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Volume 154, Issue 12

Kex2 protease converts the endoplasmic reticulum 1,2-mannosidase of into a soluble cytosolic form Free

Abstract

Cytosolic -mannosidases are glycosyl hydrolases that participate in the catabolism of cytosolic free -oligosaccharides. Two soluble -mannosidases (E-I and E-II) belonging to glycosyl hydrolases family 47 have been described in . We demonstrate that addition of pepstatin A during the preparation of cell homogenates enriched -mannosidase E-I at the expense of E-II, indicating that the latter is generated by proteolysis during cell disruption. E-I corresponded to a polypeptide of 52 kDa that was associated with mannosidase activity and was recognized by an anti-1,2-mannosidase antibody. The -mannan core trimming properties of the purified enzyme E-I were consistent with its classification as a family 47 1,2-mannosidase. Differential density-gradient centrifugation of homogenates revealed that 1,2-mannosidase E-I was localized to the cytosolic fraction and Golgi-derived vesicles, and that a 65 kDa membrane-bound 1,2-mannosidase was present in endoplasmic reticulum and Golgi-derived vesicles. Distribution of -mannosidase activity in a Δ null mutant or in wild-type protoplasts treated with monensin demonstrated that the membrane-bound 1,2-mannosidase is processed by Kex2 protease into E-I, recognizing an atypical cleavage site of the precursor. Analysis of cytosolic free -oligosaccharides revealed that cytosolic 1,2-mannosidase E-I trims free ManGlcNAc isomer B into ManGlcNAc isomer B. This is believed to be the first report demonstrating the presence of soluble 1,2-mannosidase from the glycosyl hydrolases family 47 in a cytosolic compartment of the cell.

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Keyword(s): 1-DMJ, 1-deoxymannojirimycin , AEBSF, 4-(2-aminoethyl)benzenesulfonyl fluoride , CPY, carboxypeptidase Y , E64, trans-epoxysuccinyl-l-leucyl-amido(4-guanidino)butane , ER, endoplasmic reticulum , ERAD, endoplasmic-reticulum-associated degradation , M7B, Man7GlcNAc2 isomer B , M8B, Man8GlcNAc2 isomer B , M9, Man9GlcNAc2 , MU, 4-methylumbelliferone and MUαMan, 4-methylumbelliferyl-α-d-mannopyranoside
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2008-12-01
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References

  1. Arnaud M. B., Costanzo M. C., Skrzypek M. S., Binkley G., Lane C., Miyasato S. R., Sherlock G. 2005; The Candida Genome Database (CGD), a community resource for Candida albicans gene and protein information. Nucleic Acids Res 33:D358–D363
     [Google Scholar]
  2. Bader O., Schaller M., Klein S., Kukula J., Haack K., Mühlschlegel F., Korting H. C., Schäfer, W., Hube B. 2001; The KEX2 gene of Candida glabrata is required for cell surface integrity. Mol Microbiol 41:1431–1444
     [Google Scholar]
  3. Beaufay H., Amar-Costesec A., Thines-Sempoux D., Wibo M., Robbi M., Berthet J. 1974; Analytical study of microsomes and isolated subcellular membranes from rat liver. J Cell Biol 61:213–231
     [Google Scholar]
  4. Bevan A., Brenner C., Fuller R. S. 1998; Quantitative assessment of enzyme specificity in vivo: P2 recognition by Kex2 protease defined in a genetic system. Proc Natl Acad Sci U S A 95:10384–10389
     [Google Scholar]
  5. Bischoff J., Kornfeld R. 1986; The soluble form of rat liver alpha-mannosidase is immunologically related to the endoplasmic reticulum membrane alpha-mannosidase. J Biol Chem 261:4758–4765
     [Google Scholar]
  6. Bischoff J., Moremen K. W., Lodish H. F. 1990; Isolation, characterization, and expression of cDNA encoding a rat liver endoplasmic reticulum alpha-mannosidase. J Biol Chem 265:17110–17117
     [Google Scholar]
  7. Bloom G. S., Brashear T. A. 1989; A novel 58-kDa protein associates with the Golgi apparatus and microtubules. J Biol Chem 264:16083–16092
     [Google Scholar]
  8. Bradford M. M. 1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254
     [Google Scholar]
  9. Brand A., MacCallum D. M., Brown A. J. P., Gow N. A. R., Odds F. C. 2004; Ectopic expression of URA3 can influence the virulence phenotypes and proteome of Candida albicans but can be overcome by targeted reintegration of URA3 at the RPS10 locus. Eukaryot Cell 3:900–909
     [Google Scholar]
  10. Brenner C., Fuller R. S. 1992; Structural and enzymatic characterization of a purified prohormone-processing enzyme: secreted, soluble Kex2 protease. Proc Natl Acad Sci U S A 89:922–926
     [Google Scholar]
  11. Bryant N. J., Stevens T. H. 1998; Vacuole biogenesis in Saccharomyces cerevisiae: protein transport pathways to the yeast vacuole. Microbiol Mol Biol Rev 62:230–247
     [Google Scholar]
  12. Chantret I., Frenoy J. P., Moore S. E. 2003; Free-oligosaccharide control in the yeast Saccharomyces cerevisiae: roles for peptide :  N-glycanase (Png1p) and vacuolar mannosidase (Ams1p. Biochem J 373:901–908
     [Google Scholar]
  13. Chrispeels M. J. 1983; The Golgi apparatus mediates the transport of phytohemagglutinin to the protein bodies in bean cotyledons. Planta 158:140–151
     [Google Scholar]
  14. Chrispeels M. J., Higgins T. J., Craig S., Spencer D. 1982; Role of the endoplasmic reticulum in the synthesis of reserve proteins and the kinetics of their transport to protein bodies in developing pea cotyledons. J Cell Biol 93:5–14
     [Google Scholar]
  15. Costanzi E., Balducci C., Cacan R., Duvet S., Orlacchio A., Beccari T. 2006; Cloning and expression of mouse cytosolic α-mannosidase (Man2c1. Biochim Biophys Acta 17601580–1586
     [Google Scholar]
  16. Daniel P. F., Winchester B., Warren C. D. 1994; Mammalian α-mannosidases – multiple forms but a common purpose?. Glycobiology 4:551–566
     [Google Scholar]
  17. De Gasperi R., Al Daher S., Winchester B. G., Warren C. D. 1992; Substrate specificity of the bovine and feline neutral alpha-mannosidases. Biochem J 286:47–53
     [Google Scholar]
  18. Dutta P., Majumder G. C. 1984; Enzymic characteristics of the isoenzymes of rat epididymal neutral alpha-mannosidases and their changes during development in vivo . Biochem J 218:489–494
     [Google Scholar]
  19. Fonzi W. A., Irwin M. Y. 1993; Isogenic strain construction and gene mapping in Candida albicans . Genetics 134:717–728
     [Google Scholar]
  20. Fuller R. S., Sterne R. E., Thorner J. 1988; Enzymes required for yeast prohormone processing. Annu Rev Physiol 50:345–362
     [Google Scholar]
  21. Fuller R. S., Brake A., Thorner J. 1989; Yeast prohormone processing enzyme ( KEX2 gene product) is a Ca2+-dependent serine protease. Proc Natl Acad Sci U S A 86:1434–1438
     [Google Scholar]
  22. Gillum A. M., Tsay E. Y. H., Kirsch D. R. 1984; Isolation of the Candida albicans gene for orotidine-5′-phosphate decarboxylase by complementation of S. cerevisiae ura3 and E. coli pyrF mutations. Mol Gen Genet 198:179–182
     [Google Scholar]
  23. Grard T., Saint-Pol A., Haeuw J., Alonso C., Wieruszeski J., Strecker G., Michalski J. 1994; Soluble forms of α-d-mannosidases from rat liver. Eur J Biochem 223:99–106
     [Google Scholar]
  24. Grard T., Herman V., Saint-Pol A., Kmiecik D., Labiau O., Mir A. M., Alonso C., Verbert A., Cacan R., Michalski J. C. 1996; Oligomannosides or oligosaccharide-lipids as potential substrates for rat liver cytosolic alpha-d-mannosidase. Biochem J 316:787–792
     [Google Scholar]
  25. Haeuw J. F., Strecker G., Wieruszeski J. M., Montreuil J., Michalski J. C. 1991; Substrate specificity of rat liver cytosolic alpha-d-mannosidase. Novel degradative pathway for oligomannoside type glycans. Eur J Biochem 202:1257–1268
     [Google Scholar]
  26. Harris S. L., Waters M. G. 1996; Localization of a yeast early Golgi mannosyltransferase, Och1p, involves retrograde transport. J Cell Biol 132:985–998
     [Google Scholar]
  27. Helenius A., Aebi M. 2004; Roles of N-linked glycans in the endoplasmic reticulum. Annu Rev Biochem 73:1019–1049
     [Google Scholar]
  28. Henrissat B., Davis G. 1997; Structural and sequence-based classification of glycoside hydrolases. Curr Opin Struct Biol 7:637–644
     [Google Scholar]
  29. Herscovics A. 1999a; Processing glycosidases of Saccharomyces cerevisiae . Biochim Biophys Acta 1426:275–285
     [Google Scholar]
  30. Herscovics A. 1999b; Importance of glycosidases in mammalian glycoprotein biosynthesis. Biochim Biophys Acta 1473:96–107
     [Google Scholar]
  31. Huh W. K., Falvo J. V., Gerke L. C., Carroll A. S., Howson R. W., Weissman J. S., O'Shea E. K. 2003; Global analysis of protein localization in budding yeast. Nature 425:686–691
     [Google Scholar]
  32. Jakob C. A., Bodmer D., Spirig U., Battig P., Marcil A., Dignard D., Bergeron J. J., Thomas D. Y., Aebi M. 2001; Htm1p, a mannosidase-like protein, is involved in glycoprotein degradation in yeast. EMBO Rep 2:423–430
     [Google Scholar]
  33. Jelinek-Kelly S., Herscovics A. 1988; Glycoprotein biosynthesis in Saccharomyces cerevisiae: purification of the α-mannosidase which removes one specific mannose residue from Man9GlcNAc. J Biol Chem 263:14757–14763
     [Google Scholar]
  34. Laemmli U. K. 1970; Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685
     [Google Scholar]
  35. Lesage G., Tremblay M., Guimond J., Boileau G. 2001; Mechanism of Kex2p inhibition by its proregion. FEBS Lett 508:332–336
     [Google Scholar]
  36. Massaad M. J., Herscovics A. 2001; Interaction of the endoplasmic reticulum alpha 1,2-mannosidase Mns1p with Rer1p using the split-ubiquitin system. J Cell Sci 114:4629–4635
     [Google Scholar]
  37. Merril C. R. 1990; Gel-staining techniques. In Guide to Protein Purification pp 477–488 Edited by Deutscher M. P. San Diego, CA: Academic Press;
     [Google Scholar]
  38. Mora-Montes H. M., López-Romero E., Zinker S., Ponce-Noyola P., Flores-Carreón A. 2004; Hydrolysis of Man9GlcNAc2 and Man8GlcNAc2 oligosaccharides by a purified α-mannosidase from Candida albicans . Glycobiology 14:593–598
     [Google Scholar]
  39. Mora-Montes H. M., López-Romero E., Zinker S., Ponce-Noyola P., Flores-Carreón A. 2006; Purification of soluble α1,2-mannosidase from Candida albicans CAI-4. FEMS Microbiol Lett 256:50–56
     [Google Scholar]
  40. Mora-Montes H. M., Bates S., Netea M. G., Díaz-Jiménez D. F., López-Romero E., Zinker S., Ponce-Noyola P., Kullberg B. J., Brown A. J. other authors 2007; Endoplasmic reticulum alpha-glycosidases of Candida albicans are required for N-glycosylation, cell wall integrity, and normal host–fungus interaction. Eukaryot Cell 6:2184–2193
     [Google Scholar]
  41. Mukhtar M., Logan D. A., Kaufer N. F. 1992; The carboxypeptidase Y-encoding gene from Candida albicans and its transcription during yeast-to-hyphae conversion. Gene 121:173–177
     [Google Scholar]
  42. Nakatsukasa K., Nishikawa S., Hosokawa N., Nagata K., Endo T. 2001; Mnl1p, an alpha-mannosidase-like protein in yeast Saccharomyces cerevisiae, is required for endoplasmic reticulum-associated degradation of glycoproteins. J Biol Chem 276:8635–8638
     [Google Scholar]
  43. Newport G., Kuo A., Flattery A., Gill C., Blake J. J., Kurtz M. B., Abruzzo G. K., Agabian N. 2003; Inactivation of Kex2p diminishes the virulence of Candida albicans . J Biol Chem 278:1713–1720
     [Google Scholar]
  44. Parlati F., Dominguez M., Bergeron J. J. M., Thomas D. Y. 1995; Saccharomyces cerevisiae CNE1 encodes an endoplasmic reticulum (ER) membrane protein with sequence similarity to calnexin and calreticulin and functions as a constituent of the ER quality control apparatus. J Biol Chem 270:244–253
     [Google Scholar]
  45. Ramírez M., Hernández L. M., Larriba G. 1989; A similar protein portion for two exoglucanases secreted by Saccharomyces cerevisiae . Arch Microbiol 151:391–398
     [Google Scholar]
  46. Rosa P., Mantovani S., Rosboch R., Huttner W. B. 1992; Monensin and brefeldin A differentially affect the phosphorylation and sulfation of secretory proteins. J Biol Chem 267:12227–12232
     [Google Scholar]
  47. Spiro R. G. 2004; Role of N-linked polymannose oligosaccharides in targeting glycoproteins for endoplasmic reticulum-associated degradation. Cell Mol Life Sci 61:1025–1041
     [Google Scholar]
  48. Suzuki T., Hara I., Nakano M., Shigeta M., Nakagawa T., Kondo A., Funakoshi Y., Taniguchi N. 2006; Man2C1, an α-mannosidase, is involved in the trimming of free oligosaccharides in the cytosol. Biochem J 400:33–41
     [Google Scholar]
  49. Tremblay L. O., Herscovics A. 2000; Characterization of a cDNA encoding a novel human Golgi α1,2-mannosidase (IC) involved in N-glycan biosynthesis. J Biol Chem 275:31655–31660
     [Google Scholar]
  50. Tulsiani D. R. P., Touster O. 1987; Substrate specificities of rat kidney lysosomal and cytosolic α-D-mannosidases and effects of swainsonine suggest a role of the cytosolic enzyme in glycoprotein catabolism. J Biol Chem 262:6506–6514
     [Google Scholar]
  51. Vázquez-Reyna A. B., Ponce-Noyola P., Calvo-Méndez C., López-Romero E., Flores-Carreón A. 1999; Purification and biochemical characterization of two soluble α-mannosidases from Candida albicans . Glycobiology 9:533–537
     [Google Scholar]
  52. Weng S., Spiro R. G. 1996; Endoplasmic reticulum kifunensine-resistant alpha-mannosidase is enzymatically and immunologically related to the cytosolic alpha-mannosidase. Arch Biochem Biophys 325:113–123
     [Google Scholar]
  53. Wu Y., Termine D. J., Swulius M. T., Moremen K. W., Sifers R. N. 2007; Human endoplasmic reticulum mannosidase I is subject to regulated proteolysis. J Biol Chem 282:4841–4849
     [Google Scholar]
  54. Yoshihisa T., Anraku Y. 1990; A novel pathway of import of alpha-mannosidase, a marker enzyme of vacuolar membrane, in Saccharomyces cerevisiae . J Biol Chem 265:22418–22425
     [Google Scholar]
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Kex2 protease converts the endoplasmic reticulum α1,2-mannosidase of Candida albicans into a soluble cytosolic form
Microbiology 154, 3782 (2008); https://doi.org/10.1099/mic.0.2008/019315-0
/content/journal/micro/10.1099/mic.0.2008/019315-0
/content/journal/micro/10.1099/mic.0.2008/019315-0
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