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Biochemical Genetics

, Volume 25, Issue 1–2, pp 161–174 | Cite as

Abnormal subcellular distribution of β-glucuronidase in mice with a genetic alteration in enzyme structure

  • Richard T. Swank
  • Karen Moore
  • Verne M. Chapman
Article

Abstract

Liver β-glucuronidase is structurally altered in inbred strain PAC so that a peptide subunit with a more basic isoelectric point, GUS-SN, is produced. This allele of β-glucuronidase was transferred to strain C57BL/6J by 12 backcross matings to form the congenic line B6 · PAC-Gusn. Liver β-glucuronidase activity was halved in males of the congenic strain compared to normal males. The lowered activity was specifically accounted for by a decrease in the lysosomal component. There was no alteration in the concentration of microsomal activity. This alteration in the subcellular distribution of β-glucuronidase in Gusn/Gusn mice was confirmed by two independent gel electrophoretic systems which separate microsomal and lysosomal components. β-Glucuronidase activity was likewise approximately halved in mutant spleen, lung, and brain, organs which contain exclusively or predominantly lysosomal β-glucuronidase. The loss of liver lysosomal β-glucuronidase activity was shown by immunotitration to be due to a decrease in the number of β-glucuronidase molecules in lysosomes of the congenic strain. The Gusn structural alteration likely causes the lowered lysosomal β-glucuronidase activity since the two traits remain in congenic animals. Heterozygous Gusn/Gusb animals had intermediate levels of liver β-glucuronidase. Also, the effect was specific, in that three other lysosomal enzymes were not reproducibly lower in Gusn/Gusn mice. Gusn is, therefore, an unusual example of a mutation which causes a change in the subcellular distribution of a two-site enzyme.

Key words

β-glucuronidase lysosomal microsomal subcellular congenic 

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References

  1. Adams, G. A., and Rose, J. K. (1985). Membrane-spanning domain blocks cell surface transport but not membrane anchoring of a viral glycoprotein. Mol. Cell. Biol. 51442.Google Scholar
  2. Alt, F. W., Bothwell, A. L. M., Knapp, M., Siden, E., Mather, E., Koshland, M., and Baltimore, D. (1980). Synthesis of secreted and membrane-bound immunoglobulin Mu heavy chains is directed by mRNAs that differ at their 3′ ends. Cell 20293.Google Scholar
  3. Brandt, E. J., Elliott, R. W., and Swank, R. T. (1975). Defective lysosomal secretion in kidneys of Chediak-Higashi (beige) mice. J. Cell Biol. 67774.Google Scholar
  4. Brown, J. A., Jahreis, G. P., and Swank, R. T. (1981). The synthesis and processing of β-glucuronidase in normal and egasyn deficient mouse kidney. Biochem. Biophys. Res. Commun. 99691.Google Scholar
  5. De Pierre, J. W., and Ernster, L. (1977). Enzyme topology of intracellular membranes. In Snell, E. (ed.), Annual Reviews of Biochemistry, Vol. 46 Annual Reviews, Inc., Palo Alto, Calif., pp. 201–262.Google Scholar
  6. De Robertis, E. M. (1983). Nucleocytoplasmic segregation of proteins and RNAs. Cell 321021.Google Scholar
  7. Ganschow, R. E. (1975). Simultaneous genetic control of the structure and rate of synthesis of murine glucuronidase. In Markert, C. (ed.), Isozymes IV, Genetics and Evolution, Vol. 4 Academic Press, New York, pp. 633–647.Google Scholar
  8. Ganschow, R., and Paigen, K. (1967). Separate genes determining the structure and intracellular location of hepatic glucuronidase. Proc. Natl. Acad. Sci. USA 58938.Google Scholar
  9. Hayashi, M., Nakajima, Y., and Fishman, W. H. (1964). The cytologic demonstration of β-glucuronidase employing naphthol AS-BI glucuronide and hexazonium pararosaniline. J. Histochem. Cytochem. 12293.Google Scholar
  10. Kalderon, D., Robert, B. L., Richardson, W. D., and Smith, A. E. (1984). A short amino acid sequence able to specify nuclear location. Cell 39499.Google Scholar
  11. Larusso, N. F., and Fowler, S. (1979). Coordinate secretion of acid hydrolases in rat bile. J. Clin. Invest. 64948.Google Scholar
  12. Lodish, M. F., Brall, W. A., Schwartz, A. L., Strous, G. J. A. M., and Zilberstein, A. (1981). Synthesis and assembly of membrane and organelle proteins. Int. Rev. Cytol. Suppl. 12247.Google Scholar
  13. Lusis, A. J., and Paigen, K. (1977). Mechanisms involved in the intracellular localization of mouse glucuronidase. In Ratazzi, M. C., Scandalius, J. G., and Whitt, G. S., (eds.), Isozymes: Current Topics in Biological and Medical Research, Vol. 2 Alan R. Liss, New York, pp. 63–106.Google Scholar
  14. Lusis, A. J., and Paigen, K. (1978). The large scale isolation of mouse β-glucuronidase and comparison of allozymes. J. Biol. Chem. 2537336.Google Scholar
  15. Lusis, A. J., and Swank, R. T. (1980). Regulation of location of intracellular proteins. In Goldstein, L., and Prescott, D. M. (eds.), Cell Biology: A Comprehensive Treatise, Vol. 4 Academic Press, New York, pp. 339–391.Google Scholar
  16. Masters, C. J. (1981). Interactions between soluble enzymes and subcellular structures. In Fasman, G. D. (ed.), Critical Reviews in Biochemistry, Vol. 11, pp. 105–143.Google Scholar
  17. Medda, S., and Swank, R. T. (1985). Egasyn a protein which determines the subcellular distribution of β-glucuronidase, has esterase activity. J. Biol. Chem. 26015802.Google Scholar
  18. Novak, E. K., and Swank, R. T. (1979). Lysosomal dysfunctions associated with mutations at mouse pigment genes. Genetics 92189.Google Scholar
  19. Paigen, K. (1961). The effect of mutation on the intracellular location of β-glucuronidase. Exp. Cell Res. 25286.Google Scholar
  20. Rogers, J., Early, P., Carter, C., Calame, K., Bord, M., Hood, L., and Wall, R. (1980). Two mRNAs with different 3′ ends encode membrane-bound and secreted forms of immunoglobulin μ chain. Cell 20303.Google Scholar
  21. Smith, K., and Ganschow, R. E. (1978). Turnover of murine β-glucuronidase. J. Biol. Chem. 2535437.Google Scholar
  22. Swank, R.T., and Paigen, K. (1973). Biochemical and genetic evidence for a macromolecular β-glucuronidase complex in microsomal membranes. J. Mol. Biol. 77371.Google Scholar
  23. Swank, R. T., Pfister, K., Miller, D., and Chapman, V. (1986). The egasyn gene affects the processing of oligosaccharides of lysosomal β-glucuronidase in liver. Biochem. J. 240445.Google Scholar
  24. Tomino, S., Paigen, K., Tulsiani, D. R. P., and Touster, O. (1975). Purification and chemical properties of mouse liver lysosomal (L form) β-glucuronidase. J. Biol. Chem. 2508503.Google Scholar
  25. Wood, S. (1975). A sensitive fluorometric assay for alpha-L-fucosidase. Clin. Chem. Acta 58251.Google Scholar

Copyright information

© Plenum Publishing Corporation 1987

Authors and Affiliations

  • Richard T. Swank
    • 1
  • Karen Moore
    • 2
  • Verne M. Chapman
    • 1
  1. 1.Department of Molecular BiologyRoswell Park Memorial InstituteBuffalo
  2. 2.Frederick Cancer Research FacilityFrederick

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