Developmental and regional expression of multifunctional Ca2+/calmodulin-dependent protein kinase isoforms in rat brain

Abstract

Multifunctional Ca2+/calmodulin-dependent protein kinase (CaM kinase) participates in diverse calcium signaling pathways in neurons. The alpha- and beta-CaM kinase isoforms are neuron-specific and highly abundant in rat brain. The variable domain of CaM kinase is a potential site for the generation of isoform diversity by alternative spicing of its N- and/or C-terminal segments. We used specific PCR primers which span the variable domain of either alpha- or beta-CaM kinase and isolated three new isoforms from rat brain, namely alpha B-, beta e- and beta'e-CaM kinase. alpha beta-CaM kinase contains 11 amino acids, likely inserted by alternative splicing, at the C-terminal segment of the variable domain. This insertion introduces a nuclear localization signal (NLS) that targets alpha B-CaM kinase to the nucleus of transfected cells; alpha-CaM kinase is excluded from the nucleus. The mRNA and the protein corresponding to this isoform are detected only in the diencephalon/midbrain regions. We have also identified two alternatively spliced isoforms of beta-CaM kinase that lack the 24 amino acid sequence at the N-terminal segment of the variable domain. Alternative splicing of these two isoforms occurs with a three base pair shift of the 3'-splice site. Our analysis shows that these new beta-CaM kinase isoforms are expressed primarily in early developmental stages, and we therefore term them beta e - (embryonic) and beta' e-CaM kinase. Recombinant alpha B-, beta e and beta' e-CaM kinase expressed in COS-7 cells exhibit characteristic Ca2+/calmodulin-dependent protein kinase activity and autophosphorylation.

Activity-Dependent Regulation of Calcium/Calmodulin-Dependent Protein Kinase II Localization: Figure 1.

“…Splice variants of the ␣ and the ␦ gene (␣ B -and ␦ B -CaMKII) contain a nuclear localization signal (NLS) that directs each of these to the nucleus (Srinivasan et al, 1994;Brocke et al, 1995). Not surprisingly, CaMKII-mediated changes in gene expression require nuclear localization (Ramirez et al, 1997).…”

“…A transcription factor, NeuroD, that regulates dendritic growth and morphology has been found recently to be phosphorylated by CaMKII in granule cells, but it is not known which kinase isoform is involved (Gaudilliere et al, 2004). ␣ B -CaMKII is abundant in brain but essentially restricted to the midbrain and diencephalon, regions that have approximately equal amounts of ␣-and ␣ BCaMKII (Walaas et al, 1983;Brocke et al, 1995). Holoenzymes with primarily nuclear subunits are found almost exclusively in the nucleus and vice versa, whereas both nuclear and cytosolic localization is seen at equal expression of the two isoforms (Srinivasan et al, 1994).…”

Identification and Expression of δ-Isoforms of the Multifunctional Ca 2+ /Calmodulin-Dependent Protein Kinase in Failing and Nonfailing Human Myocardium

“…4,5 Alternative splicing of the primary transcripts encoded by 4 genes (␣, ␤, ␥, and ␦) results in the generation of different CaMKII variants. 6,7 Members of the ␣-and ␤-classes are expressed mainly in neuronal tissue, whereas ␥-and ␦-CaMKII isoforms are abundantly expressed in all tissues. 6,8,9 Multifunctionality of the homomultimeric or heteromultimeric holoenzyme composed of 8 through 12 subunits 10 seems to counteract its specific cellular functions.…”

Inositol 1,4,5-Trisphosphate 3-Kinase A Associates with F-actin and Dendritic Spines via Its N Terminus

“…Despite its proposed ability to bind F-actin, some CaMKII␤ in some cells appeared cytosolic. This may be because this protein occurs in at least four splice forms (32), and some of the splicing occurs in the suggested actin-binding region (31). In summary, IP3kinA was usually found to be co-localized with CaMKII, its endogenous activator, and this co-localization occurred more often with the ␤ than the ␣ isoform.…”