IMMORTALIZATION AND CHARACTERIZATION OF A CELL-LINE EXHIBITING A SEVERE MULTIPLE SULFATASE DEFICIENCY PHENOTYPE

Multiple sulphatase deficiency (MSD) is a rare genetic defect that cau ses a simultaneous deficiency of all known sulphatases, All available evidence suggests that the deficient gene product is normally responsi ble for the post-translational modification of a conserved cysteine re sidue to 2-amino-3-oxopropionic acid and that this modification is nec essary for sulphatase activity. MSD often has an enzymically mild phen otype, with significant levels of residual sulphatase activity being d etectable. Here we identify an MSD cell line in which the residual act ivity of the sulphatases assayed was generally very low. To characteri ze the phenotype of this cell line further, immortalized lines were es tablished after transformation with simian virus 40 (SV40) T antigen. Immortalized cell lines representing normal and MSD phenotypes were th en transduced with a retroviral vector carrying the gene encoding huma n N-acetylgalactosamine-4-sulphatase. Analysis of N-acetylgalactosamin e-4-sulphatase protein synthesis and enzyme activity showed that trans duced cell lines expressed large amounts of enzyme and that the specif ic activity of this enzyme was approx. 0.5-1.5 % of normal, confirming that this cell line defines a severe phenotype for MSD. N-Acetylgalac tosamine-4-sulphatase purified from a transduced MSD cell line seemed normal on denaturing PAGE. Kinetic analysis of the purified enzyme sug gests that the residual activity is due to small amounts of normal enz yme rather than unmodified enzyme with low levels of residual activity , These cell lines and the availability of large amounts of inactive N -acetylgalactosamine-4-sulphatase from MSD cells should facilitate the further study of this disorder.