Induction and alternative splicing of δ isoform of Ca2+/calmodulin-dependent protein kinase II during neural differentiation of P19 embryonal carcinoma cells and during brain development

Abstract

Since the expression of Ca2+/calmodulin-dependent protein kinase II (CaM kinase II) is regulated during brain development, the developmental change of the enzyme was investigated during the neural differentiation of murine P19 embryonal carcinoma cells. CaM kinase II activity was induced during the differentiation of P19 cells treated with retinoic acid. Expression of the enzyme was induced 2 days after the treatment and maximized at 5 days. The enzyme activity increased about ∼8-fold. The enzyme protein was shown to differ between differentiated and undifferentiated cells. The δ isoform of CaM kinase II was found as the major isoform in P19 cells by immunoblotting and reverse transcription-polymerase chain reaction (RT-PCR). A total of four and three alternatively spliced variants of δ isoform were detected in P19 cells by RT-PCR analysis and by immunoblotting, respectively. Although multiple alternatively spliced forms have been reported, the major splice variants of δ isoform in differentiated cells were δ1 and δ9 isoforms, which were specifically detected in differentiated cells. In undifferentiated cells, the major splice variant corresponded to δ2 isoform. These results indicated that the expression of δ isoform of CaM kinase II was induced, and the splicing pattern of the isoform changed, during neural differentiation. Cell type distinctive changes of splicing pattern of δ isoform were also observed not only during differentiation of cultured neuronal cells, but also during development of rat forebrain and cerebellum. © 2000 Elsevier Science B.V.