Involvement of calcium/calmodulin-dependent protein kinase II in the induction of mPer1

Recent studies suggest that CaM kinase II is involved in light-induced phase delays and induction of Per1 and Per2 genes in the hamster suprachiasmatic nucleus (SCN) (Yokota et al., 2001). We focused on intracellular mechanisms of the CaM kinase II-induced mPer1 gene expression. Immunoblotting and immunohistochemical analyses with isoform-specific antibodies against different isoforms of CaM kinase II and CaM kinase IV showed abundant expression of the δ isoform of CaM kinase II without significant expression of CaM kinase IV in the lateral ventral region of the rat SCN. We next defined the CaM kinase II-responsive region on the mPer1 promoter using a luciferase reporter gene assay. Transfection of the constitutively-active CaM kinase 11δ greatly increased mPer1 promoter activity in NG108-15 cells and increased activity slightly but significantly in NB2A and C6 glioma cells. Similarly, transfection of a constitutivelyactive MEKK, an upstream kinase of mitogen-activated protein kinase (MAPK), greatly increased promoter activity in NB2A cells. Deletion and mutation analyses of the mPer1 promoter revealed that a 5′-GAGGGG-3′ sequence motif near exon 1B, in which several zinc finger proteins seem to bind, was essential for the CaM kinase II-induced activation of the mPer1 promoter. These results suggest that CaM kinase 11δ but not CaM kinase IV plays an essential role for mPer1 expression through the 5′-GAGGGG-3′ motif on the mPer1 promoter.