Biosynthesis and expression of polysialic acid on the neural cell adhesion molecule is predominantly directed by ST8Sia II/STX during in vitro neuronal differentiation

We have reported recently that ST8Sia II/STX as well as ST8Sia IV/PST-1 is a neural cell, adhesion molecule (NCAM)-specific polysialic acid (PSA) synthase (Kojima, N., Tachida, Y., Yoshida, Y., and Tsuji, S. (1996) J. Biol. Chem. 271, 19457-19463). To investigate which of two PSA synthase (ST8Sia II and IV) are involved in the biosynthesis of PSA associated with NCAM, the expressions of PSA, PSA synthase activity, and the genes of two PSA synthases during in vitro neuronal differentiation of mouse embryonal carcinoma P19 cells were determined. PSA was not expressed on undifferentiated cells (day 0) or cell aggregates (days 1-3) induced with retinoic acid. Expression of PSA began after cell aggregates had been dissociated and re-plated on a dish (day 4) and increased up to day 7. The expression of the mouse ST8Sia II gene was negligible in both undifferentiated and aggregated cells, it beginning at day 4 then dramatically increasing, and reaching the maximum level at days 6-7. On the other hand, transcription of the ST8Sia IV gene remained at a very low level throughout the entire period, a significant increase in its expression during differentiation not being observed. PSA synthase activity was not detected in undifferentiated or aggregated P19 cells, it increasing in parallel with ST8Sia II gene expression during differentiation. In addition, the cells at day 7 were stained with an anti-mouse ST8Sia II antiserum. Similar up-regulation of the ST8Sia II gene were observed during the differentiation of rat MNS-8 cells, which were derived from E-12 rat neuroepithelium of the neural tube and shown to differentiate into neurons. These results indicate that ST8Sia II predominantly directs PSA expression during neuronal differentiation rather than ST8Sia IV.