Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1993 Sep 15;294(Pt 3):657–662. doi: 10.1042/bj2940657

Overexpression of N-acetylgalactosamine-4-sulphatase induces a multiple sulphatase deficiency in mucopolysaccharidosis-type-VI fibroblasts.

D S Anson 1, V Muller 1, J Bielicki 1, G S Harper 1, J J Hopwood 1
PMCID: PMC1134512  PMID: 8379921

Abstract

High-titre stocks of an amphotropic retrovirus, constructed so as to express a full-length cDNA encoding the human lysosomal enzyme N-acetylgalactosamine-4-sulphatase (4-sulphatase) from the cytomegalovirus immediate early promoter, were used to infect skin fibroblasts from a clinically severe mucopolysaccharidosis type VI (MPS VI) patient. The infected MPS VI cells showed correction of the enzymic defect with the enzyme being expressed at high levels and in the correct subcellular compartment. Surprisingly this did not result in correction of glycosaminoglycan turnover as measured by accumulation of 35S in metabolically labelled cells. We demonstrate that this is apparently caused by an induced reduction of the activities of other lysosomal sulphatases, presumably due to competition for a sulphatase-specific processing mechanism by the over-expressed 4-sulphatase. The level of steroid sulphatase, which is a microsomal sulphatase, was also reduced. Infection of skin fibroblasts from a second, clinically mildly affected, MPS VI patient with the same virus also resulted in no significant change in the level of glycosaminoglycan storage. However, in this case the cause of the observed phenomenon was less clear. These results are of obvious practical importance when considering gene therapy for a sulphatase deficiency such as MPS VI and also provide possible new avenues for exploration of the processes involved in sulphatase synthesis and genetically determined multiple sulphatase deficiency.

Full text

PDF
660

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anson D. S., Bielicki J., Hopwood J. J. Correction of mucopolysaccharidosis type I fibroblasts by retroviral-mediated transfer of the human alpha-L-iduronidase gene. Hum Gene Ther. 1992 Aug;3(4):371–379. doi: 10.1089/hum.1992.3.4-371. [DOI] [PubMed] [Google Scholar]
  2. Bielicki J., Freeman C., Clements P. R., Hopwood J. J. Human liver iduronate-2-sulphatase. Purification, characterization and catalytic properties. Biochem J. 1990 Oct 1;271(1):75–86. doi: 10.1042/bj2710075. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Brooks D. A., Gibson G. J., McCourt P. A., Hopwood J. J. A specific fluorogenic assay for N-acetylgalactosamine-4-sulphatase activity using immunoadsorption. J Inherit Metab Dis. 1991;14(1):5–12. doi: 10.1007/BF01804381. [DOI] [PubMed] [Google Scholar]
  4. Danos O., Mulligan R. C. Safe and efficient generation of recombinant retroviruses with amphotropic and ecotropic host ranges. Proc Natl Acad Sci U S A. 1988 Sep;85(17):6460–6464. doi: 10.1073/pnas.85.17.6460. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Eto Y., Wiesmann U. N., Carson J. H., Herschkowitz N. N. Multiple sulfatase deficiencies in cultured skin fibroblasts. Occurrence in patients with a variant form of metachromatic leukodystrophy. Arch Neurol. 1974 Feb;30(2):153–156. doi: 10.1001/archneur.1974.00490320041005. [DOI] [PubMed] [Google Scholar]
  6. Freeman C., Hopwood J. J. Sanfilippo D syndrome: estimation of N-acetylglucosamine-6-sulfatase activity with a radiolabeled monosulfated disaccharide substrate. Anal Biochem. 1989 Feb 1;176(2):244–248. doi: 10.1016/0003-2697(89)90303-5. [DOI] [PubMed] [Google Scholar]
  7. Gibson G. J., Saccone G. T., Brooks D. A., Clements P. R., Hopwood J. J. Human N-acetylgalactosamine-4-sulphate sulphatase. Purification, monoclonal antibody production and native and subunit Mr values. Biochem J. 1987 Dec 15;248(3):755–764. doi: 10.1042/bj2480755. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Glaser J. H., Sly W. S. Beta-glucuronidase deficiency mucopolysaccharidosis: methods for enzymatic diagnosis. J Lab Clin Med. 1973 Dec;82(6):969–977. [PubMed] [Google Scholar]
  9. Hopwood J. J., Elliott H. Detection of Morquio A syndrome using radiolabelled substrates derived from keratan sulphate for the estimation of galactose 6-sulphate sulphatase. Clin Sci (Lond) 1983 Sep;65(3):325–331. doi: 10.1042/cs0650325. [DOI] [PubMed] [Google Scholar]
  10. Hopwood J. J., Elliott H. Diagnosis of Sanfilippo type A syndrome by estimation of sulfamidase activity using a radiolabelled tetrasaccharide substrate. Clin Chim Acta. 1982 Aug 18;123(3):241–250. doi: 10.1016/0009-8981(82)90168-1. [DOI] [PubMed] [Google Scholar]
  11. Hopwood J. J., Elliott H., Muller V. J., Saccone G. T. Diagnosis of Maroteaux-Lamy syndrome by the use of radiolabelled oligosaccharides as substrates for the determination of arylsulphatase B activity. Biochem J. 1986 Mar 15;234(3):507–514. doi: 10.1042/bj2340507. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jezyk P. F., Haskins M. E., Patterson D. F., Mellman W. J., Greenstein M. Mucopolysaccharidosis in a cat with arylsulfatase B deficiency: a model of Maroteaux-Lamy syndrome. Science. 1977 Nov 25;198(4319):834–836. doi: 10.1126/science.144321. [DOI] [PubMed] [Google Scholar]
  13. Kolodny E. H., Mumford R. A. Human leukocyte acid hydrolases: characterization of eleven lysosomal enzymes and study of reaction conditions for their automated analysis. Clin Chim Acta. 1976 Jul 15;70(2):247–257. doi: 10.1016/0009-8981(76)90426-5. [DOI] [PubMed] [Google Scholar]
  14. LEABACK D. H., WALKER P. G. Studies on glucosaminidase. 4. The fluorimetric assay of N-acetyl-beta-glucosaminidase. Biochem J. 1961 Jan;78:151–156. doi: 10.1042/bj0780151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  16. Miller A. D., Buttimore C. Redesign of retrovirus packaging cell lines to avoid recombination leading to helper virus production. Mol Cell Biol. 1986 Aug;6(8):2895–2902. doi: 10.1128/mcb.6.8.2895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Miller A. D., Rosman G. J. Improved retroviral vectors for gene transfer and expression. Biotechniques. 1989 Oct;7(9):980-2, 984-6, 989-90. [PMC free article] [PubMed] [Google Scholar]
  18. Peters C., Rommerskirch W., Modaressi S., von Figura K. Restoration of arylsulphatase B activity in human mucopolysaccharidosis-type-VI fibroblasts by retroviral-vector-mediated gene transfer. Biochem J. 1991 Jun 1;276(Pt 2):499–504. doi: 10.1042/bj2760499. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Rommerskirch W., von Figura K. Multiple sulfatase deficiency: catalytically inactive sulfatases are expressed from retrovirally introduced sulfatase cDNAs. Proc Natl Acad Sci U S A. 1992 Apr 1;89(7):2561–2565. doi: 10.1073/pnas.89.7.2561. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Steckel F., Hasilik A., von Figura K. Synthesis and stability of arylsulfatase A and B in fibroblasts from multiple sulfatase deficiency. Eur J Biochem. 1985 Aug 15;151(1):141–145. doi: 10.1111/j.1432-1033.1985.tb09078.x. [DOI] [PubMed] [Google Scholar]
  21. Taylor J. A., Gibson G. J., Brooks D. A., Hopwood J. J. Human N-acetylgalactosamine-4-sulphatase biosynthesis and maturation in normal, Maroteaux-Lamy and multiple-sulphatase-deficient fibroblasts. Biochem J. 1990 Jun 1;268(2):379–386. doi: 10.1042/bj2680379. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wasteson A. A method for the determination of the molecular weight and molecular-weight distribution of chondroitin sulphate. J Chromatogr. 1971 Jul 8;59(1):87–97. doi: 10.1016/s0021-9673(01)80009-1. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES