Neuronal expression of the chondroitin sulfate proteoglycans receptor-type protein-tyrosine phosphatase β and phosphacan

• DOI: 10.1016/j.neuroscience.2004.11.017
• Publication Date: 2005-01-29T12:22:16Z
• Authors: N. Hayashi; S. Miyata; M. Yamada; K. Kamei; A. Oohira
• Keywords: Neurons; Protein Tyrosine Phosphatase, Non-Receptor Type 1; 0303 health sciences; Cell-Free System; Receptor-Like Protein Tyrosine Phosphatases, Class 5; Brain; Gene Expression Regulation, Developmental; Nerve Tissue Proteins; Rats; 3. Good health; 03 medical and health sciences; Chondroitin Sulfate Proteoglycans; Astrocytes; Animals; Protein Tyrosine Phosphatases; Rats, Wistar; Cells, Cultured

Receptor-type protein-tyrosine phosphatase beta (RPTPbeta) and its spliced variant phosphacan are major components of chondroitin sulfate proteoglycans in the CNS. In this study, expression and localization of RPTPbeta and phosphacan were examined in developing neurons by immunological analyses using 6B4, 3F8, and anti-PTP antibodies and reverse transcription-polymerase chain reaction (RT-PCR). Light microscopic immunohistochemistry showed that 6B4 RPTPbeta/phosphacan immunoreactivity was observed around neurons in the cortical plate. Further ultrastructural observation showed that 6B4 RPTPbeta/phosphacan immunoreactivity was observed mainly at the membrane of migrating neurons and radial glia. Immunocytochemical analysis revealed that RPTPbeta immunoreactivity was observed in cultured cerebral, hippocampal, and cerebellar neurons in addition to type-1 and type-2 astrocytes. Western analysis further demonstrated that the shorter receptor form of RPTPbeta (sRPTPbeta) was detected from cell lysate of cortical and hippocampal neurons using 6B4 and anti-PTP antibodies, while sRPTPbeta of cerebellar neurons and type-1 astrocytes was recognized only by anti-PTP antibody. Phosphacan was detected from neuronal culture supernatants of cortical, hippocampal, and cerebellar neurons, but not from type-1 astrocytes using 6B4 and 3F8 antibodies. RT-PCR analysis demonstrated the prominent expression of sRPTPbeta and phosphacan mRNAs in cortical neurons, and that of sRPTPbeta mRNA in type-1 astrocytes. During culture development of cortical neurons, the immunoreactivity of 6B4 sRPTPbeta was observed entirely on the neuronal surface including somata, dendrites, axons, and growth cones at earlier stages of cortical neuronal culture such as stages 2 and 3, while, after longer culture, 6B4 sRPTPbeta immunoreactivity in stages 4 and 5 neurons was detected at dendrites and somata and disappeared from axons, and was not observed over axonal terminals and postsynaptic spines. These results demonstrate that neurons are able to express sRPTPbeta on their cellular surface and to secrete phosphacan, and neuronal expression of sRPTPbeta may modulate neuronal differentiation including neuritogenesis and synaptogenesis.