Human
neuroblastoma GOTO cell lines were established that stably express recombinant human
lysosomal protective protein/
cathepsin A (
PPCA)
cDNA by transfection. Intracellular
cathepsin A (
acid serine carboxypeptidase) activity increased four-fold compared with in those of the parent and mock-transfected cell lines. The immunoreactive 54 kDa precursor/
zymogen and mature 32/20 kDa two-chain forms were produced in the cells. The amount of the latter form expressed in the GOTO cells was significantly larger than those in the
PPCA-overexpressing CHO cell lines previously established. The intracellular
proteins showed a typical lysosomal granular distribution and the glycosylated 54 kDa precursor was secreted into the culture medium without the addition of an alkalizing agent. The
PPCA-overexpressing cell lines also retained the ability to differentiate bi-directionally as well as the parent cells; into neuronal cells on induction by dibutyryl cAMP in serum-free medium and into Schwannian cells on induction by
bromodeoxyuridine. During the course of differentiation into neuronal and Schwannian cells, the intracellular
cathepsin A activity further increased two and five times, respectively, which was associated with an increase in the expression of the 32/20 kDa two-chain form. The glycosylated precursor
proteins were taken up via the
mannose 6-phosphate receptors, and the
cathepsin A, alpha-
neuraminidase and
beta-galactosidase (beta-Gal) activities deficient in the fibroblasts derived from a patient with
PPCA deficiency (
galactosialidosis) were restored. These results suggest that the bi-directional differentiation of GOTO cell lines stably expressing the recombinant human
PPCA gene could be a model system for analyzing the functions of
PPCA in peripheral neuronal cells and Schwannian cells as well as the recombinant
PPCA could be a useful source for
enzyme replacement therapy (ERT) for
galactosialidosis patients.