Cellular Signaling through Multifunctional Ca2+/Calmodulin-dependent Protein Kinase II

Ca2+/CaM-dependent protein kinase autoinhibitory domain α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic receptor calmodulin cytoplasmic polyadenylation element cytoplasmic polyadenylation element-binding protein cAMP response element-binding protein disc large protein glutamate receptor subunit 1 of AMPA-R αCaM-KII association protein long term potentiation N-methyld-aspartate receptor neuronal nitric oxide synthase postsynaptic density protein kinase A Psd-Dlg-Zo-1 synapse-associated protein synaptic GTPase-activating protein untranslated region Ca2+/CaM-dependent protein kinase II (CaM-KII)1is a ubiquitously expressed protein kinase that transduces elevated Ca2+ signals in cells to a number of target proteins ranging from ion channels to transcriptional activators. CaM-KII has a unique holoenzyme structure and autoregulatory properties that allow it to give a prolonged response to transient Ca2+ signals and to sense cellular Ca2+ oscillations. In neurons CaM-KII is highly expressed and localized with certain subcellular structures. Upon activation it can translocate to excitatory synapses where it regulates a number of proteins involved in synaptic transmission and its downstream signaling pathways. Elevated intracellular free calcium (Ca2+i), in response to agonist stimulation or cell depolarization, is highly regulated and involves influx through voltage- and ligand-gated Ca2+-permeable ion channels, release from intracellular stores through ryanodine- and inositol 1,4,5-trisphosphate-sensitive channels, sequestration by Ca2+ pumps and exchangers, and signaling through specific Ca2+ transducer proteins (1Bootman M.D. Berridge M.J. Cell. 1995; 83: 675-678Abstract Full Text PDF PubMed Scopus (393) Google Scholar). Changes in intracellular calcium can display variable responses ranging from highly localized, transient elevations within subcellular structures (e.g. a dendritic spine of a neuron) to Ca2+ waves that spread throughout the cell including the nucleus. The most ubiquitous calcium-sensing protein is calmodulin (CaM), which contains four “EF” hand motifs with high specificity for binding Ca2+. The Ca2+/CaM complex interacts with and modulates the functionality of a large number of proteins (2Van Eldik L. Watterson M. Calmodulin and Signal Transduction. Academic Press, New York1998Google Scholar) including several Ser/Thr protein kinases (CaM-Ks). This review, which is part of a series on Ca2+/CaM-dependent protein kinases and phosphatase, will consider one member of this family, the multifunctional CaM-KII. The CaM-KII family is encoded by four genes (α, β, γ, and δ) that also exhibit alternative splicing. The γ and δ isoforms are expressed in most tissues, whereas the α and β isoforms are most prominent in neural tissues and comprise up to 2% of total protein in hippocampus. This review will focus on selected advances over the past 5 years, especially for neuronal CaM-KII, and readers are referred to an earlier comprehensive review for general background information (3Braun A.P. Schulman H. Annu. Rev. Physiol. 1995; 57: 417-445Crossref PubMed Scopus (738) Google Scholar). The various CaM kinase II subunits are comprised of an N-terminal catalytic region, a central regulatory domain containing an autoinhibitory domain (AID) and Ca2+/CaM binding motif, a variable sequence, and the C-terminal subunit association domain (4Soderling T.R. Biochim. Biophys. Acta. 1996; 1297: 131-138Crossref PubMed Scopus (62) Google Scholar) (Fig. 1 A). The holoenzyme is an oligomeric protein comprised of twelve 50–60-kDa subunits arranged as two stacked hexameric rings (5Kolodziej S.J. Hudmon A. Waxham M.N. Stoops J.K. J. Biol. Chem. 2000; 275: 14354-14359Abstract Full Text Full Text PDF PubMed Scopus (141) Google Scholar, 6Kanaseki T. Ikeuchi Y. Sugiura H. Yamauchi T. J. Cell Biol. 1991; 115: 1049-1060Crossref PubMed Scopus (175) Google Scholar). The C-terminal association domains form the central core of each ring with the N-terminal catalytic domains projecting outward (Fig. 1 B). In the absence of bound Ca2+/CaM, the CaM-KII is maintained in an inactive conformation because of an interaction of the AID with the catalytic domain of its own subunit. Three distinct molecular models, based on structure/function studies, have been presented for how the AID might suppress catalytic activity (7Cruzalegui F.H. Kapiloff M.S. Morfin J.P. Kemp B.E. Rosenfeld M.G. Means A.R. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 12127-12131Crossref PubMed Scopus (80) Google Scholar, 8Brickey D.A. Bann J.G. Fong Y.L. Perrino L. Brennan R.G. Soderling T.R. J. Biol. Chem. 1994; 269: 29047-29054Abstract Full Text PDF PubMed Google Scholar, 9Yang E. Schulman H. J. Biol. Chem. 1999; 274: 26199-26208Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar), but definitive proof from a crystal structure has yet to be obtained. The Ca2+/CaM complex binds to a sequence that partially overlaps the AID (Fig.1 A), presumably causing a conformational change and thereby disrupting interaction of the AID with the catalytic domain and producing kinase activation. Interestingly, the sensitivity of CaM-KII to activation by Ca2+/CaM depends on the subunit composition of the holoenzyme (10Brocke L. Chiang L.W. Wagner P.D. Schulman H. J. Biol. Chem. 1999; 274: 22713-22722Abstract Full Text Full Text PDF PubMed Scopus (131) Google Scholar). Upon activation by Ca2+/CaM binding, the kinase undergoes an immediate autophosphorylation on Thr-286 (numbering will be based on the α isoform) (3Braun A.P. Schulman H. Annu. Rev. Physiol. 1995; 57: 417-445Crossref PubMed Scopus (738) Google Scholar). This autophosphorylation occurs within the oligomeric complex (i.e. intramolecular) but between adjacent subunits (intersubunit) that have bound Ca2+/CaM (11Hanson P.I. Meyer T. Stryer L. Schulman H. Neuron. 1994; 12: 943-956Abstract Full Text PDF PubMed Scopus (393) Google Scholar, 12Mukherji S. Soderling T.R. J. Biol. Chem. 1994; 269: 13744-13747Abstract Full Text PDF PubMed Google Scholar). An interesting consequence of the oligomeric structure of CaM-KII discussed above is that it would restrict intramolecular autophosphorylation of Thr-286 within each of the two hexameric rings. This rapid autophosphorylation on Thr-286 has two important regulatory consequences. 1) The subsequent dissociation rate for Ca2+/CaM upon removal of Ca2+ is decreased by several orders of magnitude (13Meyer T. Hanson P.I. Stryer L. Schulman H. Science. 1992; 256: 1199-1202Crossref PubMed Scopus (515) Google Scholar), and 2) even after full dissociation of Ca2+/CaM, the kinase retains partial activity (i.e. Ca2+/CaM-independent or constitutive activity). Presumably the complex holoenzyme structure of CaM-KII (see Fig. 1 B) endows the kinase with this unique regulatory property. Thus, transient elevation of intracellular Ca2+can give a prolonged response through the constitutive activity of autophosphorylated CaM-KII, and this property appears to be critical for certain physiological functions of CaM-KII as discussed later. The efficiency of synaptic transmission between neurons can be modulated, a process known as synaptic plasticity. At excitatory synapses using glutamate as neurotransmitter, synaptic plasticity such as long term potentiation (LTP) is triggered by increased Ca2+ in postsynaptic spines (14Yang S.N. Tang Y.G. Zucker R.S. J. Neurophysiol. 1999; 81: 781-787Crossref PubMed Scopus (417) Google Scholar) and is dependent on the frequency of afferent stimulation (15Mayford M. Wang J. Kandel E.R. O'Dell T.J. Cell. 1995; 81: 891-904Abstract Full Text PDF PubMed Scopus (449) Google Scholar). How is the frequency of Ca2+i oscillations in the postsynaptic spine decoded? CaM-KII has been touted as a decoding mechanism (Fig.2) because of its unique activation properties as discussed above and its localization in dendritic spines in an organelle called the postsynaptic density (PSD) (16Kennedy M.B. Curr. Opin. Neurobiol. 1993; 3: 732-737Crossref PubMed Scopus (129) Google Scholar). The magnitude of constitutive CaM-KII activity because of autophosphorylation of Thr-286 on adjacent subunits in the oligomeric holoenzyme should depend on the duration, amplitude, and frequency of elevated Ca2+i, and a recent in vitro study (17De Koninck P. Schulman H. Science. 1998; 279: 227-230Crossref PubMed Scopus (1088) Google Scholar) shows this to be the case. The abilities of CaM-KII to decode the frequency of synaptic stimulation and to give a prolonged readout beyond the initial stimulus are two characteristics required for a molecule involved in generation of synaptic plasticity. In many cells CaM-KII is largely soluble and widely distributed throughout the cell, but discrete subcellular pools of CaM-KII have recently become recognized. Localization of signaling enzymes close to their substrates has, in general, important regulatory consequences (18Scott J.D. Pawson T. Sci. Am. 2000; 282: 72-79Crossref PubMed Scopus (65) Google Scholar), especially for broad specificity enzymes such as CaM-KII. Thus, characterization of mechanisms for subcellular localization of CaM-KII and its physiological roles are intense areas of investigation. Alternative splice variants of α, δ, and γ isoforms contain a nuclear localization signal (19Srinivasan M. Edman C.F. Schulman H. J. Cell Biol. 1994; 126: 839-852Crossref PubMed Scopus (238) Google Scholar, 20Takeuchi Y. Yamamoto H. Matsumoto K. Kimura T. Katsuragi S. Miyakawa T. Miyamoto E. J. Neurochem. 1999; 72: 815-825Crossref PubMed Scopus (34) Google Scholar), and nuclear CaM-KII is likely to play a role in Ca2+-mediated transcriptional regulation of genes such as brain-derived neurotrophic factor (21Takeuchi Y. Yamamoto H. Miyakawa T. Miyamoto E. J. Neurochem. 2000; 74: 1913-1922Crossref PubMed Scopus (33) Google Scholar) and atrial natriuretic factor (22Ramirez M.T. Zhao X.L. Schulman H. Brown J.H. J. Biol. Chem. 1997; 272: 31203-31208Abstract Full Text Full Text PDF PubMed Scopus (175) Google Scholar) through phosphorylation of transcription factors including CCAAT/enhancer-binding protein (C/EBP) (23Wegner M. Cao Z. Rosenfeld M.G. Science. 1992; 256: 370-373Crossref PubMed Scopus (308) Google Scholar, 24Yano S. Fukunaga K. Takiguchi M. Ushio Y. Mori M. Miyamoto E. J. Biol. Chem. 1996; 271: 23520-23527Abstract Full Text Full Text PDF PubMed Scopus (38) Google Scholar). It is intriguing that CaM kinases I and IV can phosphorylate a Ser adjacent to the nuclear localization signal and prevent nuclear localization of the CaM-KII, but whether this occurs physiologically is uncertain (25Heist E.K. Srinivasan M. Schulman H. J. Biol. Chem. 1998; 273: 19763-19771Abstract Full Text Full Text PDF PubMed Scopus (98) Google Scholar). CaM-KI exhibits broad cellular distribution and is largely cytoplasmic, whereas CaM-KIV has a rather restricted tissue distribution and exists as both cytosolic and nuclear isoforms (26Soderling T.R. Trends Biochem. Sci. 1999; 24: 232-235Abstract Full Text Full Text PDF PubMed Scopus (442) Google Scholar). CaM-KII may also exert negative effects on transcription through phosphorylation of the transcription factor CREB. Surprisingly, although the activation site (Ser-133) in CREB can be efficiently phosphorylated by CaM-KII, it simultaneously phosphorylates another site (Ser-142) that exerts a dominant negative role (27Sun P. Enslen H. Myung P.S. Maurer R.A. Genes Dev. 1994; 8: 2527-2539Crossref PubMed Scopus (649) Google Scholar). Thus, it is possible that nuclear CaM-KII can inhibit CREB-dependent transcription meditated by kinases such as PKA, but this needs to be verified under physiological conditions. Ca2+-stimulated gene transcription through CREB is mediated in part by CaM-KIV (28Bito H. Deisseroth K. Tsien R.W. Cell. 1996; 87: 1203-1214Abstract Full Text Full Text PDF PubMed Scopus (977) Google Scholar,29Ahn S. Ginty D.D. Linden D.J. Neuron. 1999; 23: 559-568Abstract Full Text Full Text PDF PubMed Scopus (149) Google Scholar). Anchoring proteins that localize PKA (18Scott J.D. Pawson T. Sci. Am. 2000; 282: 72-79Crossref PubMed Scopus (65) Google Scholar) and protein kinase C (30Mochly-Rosen D. Gordon A.S. FASEB J. 1998; 12: 35-42Crossref PubMed Scopus (510) Google Scholar) close to physiological substrates have been well characterized, and recent studies indicate a similar regulatory scheme for CaM-KII. An anchoring protein, αKAP, has recently been identified that localizes skeletal muscle CaM-KII to the sarcoplasmic reticulum. This unique protein contains a hydrophobic N terminus fused to the C-terminal association domain of CaM-KII (31Bayer K.U. Lohler J. Harbers K. Mol. Cell. Biol. 1996; 16: 29-36Crossref PubMed Google Scholar). The C-terminal association domain of αKAP can form heteromers with the full-length CaM-KII subunit, and the hydrophobic N terminus of αKAP directs the resulting kinase complex to the sarcoplasmic reticulum membrane (32Bayer K.U. Harbers K. Schulman H. EMBO J. 1998; 17: 5598-5605Crossref PubMed Scopus (112) Google Scholar). Likely substrates for CaM-KII in the sarcoplasmic reticulum include the ryanodine receptor (33Witcher D.R. Kovacs R.J. Schulman H. Cefali D.C. Jones L.R. J. Biol. Chem. 1991; 266: 11144-11152Abstract Full Text PDF PubMed Google Scholar), phospholamban (34Wegener A.D. Simmerman H.K. Lindemann J.P. Jones L.R. J. Biol. Chem. 1989; 264: 11468-11474Abstract Full Text PDF PubMed Google Scholar), and the Ca2+-ATPase pump (35Xu A. Hawkins C. Narayanan N. J. Biol. Chem. 1993; 268: 8394-8397Abstract Full Text PDF PubMed Google Scholar). In brain, there is evidence for colocalization of the CaM-KII β isoform with the cytoskeleton. Upon stimulation of the Ca2+-permeable NMDA-R ion channel in hippocampal neurons, CaM-KII appears to dissociate from F-actin and undergo translocation to membranous fractions including the PSD (36Shen K. Meyer T. Science. 1999; 284: 162-166Crossref PubMed Scopus (541) Google Scholar). The PSD, a complex of postsynaptic membrane proteins involved in mediating and modulating synaptic transmission, is held together and to the cytoskeleton through anchoring proteins of the PDZ/SAP family (16Kennedy M.B. Curr. Opin. Neurobiol. 1993; 3: 732-737Crossref PubMed Scopus (129) Google Scholar, 37Kennedy M.B. Brain Res. Rev. 1998; 26: 243-257Crossref PubMed Scopus (161) Google Scholar). CaM-KII is a major constituent of the PSD where it is anchored in part through the protein densin-180 (38Strack S. Robison A.J. Bass M.A. Colbran R.J. J. Biol. Chem. 2000; 275: 25061-25064Abstract Full Text Full Text PDF PubMed Scopus (85) Google Scholar). This interaction of CaM-KII with densin-180 does not appear to depend on the activation state of the kinase. In contrast, additional CaM-KII can associate with the PSD through interaction with the NMDA-type glutamate-gated ion channel, but this translocation appears to require activation of the kinase and its autophosphorylation on Thr-286 (39Strack S. Choi S. Lovinger D.M. Colbran R.J. J. Biol. Chem. 1997; 272: 13467-13470Abstract Full Text Full Text PDF PubMed Scopus (252) Google Scholar, 40Strack S. Colbran R.J. J. Biol. Chem. 1998; 273: 20689-20692Abstract Full Text Full Text PDF PubMed Scopus (387) Google Scholar). Translocation to the PSD occurs in hippocampal slices upon treatments that activate CaM-KII, and it promotes the phosphorylation of CaM-KII substrates in the PSD such as the AMPA-R (39Strack S. Choi S. Lovinger D.M. Colbran R.J. J. Biol. Chem. 1997; 272: 13467-13470Abstract Full Text Full Text PDF PubMed Scopus (252) Google Scholar, 41Leonard A.S. Lim I.A. Hemsworth D.E. Horne M.C. Hell J.W. Proc. Natl. Acad. Sci. U. S. A. 1999; 96: 3239-3244Crossref PubMed Scopus (334) Google Scholar). The anchoring interaction appears to occur between the catalytic domain of CaM-KII and residues 1290–1309 in the cytosolic tail of the NR2B subunit of the NMDA-R (42Strack S. McNeill R.B. Colbran R.J. J. Biol. Chem. 2000; 275: 23798-23806Abstract Full Text Full Text PDF PubMed Scopus (277) Google Scholar). Translocation would localize CaM-KII a critical site of Ca2+ influx the dendritic spine because the channel has Ca2+ and its activation is required for several of synaptic plasticity. In to CaM-KII to a site of Ca2+ this translocation also the kinase in close to several important neuronal substrates (Fig. as discussed is evidence that activation of CaM-KII in neurons of region in is involved in the of T.R. Trends 2000; 23: Full Text Full Text PDF PubMed Scopus Google Scholar, R.A. 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A. 1999; 96: PubMed Scopus Google Scholar). of neurons that exhibit potentiation an in A. 1998; PubMed Scopus Google Scholar). with also a role of CaM-KII phosphorylation of as an important of in the AMPA-R subunit phosphorylated by CaM-KII, a specific in D. N. A. T. P. J. M. P. Science. 1999; 284: PubMed Scopus Google Scholar). a in αCaM-KII in a that is in K. N. A. Science. 1998; 279: PubMed Scopus Google Scholar). This does not activation of CaM-KII through binding of Ca2+/CaM, but it generation of constitutive activity by Thus, CaM-KII phosphorylation of with potentiation of is to to the of T.R. Trends 2000; 23: Full Text Full Text PDF PubMed Scopus Google Scholar, R.A. Science. 1999; PubMed Scopus Google Scholar). PSD proteins can also be phosphorylated by CaM-KII. the NMDA-R that as an for CaM-KII can be phosphorylated by CaM-KII M.J. A.J. M.B. J. Biol. 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