Neuroblastomas and cell lines derived from these tumors bear the oncodevelopmental antigen polysialic acid (PSA) bound to the neural cell adhesion molecule. Polysialyation of neural cell adhesion molecule can be achieved by two different polysialyltransferases, ST8SiaII and ST8SiaIV. This study was undertaken to investigate the pattern of polysialyltransferases expressed in the human neuroblastoma cell line SH-SY5Y. Reverse transcription-PCR showed simultaneous expression of the two enzymes, and in situ hybridization demonstrated that the polysialyltransferase mRNA expression parallels immunoreactivity with the PSA-specific monoclonal antibody 735. After retinoic acid-induced differentiation, only the PSA-positive, neuron-like cell type gave clear signals for ST8SiaII and ST8SiaIV in in situ hybridization, whereas both signals were drastically reduced in the weakly PSA-positive substrate adherent phenotype. Like the SH-SY5Y cells, a primary, PSA-positive neuroblastoma specimen revealed expression of the two polysialyltransferases. To investigate the role of PSA for cell growth and differentiation, SH-SY5Y cells were treated with the PSA-specific endo-N-acetylneuraminidase E. Although loss of PSA was accompanied with a marked reduction of cell growth, it did not interfere with retinoic acid-induced differentiation. Together, our results suggest that PSA surface expression is regulated on the level of polysialyltransferase transcription. Moreover, the similarity to the primary neuroblastoma tissue makes SH-SY5Y cells a suitable model system to examine further the role of polysialylation in tumor cell growth and the orchestration of PSA synthesis in neuroblastoma.