomal independent(CI)MPRandthesmallerreceptor as thecationenzymes and in endocytosis of extracellular phosphorylatedlysosomal dependent (CD) MPR. The cloning of cDNAs for the CI-MPR from enzymes. First, cultured cells that either lack endogenous CI-MPR bovine (lo), human (11, 12), and rat sources (13) and cDNAs for the (25) or are depleted of CI-MPR by treatment with anti-CI-MPR CD-MPR frombovine (14) and human sources (15) has provided antiserum (26,27) secrete-70% of their newly synthesized lysosomal insights into the relationshipbetween these two different proteins. enzymes and do not endocytose extracellular phosphorylated lysosoSequence analyses, combined with proteolysis experimentsof the mal enzymes. Second, the defective sorting and endocytosis phenoreceptor in membranes (16, 17), indicate that the bovine CI-MPR precursor consists of a 44-residue amino-terminal signal sequence, a ' B. Wevtlund and S. Kornfeld, unpublished data.

Mannose 6-Phosphate Receptors and Lysosomal EnzymeTargeting

12115

Minireview: Man-6-P Receptors and Lysosomal Enzyme Targeting

12116 NH2

lgnal sequence

zymes. In these instances, IGF-I1 might act by inhibiting lysosomal enzyme binding to cell surface receptors thereby preventing the recapture of secreted lysosomal enzymes (34) or by altering the intracellular trafficking and distribution of the CI-MPR, possibly resulting in less efficient sorting in the Golgi (39). Interestingly, the chicken CI-MPR does not bind either human or chicken IGF-I1 (40). Nevertheless, chicken fibroblasts are highly responsive to IGF-11, possibly via binding to the IGF-I receptor (41).

Other Growth Factors Bind to the MPR Recently a number of secreted glycoproteins has been shown to contain the Man-6-P recognition marker (42-47). Presumably these ligands are secreted because they have a low affinity for the MPR and do not effectively compete with the bulk of lysosomal enzymes at the intracellular sorting site (48). Three of these proteins have been identified as lysosomal enzymes that are secreted only under special circumstances (42-44). Another is porcine thyroglobulin which is secreted by thyroid follicle cells and then recaptured for degradation in lysosomes (45). The other proteins appear to be growth sequence factors with no known lysosomal enzyme activity. Proliferin is a Extracellular Portion prolactin-related proteinpostulated to be an autocrine growth factor while transforming growth factor-81 precursor is the proform of a Cytoplasmic hormone that has multiple effects on cell growth and differentiation COOH Portton (46, 47). Both of these proteins can bind to the CI-MPR a t the cell CdOH CdOH surface via their Man-6-Pmoieties. Their internalization could result MPR~’ MPRCo in activation in endosomes or degradation in lysosomes. These findFIG.1. Schematic representation of the MPRs. 0,N-linked glycosylation sites known to be used; 0,potential glycosylation sites. The insertion in domain ings indicate that thephosphomannosyl recognition system may have a broader biologic rolethan previously recognized. 13 of the CI-MPR represents theregion that is similar to the typeI1 repeats of Repeatmg Units

a

fibronectin. Modified from Ref. 14.

Receptor Trafficking type of the cells that lack endogenous CI-MPR can be corrected by transfection with a CI-MPR cDNA (28, 29). The residual sorting found in the CI-MPR defective cells appears to be mediated by the CD-MPR. Supporting this, when such cells are treated with an antiCD-MPR antiserum, secretion of lysosomal enzymes is increased (27). These results indicate that both receptors function in lysosomal enzyme sorting, with the CI-MPR handling most of the traffic. One clear difference between the two receptors relates to the endocytosis of extracellular lysosomal enzymes. While both receptors cycle to the plasma membrane (30), only the CI-MPR is capable of binding and internalizing lysosomal enzymes (27). This difference, however, does not resolve the fundamental question ofwhy two separate MPRs exist.

Lysosomal enzymes can be targeted to the lysosome by either one of two pathways: a directintracellular route (“biosynthetic” pathway) or an endocytic pathway, with the former being the major pathway (Fig. 2). In the biosynthetic pathway, the formation of the active phosphomannosyl monoester on lysosomal enzymes occurs in thecis (early) Golgi compartment (1, 49). This raised the possibility that lysosomal enzymes might bind to theCI-MPR inthe early Golgi and either pass through the Golgi as a complex or exit the Golgi at this point. Indeed, data consistent with the latter possibility have come from immunocytochemical studies demonstrating that in some, but not all, cells the CI-MPR is concentrated in the cis Golgi with very low levels in the trans (late) Golgi (50). However, sorting in most cells has been postulated to occur in alate Golgi compartment, based on the following observations. A number of lysosomal enzymes have The CI-MPRand the IGF-11 Receptor Are the Same Proteinbeen shown to contain terminally processed oligosaccharides (51,52),

When Morgan et al. (11)cloned and sequenced the human insulinlike growth factor I1 (IGF-11) receptor, they made the surprising discovery that its sequence corresponds to that of the bovine CIMPR. The identity of the CI-MPR and theIGF-I1 receptor has been confirmed by biochemical studies which show that this protein can RER‘ bind phosphomannosyl residues and IGF-11, a nonglycosylated polyLysosome peptide hormone, simultaneously (31,32). However, each ligand may influence the binding of the other ligand to the receptor (13, 33, 34). The stoichiometry of IGF-I1 binding to the CI-MPR has been determined to be 1 mol of IGF-II/polypeptide chain (31). In contrast, the CD-MPR does not bind IGF-I1 (31,33). The biological significance of the finding that the CI-MPR and the IGF-I1 receptor are the same protein is still uncertain. The CI-MPR is known to bind and internalize IGF-I1 at the cell surface, resulting in the degradation of this ligand (35). In this manner it may serve to clear IGF-I1 from the circulation. One critical question is whether IGF-I1 binding to the CI-MPR results in signal transduction. This has been difficult to determine since IGF-I1 also binds to the IGF-I Endosome c“““”””receptor, a member of the tyrosine kinase family of receptors known to transmit signals across the plasma membrane. There are several reports suggesting that IGF-I1 mediates a response through its own Plasma Membrane receptor (5) including one in which IGF-I1 promoted cell proliferation FIG.2. Model for lysosomal enzyme targeting to lysosomes. Lysosomal in cells that lack the IGF-I receptor but contain the CI-MPR (36). In addition, IGF-I1 stimulated production of inositol trisphosphate in enzymes (0)and secretory proteins (0) are synthesized intheRERand membranes from renal proximal tubular cells, and this effect was transported to the Golgi where the lysosomal enzymes acquire phosphomanMost of the lysosomal enzymes bind to MPRs ( y ) in potentiated by Man-6-P (37). In these instancesthe CI-MPRmay be nosy1 residues (0-P). and/or early are translocated to late the trans Golgi network transducing a signal upon IGF-I1 binding, perhaps via coupling to a endosomes where they are (TGN) and due to the intraorganelle acidification. discharged pertussis toxin-sensitive G protein (38, 39). Another possibility is The enzymes are then packaged into lysosomes while the receptors cycle back that IGF-I1 may modulate the trafficking of lysosomal enzymes. This to theGolgi or to theplasma membrane. In addition to the pathways shown, it could be important for processes such as tissue and bone remodeling is likely that MPRs also cycle to the plasma membrane from the Golgi (via during development where it is necessary to secrete lysosomal en- leak into secretory vesicles) and from late endosomes.

J

Minireview: Man-6-P Receptors and indicating that these enzymes have traversed the entire Golgi complex since the glycosyltransferases responsible for terminal glycosylation reside in the trans Golgi elements (53). In addition, studies of the kinetics of receptor trafficking have demonstrated that the MPRs return to thelast Golgi compartment, the transGolgi network, much more frequently than they cycle to theearly Golgi compartments (30, 54). Furthermore, CI-MPRs and lysosomal enzymes have been localized to clathrin-coated vesicles in the region of the trans Golgi network (55,56). Takentogether, these data indicate that lysosomal enzymes are sorted from other classes of proteins in the trans Golgi network (7). Immunolocalization studies and biochemical analyses reveal very low or undetectable amounts of the CI-MPR in lysosomes while, in contrast, a significant amount of the receptor is found in endosomal structures (18, 55, 57-59). This has led to the concept that Golgiderived vesicles containing lysosomal enzyme-receptor complexes are delivered to acidic prelysosomal/endosomal compartments rather than to lysosomes (57-60). The low pH of the endosomal compartment would cause the complex to dissociate and the released lysosomal enzymes could be packaged into lysosomes while the CI-MPRs could recycle out of this compartment. (The variation in the pH of the different compartments assures the proper vectorial transport of ligand. The receptor binds ligand at neutral pH anddischarges ligand at acidic pH; the Golgi is near neutralitywhile the endosome is acidic. Consequently, the receptor will bind lysosomal enzymes in the Golgi and discharge them in endosomes.) Griffiths et al. (57) have described an acidic late endosomal structure in normal rat kidney cells which may serve as a site where lysosomal enzymes dissociate from the CIMPRs. This compartment is located in the vicinity of the Golgi complex, contains lysosomal enzymes as well as a lysosomal membrane protein (lgp120), and is enriched in CI-MPRs. This structure is distinct from the transGolgi network and early endosomes. Studies by Geuze et al. (58) also have provided evidence that the CI-MPRis segregated from lgp120 and presumably from lysosomal enzymes in prelysosomal/endosomal structures. However, it is currentlyunknown if the newly synthesized lysosomal enzymes are delivered to early endosomes, late endosomes, or both types of endosomes. The finding of CI-MPRs in early endosomes does not resolve this issue since these molecules may bederived from the plasma membrane. Extracellular lysosomal enzymes may also be delivered to the lysosome viathe endocytic pathway. A small proportion of lysosomal enzymes is typically secreted by cells (1, 2). Some of these enzymes may bind to cell surface CI-MPRs and be internalized via clathrincoated pits and vesicles (55, 61). The internalized acid hydrolases enter acidified endosomal compartments where they dissociate from the CI-MPRs and are delivered to lysosomes. Measurements of the number and half-life of MPRs and the rateof ligand internalization indicate that theMPRs are reutilized and can undergo many rounds of ligand delivery (62). In addition, studies using antibodies (26, 60, 63) or galactosyltransferase (30) to label receptors on the cell surface indicate that all of the MPRs in the cell are in rapid equilibrium. This suggests that thereis only one pool of receptor and thata single CI-MPR functions in both the endocytic and biosynthetic pathways (60).

Signals for Receptor Sorting and Endocytosis Onemodel for trafficking of the CI-MPR is that the receptor contains multiple signals that mediate its departure from and arrival a t three different destinations: the Golgi, plasma membrane, and endosomes. The cloning of the cDNA encoding the CI-MPR has provided a tool that is proving to be useful in the characterization of these signals. One approach that has been taken is to transfect CIMPR-deficient cells with either normal CI-MPR cDNA or cDNAs containing mutations in the cytoplasic domain and then assay for lysosomal enzyme sorting and endocytosis (29, 64). The normal receptor functioned well in the sorting of newly synthesized lysosomal enzymes and in the endocytosis of exogenous lysosomal enzymes (28, 29). Mutant receptors with either 40 or 89 residues deleted from the carboxyl terminus of the 163-amino acid cytoplasmic tail mediated endocytosis as well as the wild-type receptor but were significantly impaired in sorting (29).This result shows that one aspect of receptor trafficking (sorting) can be perturbed without, affecting the other aspect (endocytosis) and suggests that the signals for return to or departure from the Golgi are disrupted in these mutants. When the cytoplasmic tail was truncated to contain 7 or 20 residues, the resultant receptors accumulated on the cell surface and were thus defective in both endocytosis and sorting. These receptors are pre-

Lysosomal Enzyme Targeting

12117

sumably unable to enter clathrin-coated pits efficiently. A mutant receptor containing a cytoplasmic tail with alanine residues at positions 24 and 26 instead of the normal tyrosine residues was found to be defective in endocytosis, thereby demonstrating that these two tyrosine residues are required for rapid internalization of the CIMPR. The importance of tyrosine residues in the cytoplasmic tails of proteins involved in receptor-mediated endocytosis is emerging as a general theme (65). The first evidence for this came from studies on the LDL receptor (66). When a single tyrosine was changed to other charged or uncharged aliphatic residues, the resultant mutant LDL receptor was unable to enter coated pits efficiently. In contrast, substitution of the tyrosine with other aromatic amino acids resulted in a functional receptor, thus ruling out tyrosine phosphorylation as the signal for internalization. The introduction of a proline residue next to thetyrosine significantly decreased the efficiency of internalization, suggesting that the environment surrounding the tyrosine residue plays a role in the internalization of the LDL receptor. This is also corroborated by studies on a mutant influenza virus hemagglutinin. The introduction of a single tyrosine residue into the cytoplasmic tail of the hemagglutinin caused this protein, which is normally excluded from coated pits, to be rapidly internalized via clathrin-coated pits (67). The location of the tyrosine residue in this short (10-amino acid) cytoplasmic domain is critical since placement of tyrosines at two other positions did not result in rapid endocytosis of hemagglutinin. Inspection of the sequences of the recycling receptors in the region of their tyrosine residues reveals that while the surrounding residues are quite hydrophilic, no simple pattern of sequence identity is apparent. Further work is needed to define what constitutes the internalization signal. How may specificamino acid sequences in the cytoplasmic domain of the CI-MPR mediate its routing? One possibility is that cellular components recognize these signals and direct the receptor to its proper destination. Recent studies have begun to identify these components. Clathrin-coated pits in the plasma membrane contain a complex of 100.50.16-kDa polypeptides (65). These polypeptides, which have been termed “adaptors,” bind clathrin and also interact with endocytic receptors (68). Pearse and co-workers (68, 69) have shown that the plasma membrane-derived adaptor proteins interact with the cytoplasmic tails of the CI-MPR, LDL receptor, and the poly-Ig receptor, but not with a mutant CI-MPR cytoplasmic tail that lacks the two tyrosine residues a t positions 24 and 26. These results indicate that tyrosine residues in the cytoplasmic tails of these endocytic receptors are necessary for theirinteraction with the plasma membrane-derived adaptor proteins and suggest that this interaction is what mediates the entry of selected receptors into clathrin-coated pits.A different set of adaptor polypeptides (100.47. 19-kDa complex) has been isolated from clathrin-coated pits in the Golgi region (65). The Golgi-derived adaptor proteins bind to the cytoplasmic tail of the CI-MPR but not to that of the LDL receptor (68,69). Furthermore, the mutant CI-MPR tail that lacked tyrosines still interacted with the Golgi-derived adaptor proteins. Thus, both classes of adaptor proteins are likely to be involved in the routing of theCI-MPR, with one set directing departure from the plasma membrane and theother set directing departure from the Golgi. The identification of these two adaptor protein complexes, each of which is located at a unique site along the receptor’s targeting pathway and interacts with distinct signals on the receptor, provides a first step toward understanding the mechanism by which the CIMPR is routed. In addition, the selective recycling of the CI-MPR to the trans Golgi network has been reconstituted in vitro (70). The properties of receptor movement in this cell-free system indicate a vesicular transport mechanism. The ability to reconstitute in vitro this portion of the CI-MPRs intracellular pathway should aid in the identification of the cellular components involved in this targeting process. Furthermore, the recent isolation of endosomal structures enriched in MPRs should be useful in the characterization of other molecules involved in routing the receptors (71).

Potential Regulatorso f Receptor Trafficking Several studies have analyzed the effect of ligand binding on CIMPR trafficking. Alterations in the steady state distribution of the CI-MPR have been found in some (72,73) but not all (74, 75) studies where lysosomal enzyme synthesis is inhibited or where the dissociation of receptor-ligand complexes is prevented. On the other hand, studies of the kinetics of receptor movement under these conditions have shown that receptor occupancy does not have a significant effect

12118

Minireview: Man-6-P Receptors and Lysosomal

Enzyme Targeting

Nolan, C. M., Creek, K. E., Grubb, J. H., and Sly, W. S. (1987) J. Cell. Riochem. 127-151 .... .. . 35. _ ~ , " "Stein, M., Zijderhand-Bleekemolen,J. E., Geuze, H., Hasilik, A,, and von Figura, K. (1987) EMBOJ. 6 , 2677-2681 Kvle. J. W.. Nolan. C. M.. Oshima.. A,.. and Slv. " _W. S. (1988) J. Biol. Chem. -263, 16230;16235 Lobel. P.. Funmoto. K.. Ye. R. D.. Griffiths., G.., and Kornfeld., S. (1989) . . Celi57, 787-796 ' ' Duncan, J., and Kornfeld, S. (1988) J . Cell Bid. 106,617-628 Tong, P. Y., Tollefsen, S. E., and Kornfeld, S. (1988) J. Biol. Chem. 2 6 3 , 2585-2588 Waheed, A,, Braulke, T., Junghans, U., and von Figura, K. (1988) Biochem. Biophys. Res. Commun. 1 5 2 , 1248-1254 Kiess, W., Blickenstaff, G. D., Sklar, M. M., Thomas, C. L., Nissley, S. P., and Sahagian, G. G. (1988) J. Biol. Chem. 263,9339-9344 Kiess, W., Thomas, C. L., Greenstein, L. A., Lee, L., Sklar, M. M., Rechler, M. M., Sahagian, G. G., and Nissley, S. P. (1989) J. Biol. Chem. 264, 4710-4714 Oka, Y., Rozek, L. M., and Czech, M. P. (1985) J. Biol. Chem. 260,9435 Tally, M., Li, C. H., and Hall, K. (1987) Biochem. Biophys. Res. Commun. 148,811-816 Rogers, S. A., and Hammerman, M. R. (1989) J. Biol. Chem. 264,4273 Kojima, I., Nishimoto, I., Iiri, T., Ogata, E., and Rosenfeld, R. (1988) Biochem. Biophys. Res. Commun. 154,9-19 39. Braulke, T., Tippmer, S., Neher, E., and von Figura. K. (1989) EMBO J.

26. o n t h e r a t e of receptor movement (30, 54, 76). These latter results indicate that the CI-MPR shuttles constitutively between the cell 27. surface and intracellular compartments. However, the presence or 28. of receptor absence of ligand might inducesmall changes in the rates movement to various compartments, thereby affecting the steady 29. state distributionof receptor without having much of a n effect on the 30. overall rate of receptor movement. In some specialized cell types, 31. insulin and other growth factors induce a dramatic redistribution of t h e CI-MPR a n d t h e glucose transporter from endosomal compart32. m e n t s t o t h e cell surface (77, 78, a n d op. cit.). This redistribution is associated with a decreasein the phosphorylation state of the plasma 33. membrane-associated CI-MPRs (78). It is not known if this change 34. in phosphorylation is the cause or a consequence of the redistribution. This can now be tested directly by mutating the potential phosphor- 35. ylation sites on the receptor and determining if the response to growth 36. factors is maintained. 37. 38.

Conclusions

Thecloning of t h e MPRs hasprovidednewinsightintothe structure and functionof the receptors, yet many questions remain. 40. 41. W h a t is the purpose of having two distinct receptors for lysosomal enzyme trafficking? Are they redundant or do they function in the delivery of lysosomal enzymes to different compartments? Why does 42. 43. the same receptor bind lysosomal enzymes andIGF-II? Is lysosomal enzyme trafficking modulated byIGF-II? Does this receptor mediate 44. in the transport of signaltransduction,ordoesitonlyfunction to lysosomal enzymes,IGF-11, and perhaps other unidentified ligands t h e lysosome? W h a t a r e t h e s i g non a l sthe receptors that are involved45. 46. that direct the in routing, and what are the cellular components 47. receptor's movement? Studies involving the expression of mutated MPRs, the interaction of receptors with components of clathrin48. in uitro reconstitution of vesicular transport coated pits, and the all contributing to the under- 49. between the various compartments are 50. level. standing of this transport system at the molecular

REFERENCES 1. von Figura, K., and Hasilik, A. (1986) Annu. Reu. Biochem. 55,167-193 2. Kornfeld, S. (1987) FASEB J.1,462-468 3. Nolan, C. M., and Sly, W. S. (1987) Adu. Exp. Med. Biol. 2 2 5 , 199-212 4. Robbins, A. R. (1988) in Protein Transfer and OrganelleBiogenesrs,pp. 463-520, Academic Press, Orlando, FL 5. Roth, R. A. (1988) Science 239,1269-1271 6. Pfeffer, S. R. (1988) J. Membr. Biol. 103,7-16 7. Griffiths, G., and Simons, K. (1986) Science 234,438-443 8. Tong, P. Y., Gregory, W., and Kornfeld, S. (1989) J . Biol. Chem. 2 6 4 , 7962-7969 9. Tong, P.Y., and Kornfeld, S. (1989) J. Biol. Chem. 264,7970-7975 10. Lobel, P., Dahms, N. M., and Kornfeld, S. (1988)J. Biol. Chem. 263,25632570 11. Morgan, D. O., Edman, J. C., Standring, D. N., Fried, V. A,, Smith, M. C., Roth, R. A., and Rutter, W. J. (1987) Nature 329,301-307 12. Oshima. A,. Nolan. C. M.. Kvle. J. W.. Grubb.. J. H... and Slv. " . W. S. (1988) J. Biol. Chem. 263,2553-2562 13. MacDonald, R. G., Pfeffer, S. R., Coussens, L., Tepper, M. A,, Brocklehank, C. M.. Mole. J. E.. Anderson. J. K.. Cben.. E... Czech. M. P.. and Ullrich. A. (1988) S&nce 2 3 9 , 113411137 14. Dahms, N. M., Lobel, P., Breitmeyer, J., Chkgwin, J. M., and Kornfeld, S. (1987) Cell 5 0 , 181-192 15. Pohlmann, R., Nagel, G., Schmidt, B., Stein, M., Lorkowski, G., Krentler, '

16. 17. 18.

19. 20. 21. 22. 23. 24. 25.

C., Cully, J., Meyer, H. E., Grzeschik, K. H., Mersmann, G., Hasilik, A., and von Figura, K. (1987) Proc. Natl. Acad. Sci. U. S. A. 84,5575-5579 Sahagian, G. G., and Steer, C. J. (1985) J. Biol. Chem. 260,9838-9842 von Figura, K., Gieselmann, V., and Hasilik, A. (1985) Biochem. J . 225, 543-547 Sahagian, G. G., and Neufeld, E. F. (1983) J . Biol. Chem. 258,7121-7128 Corvera, S., and Czech,M. P. (1985) Proc. Natl. Acad.Sci. U. S. A. 8 2 , 7314-7318 Goldstein. J. L., Brown, M. S., Anderson, R. G. W., Russell, D. W., and Schneider, W. J. (1985) Annu. Reu. Ce:l Biol. 1 , 1-39 Stein, M., Braulke, T., Krentler, C., Hasilik, A,, and von Figura, K. (1987) Biol. Chem. Hoppe-Seyler 368,937-947 Dahms, N. M., and Kornfeld, S. (1989) J. Biol. Chem. 264,11458-11467 Stein, M., Meyer, H. E., Hasilik, A,, and von Figura, K. (1987) Biol. Chem. Hoppe-Seyler 368,927-936 Perdue, J. F., Chan. J. K., Thibault, C., Radaj, P., Mills, B., and Daughaday, W. H. (1983) J . Biol. Chem. 258,7800-7811 Gabel, C. A,, Goldberg, D. E., and Kornfeld, S. (1983) Proc. Natl. Acad. Sci. U. S. A. 80, 775-179

51. 52. 53. 54.

'

_"

8 . fi81-68fi _,"_

Canfield, W. M., and Kornfeld, S. (1989) J. Biol. Chem. 2 6 4 , 7100-7103 Rechler, M.M.,Zapf, J., Nissley, S. P., Froesch, E. R., Moses, A. C., Podskalnv. J. M.. Schillinz. E. E.. and Humbel. R. E. (1980) . . Endocrinology 107,-1451-1459 Sahagian, G. G., and Gottesman, M. M. (1982) J . B i d . Chem. 257,11145 Baumbach, G. A,, Saunders, P. T. K., Bazer, F. W., and Roberts, R. M. (1984) Proc. Natl. Acad. Scc. U. S. A. 81,2985-2989 Capony F. Morisset, M., Barrett, A. J., Capony, J. P., Broquet, P., Vignon, F., Ciambon, M., Louisot, P., andRochefort, H. (1987) J. Cell Biol. 1 0 4 , I .

3 "' v L I

I

-"-

C)fic)

Herzog, V., Neumuller, W., and Holzmann, B. (1987) EMBO J. 6,555-560 Lee, S.-J., and Nathans, D. (1988) J. Biol. Chem. 263,3521-3527 Kovacina, K. S., Steele-Perkins, G., Purchio, A. F., Lioubin, M., Miyazona, K., Heldin, C.-H., and Roth, R. A. (1989) Biochem. Biophys. Res. Commun. 160,393-403 Dong, J., Prence, E. M., and Sahagian, G. G. (1989) J. Biol. Chem. 2 6 4 , 7377-7383 Lazzarino, D. A., and Gabel, C. A. (1988) J . Biol. Chem. 263,10118-10126 Brown, W. J., and Farquhar, M. G. (1987) Proc. Nutl. Acad. Sci. U.S. A. 84,9001-9005 Vladutiu, G. D. (1983) Biochim. Biophys. Acta 760,363-370 Fedde, K. N., and Sly, W. S. (1985) Biochem. Biophys. Res. Commun. 1 3 3 , 614-620 Kornfeld, R., and Kornfeld, S. (1985) Annu. Rev. Biochem. 54,631-664 Jin, M., Sahagian, G. G., and Snider, M. D. (1989) J. Biol. Chem. 2 6 4 , 7fi7.5-7fiun

55. Geuze. H. J.. Slot. J. W.. Strous. G. J. A. M.. Hasilik.. A... and von Firmra, K. (1985) J . Cell Biol. 101.2253-2262 56. Lemansky, P., Hasilik, A,, von Figura, K., Helmy, S., Fishman, J., Fine, R. E., Kedersha, N. L., and Rome, L. H. (1987) J. Cell Biol. 1 0 4 , 1743 57. Griffiths. G.. Hoflack. B.. Simons.. K.., Mellman., I.., and Kornfeld. S. (1988) Cell 52,329-341 58. Geuze, H. J., Stoorvogel, W., Strous, G. J., Slot, J. W., Bleekemolen, J. E., and Mellman, I. (1988) J. Cell B~ol.1 0 7 , 2491-2501 59. Brown, W. J., Goodhouse, J., and Farquhar, M. G. (1986) J. Cell Biol. 1 0 3 , 1235-1247 60. Sahagian, G. G. (1984) Biol. Cell. 51,207-214 61. Willingham, M.C., Pastan, I. H. Sahagian, G. G., Jourdian, G. W., and Neufeld, E.F. (1981) Proc. Nati. Acad. Sci. U. S. A. 78,6867-6871 62. Gonzalez-Norleea. A.. Grubb. J. H., Talkad. V., and Sly, W. S. (1980) J. Cell Biol. 86,-839-852 63. Gartune. C.. Braulke., T.., Hasilik., A,.. and von Firmra. - . K. (1985) EMBO J. 4 , 1775-1730 64. Wntanahe. H.. Kvle. J. W.. Nolan. C. M.. and Slv. W. S. (1988) J . Cell Biol. 1 0 7 , 3i09(A&L 550) ' 65. Brodsky, F. M. (1988) Science 242,1396-1402 66. Davis, C. G., van Driel, I. R., Russell, D. W., Brown, M. S., and Goldstein, J. L. (1987) J. Biol. Chem. 262,4075-4082 67. Lazarovitis, J., and Roth, M. (1988) Cell 5 3 , 743-752 68. Pearse, B. M. F. (1988) EMBO J . 7,3331-3336 69. Glickman, J. N.. Conibear, E.. and Pearse, B. M. F. (1989) EMBO J. 8, 1041-1047 70. Goda, Y., and Pfeffer, S. R. (1988) Cell 55,309-320 71. Messner, D., Griffiths, G., and Kornfeld, S. (1989) J. Cell Biol., in press 72. Brown, W. J., and Farquhar, M. G. (1984) Proc. Natl. Acad. Sei. U. S. A. 81,5135-5139 73. Brown. W. J.. Constantinescu., E.,. and Farquhar. . . M. G. (1984) J . Cell Biol. 9 9 , 320-326 74. Pfeffer, S. R. (1987) J . Cell Biol. 1 0 5 , 229-234 15. Braulke, T., Gartung, C., Hasilik, A,, and von Figura, K. (1987) J. Cell Biol. 1 0 4 1735-1742 76. Oka, Y:, and Czech, M. P. (1986) J. Biol. Chem. 261,9090-9093 77. Appell, K. C., Simpson, I. A., and Cushman, S. W. (1988) J. Biol. Chem. 263,10824-10829 78. Corvera, S., Folander, K., Clairmont, K. B., and Czech, M. P. (1988) Proc. Natl. Acad. Sci. U. S. A. 8 5 , 7567-7571 ~

~~~

""

~I

~~~

~

~

I

I