CaMKII

CaMKII. Calcium/Calmodulin Dependent Protein Kinase II. Ver. 0.3. This calcium calmodulin-dependent protein kinase is composed of four different chains: alpha, beta, gamma, and delta. An enzyme belongs to the serine/threonine protein kinase family, and to the Ca(2+)/calmodulin-dependent protein kinase subfamily.

https://www.genecards.org/cgi-bin/carddisp.pl?gene=CAMK2A

https://www.genecards.org/cgi-bin/carddisp.pl?gene=CAMK2B

https://www.genecards.org/cgi-bin/carddisp.pl?gene=CAMK2G

https://www.genecards.org/cgi-bin/carddisp.pl?gene=CAMK2D

Ca2+–calmodulin-dependent kinase isoforms in human skeletal muscle. (Rose 2006) [1]

Detection of multifunctional Ca2+–calmodulin-dependent kinase isoforms in human skeletal muscle. Rat brain (RB) and human skeletal muscle (HSkM) extract proteins were immunoblotted using isoform-specific CaM kinase antibodies. For skeletal muscle CaMKII the bands at 55–60 kDa probably correspond to γ/δ isoforms, whereas the 73 kDa band is likely to be the muscle-specific β isoform splice variant termed βM (Bayer et al. 1998; Damiani et al. 2000).

The relative estimates of the amount of CaMKII isoforms in human skeletal muscle based on proteomics data from (Murgia 2017) [2]

CaMKII subunit alpha	CAMK2A	348	nmol/L
CaMKII subunit beta	CAMK2B	482	nmol/L
CaMKIII subunit delta	CAMK2D	457	nmol/L
CaMKII subunit gamma	CAMK2G	203	nmol/L

Summary remarks

Unlike many other protein kinases, CaMKII does not contain a phosphorylatable residue analogous to the activation domains of CaMKI and CaMKIV. Thus, CaMKII is not phosphorylated or activated by CaMKK2. Transient elevation of intracellular Ca2+ can give a prolonged response through the constitutive activity of autophosphorylated CaMKII.

The different kinase isoforms have different CaM binding affinity and different CaM dependence for their autophosphorylation.

There is not enough data yet, so it does not make sense to consider different kinase isoforms for simulations in the model. In order to make sense to divide the isoforms in the current model, it is necessary that the following criteria be fulfilled: significantly different kinetics of isoforms, different targets (substrates) for different isoforms, and the presence of studies on the activity of different isoforms in human muscles, which can be used for model validation. These criteria are not met up to date, except for different kinetics.

Structure

(Hudmon 2002) [3] CaMKII is derived from a family of four homologous but distinct genes, with over 30 alternatively spliced isoforms described at present.

(Soderling 2001) [4]. Unlike many other protein kinases, CaM-KII does not contain a phosphorylatable residue analogous to the activation domains of CaM-KI and CaM-KIV. Thus, CaM-KII is not phosphorylated or activated by CaM-KK2. However, the unique holoenzyme structure of CaM-KII endows it with unusual regulatory properties required for sensing and transducing various types of intracellular Ca2+ signals. Upon activation by Ca2+/CaM binding, the kinase undergoes immediate autophosphorylation on Thr286 (numbering based on the R isoform). (59) This autophosphorylation occurs within the oligomeric complex (i.e., intramolecular) but between adjacent subunits (intersubunit) that have bound Ca2+/CaM. (63,64) This rapid autophosphorylation on Thr286 has two important regulatory consequences: (1) the subsequent dissociation rate for Ca2+/CaM upon removal of Ca2+ is decreased by several orders of magnitude, (65) and (2) even after full dissociation of Ca2+/CaM, the kinase retains partial activity (i.e., Ca2+/CaM-independent or constitutive activity). Presumably, the stacked hexameric ring structure of the CaM-KII holoenzyme restricts intersubunit autophosphorylation to within each ring. 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.

Linear diagram of a prototypical CaMKII subunit The catalytic domain is autoinhibited by a pseudosubstrate autoregulatory sequence that is disinhibited following Ca2+/CaM binding. The association domain produces the native form of the enzyme, a multimeric holoenzyme composed of 12 subunits. Isoform differences present in the α, β, γ and δ isoforms of CaMKII are contributed primarily by a region of multiple alternatively spliced sequences, termed variable inserts, which reside in the association domain. Conserved sites of autophosphorylation are indicated in the autoregulatory region.

Basal autophosphorylation at threonine 306 blocks calmodulin binding, resulting in inactivation of CaMKII (Colbran 1993) [5].

(Soderling 2001) [4]. In the absence of bound Ca2+/CaM, the CaM-KII is maintained in an inactive conformation due to an interaction of an AID with the catalytic domain of its own subunit. Interestingly, the sensitivity of CaMKII to activation by Ca2+/CaM is dictated by the subunit composition of the holoenzyme. (Brocke ‎1999) [6].

The holoenzyme model (Rosenberg 2005) [7]. Iinactive CaMKII forms tightly packed autoinhibited assemblies that convert upon activation into clusters of loosely tethered and independent kinase domains.

The domains of human CaMKII-α (Bhattacharyya 2016) [8]. The architecture of the CaMKII holoenzyme is shown on the right. The hub domain forms a dodecamer or tetradecamer, and the kinase domains are flexibly linked to it by the autoinhibitory segment.

The mass spectra demonstrate that the isolated hub assembly exists as a ~ 1:1 mixture of dodecamers and tetradecamers in solution.

The principal difference between CaMKII-α and CaMKII-β isoforms is the length of the linker, which is 218 residues long in CaMKII-β, compared to 30 residues in CaMKII-α. …the phenomenon of activation-triggered subunit exchange is not limited to just the α isoform of CaMKII. CaMKII-β can also exchange subunits with CaMKII-α in an activation-dependent manner, leading to the formation of CaMKII heterooligomers.

CaMKII undergoes multiple autoregulatory states that may have an impact on its function following Ca2+/CaM activation.

The multimeric holoenzyme structure of CaMKII is depicted as a 6-mer for simplicity, with activated catalytic subunits illustrated in red. In addition to the Ca2+/CaM-dependent form of the enzyme phosphorylating physiological targets throughout the cell, autophosphorylation produces functional changes in CaMKII (autonomous activity, CaM trapping, CaM capping) that may alter further its activity, regulation, and function.

The autophosphorylation of CaMKII requires coincident Ca2+/CaM binding between neighbouring subunits within the holoenzyme. The catalytic/autoregulatory region is shown for each subunit. In the absence of Ca2+/CaM, the blue autoregulatory domain inhibits catalytic activity (inactivity illustrated in red). Ca2+/CaM binding to each subunit independently disinhibits the autoregulatory region to produce catalytic activity (green). Activated subunits may then act as both kinase and substrate in an intersubunit ± intraholoenzyme autophosphorylation reaction of Thr286 (shown as ` T ' within the red circle). Only the autophosphorylated subunit retains activity in the absence of Ca2+/CaM. Although this autonomous activity is uncoupled from its dependence on Ca2+/CaM, this enzymic state is sensitive to phosphatase activity. (Bhattacharyya 2016) [8]. Phosphorylation on Thr 286 increases the affinity of CaMKII for Ca2+/CaM by more than 1000-fold, by fully or partially releasing the autoinhibitory segment from the kinase domain (Meyer et al., 1992).

A model encompassing the biophysical and enzymic characteristics of CaMKII displays a non-linear Ca2+ spike frequency-dependence in the generation of autonomous activity (left) and the role of coincident CaM binding, autophosphorylation and CaM trapping in Ca2+-spike frequency detection (right).

Inactive subunits within CaMKII holoenzymes (shown as 6-mers for simplicity) are represented by open circles. Ca2+/CaM (blue circle) binding activates a given subunit (shown in red) and coincident Ca2+/CaM binding results in autophosphorylation and CaM trapping ('P' inside a filled dark red circle). During low-frequency Ca2+ spikes, CaM completely dissociates between Ca2+ spikes, effectively producing 'naive' CaMKII subunits at each inner spike interval. However, at high Ca2+ spike frequencies, CaM does not completely dissociate between Ca2+ spikes, which increases the probability that coincident CaM binding, autophosphorylation, and CaM trapping may occur; a process that effectively increases the probability that a neighboring subunit will also bind Ca2+/CaM during successive spikes to produce the dramatic non-linear increase in autonomous activity.

Action

(Sacchetto et al., 2005) http://www.bio.unipd.it/bam/PDF/15-1/04569SacchettoDamiani.pdf

In slow-twitch muscle, CaMKII is mainly involved in regulation of SR Ca2+-transport, as indicated by phosphorylation of SERCA2a isoform of Ca2+-ATPase and of SERCA2a-regulatory protein phospholamban. In fast-twitch muscle, the main role of CaMKII seems to be the control of Ca2+- release. Much experimental evidence seems to negate that ryanodine receptor/SR Ca2+- release channel of skeletal muscle (RyR1) is a substrate of phosphorylation of endogenous CaMKII.

(Rose 2006) [1]. In response to exercise, there was a transient increase in autonomous CaMKII activity and phosphorylation. In particular, when measured from skeletal muscle lysates, autonomous CaMKII activity was 1.3±0.2% of maximal at rest, and increased to 11.3±1.4% (12±3-fold) of maximal after 1 min of exercise but then decreased to 2–4% (1- to 3-fold higher than basal) of maximal by 10 min of exercise and remained at this level for the remaining 90 min of exercise.

The Ca2+–calmodulin-dependent protein kinase II phosphorylation during exercise has the same pattern.

Despite these changes in CaMKII activity, the PLN phosphorylation (CaMKII substrate) plateaus after 10 min of exercise

(Rose 2006) [1]. PLN phosphorylation at Thr17 may be representative of CaMKII activity in vivo and plateaus after 10 min of exercise… the effect of exercise intensity was examined using an exercise bout with 10 min stages of incrementally graded exercise intensity. Skeletal muscle PLN phosphorylation at Thr17 was approximately 1- to 3-fold higher with exercise intensities of 35% and 60% V˙O2peak, but was higher still (3- to 5-fold) at 85% V˙O2peak when compared with rest. PLN phosphorylation at Thr17 had returned to resting levels at 30 min post-exercise.

(Rose 2006) [1]. In vitro autonomous Ca2+–calmodulin-dependent protein kinase II activity is positively related to phosphorylation at Thr287 Correlation between Ca2+–calmodulin-dependent protein kinase II (CaMKII) phosphorylation at Thr287 and autonomous CaMKII activity from skeletal muscle samples taken before and during exercise (A; r 2 = 0.884, P < 0.01) and from basal skeletal muscle extracts (n = 4) manipulated to differing levels of phosphorylation by incubation with differing amounts of Ca2+ and calmodulin (B; r 2 = 0.996, P < 0.01).

(Rose 2006) [1]. It should be noted that the increase in autonomous activity was only up to 2–4% of Ca2+–CaM-stimulated activity during the majority of exercise time, meaning that the rise in activity during the Ca2+ ‘spike’ may be quantitatively more important in the overall increase in CaMKII activity. Indeed, the patterns of CaMKII phosphorylation and phosphorylation of the putative CaMKII substrate PLN, which may be indicative of in vivo CaMKII activity during exercise, were remarkably different. The transient nature of the increase in autophosphorylation with exercise is difficult to explain, but may be related to differences in Ca2+ uptake and release kinetics by the sarcoplasmic reticulum and thus Ca2+–CaM binding, or a differential action of CaMKII phosphatases, on CaMKII over time. Indeed, skeletal muscle CaMKII may indirectly regulate its own dephosphorylation by increasing protein phosphatase-1c (PP1c) activity through phosphorylation of the PP1c-targeting subunit GM (Sacchetto et al. 2005b).

Location

(Sacchetto et al., 2005) http://www.bio.unipd.it/bam/PDF/15-1/04569SacchettoDamiani.pdf

CaMKII is localized on the cytoplasmic side of SR membranes, and is ubiquitously distributed between longitudinal SR and junctional SR.

(Chin 2005) [9], The CaMKII isoform designated CaMKII-βM is predominantly associated with skeletal muscle SR and is anchored there by a truncated, non-kinase protein α-KAP, which is a splice variant of CaMKII- α (4). Furthermore, the various CaMKII- βM splice variants (-β, -βM and - βe’) have similar specific activity but differential sensitivities to Ca2+ due to differences in their CaM activation constant and thus initial rates of autophosphorylation (5).

The majority (i.e. ∼80%) of CaMKII that is expressed in human skeletal muscle is localized to the soluble cytosolic fraction. The majority of work that has been conducted has examined the functional properties and substrates of membrane-associated CaMKII (present study; Sacchetto et al. 2005a). Thus, it is clear that future work should focus on examining the potential substrates and functions of cytosolic CaMKII. (Rose 2006) [1]

Pathways

Upstream

Ca2+/CaM, (Hudmon 2002) [3].

PP1, protein phosphatase-1, (Sacchetto 2005b) [10].

Perm1 (PGC-1 and ERR induced regulator, muscle 1) associates with skeletal muscle CaMKII and promotes CaMKII activation, (Cho 2019) [11].

(Coultrap 2014) [11]. Two novel mechanisms for generating CaMKII autonomy include oxidation and S-nitrosylation, the latter requiring both Cys280 and Cys289 amino acid residues of the brain isoform, CaMKIIα. … nitric oxide (NO)-signaling generated autonomy also for the CaMKIIβ isoform. …oxidation of the Met280/281 residues is sufficient to generate autonomy for most CaMKII isoforms. Thus, all CaMKII isoforms can be regulated by physiological NO-signaling, but CaMKIIα regulation by oxidation and S-nitrosylation is more stringent.

(Morales-Alamo 2012) [12]. RNOS may also activate CaMKII through modification of the Met281/282 pair within the regulatory domain, blocking reassociation with the catalytic domain and preserving kinase activity via a similar but parallel mechanism to Thr286 autophosphorylation (13). Thus, reducing RNOS by antioxidant administration could result in lower CaMKII activity by a mechanism unrelated to sarcoplasmic Ca2+.

(Huang 2014) [13]. …We show here that this activation of CaMKII depends, in part, on dephosphorylation of CaMKII at novel sites (Thr393/Ser395) and that this is mediated by metabolic activation of protein phosphatase 2A in complex with the B55β targeting subunit. This represents a novel locus of CaMKII control.

Downstream

PLN, SRF, GS, NR2B, SERCA, CREB, c-fos, FAP, AP-1, GAPDH, GSK-3β, caspase-2

Phospholamban (Sacchetto et al., 2005) http://www.bio.unipd.it/bam/PDF/15-1/04569SacchettoDamiani.pdf

Phospholamban (PLN) was the only identified substrate of CaMKII in human skeletal muscle (Margreth et al. 2000) www.bio.unipd.it/bam/PDF/10-4/00442Margreth.pdf, (Rose 2006) [1].

SRF (Flück ‎2000) [14], Isoforms of CaMKII enriched from skeletal muscle phosphorylated SRF in vitro (Fluck 2000b) [15].

GS SR-associated glycogen synthase (GS) was phosphorylated in vitro by SR-bound Ca2+-calmodulin-dependent protein kinase (CaMKII) (Sacchetto 2007) [16].

CaMKII was shown to phosphorylate serum response factor (SRF). Other known substrates of CaMKII include NR2B subunit of the N-methyl-D-aspartate (NMDA) receptor (44), NF-B (24), cardiac calcium release channel (20), SERCA (57), and CREB (45). Other known intermediates for CaMKII signaling include c-fos, FAP, and AP-1 (51). Thus CaMKII may activate transcription directly through posttranslational modification of transcription factors (i.e., SRF, CREB, c-fos, FAP, AP-1). (Chin 2005) [9].

RyR1 is a substrate of phosphorylation of endogenous CaMKII. (Sacchetto et al., 2005) http://www.bio.unipd.it/bam/PDF/15-1/04569SacchettoDamiani.pdf

It has also been shown that skeletal muscle CaMKII can phosphorylate glyceraldehyde-3-phosphate dehydrogenase (Sacchetto et al. 2005b; Singh et al. 2004)… and the glycogen and protein phosphatase-1c targeting subunit GM (Sacchetto et al. 2005b) in a Ca2+–CaM-dependent manner. (Rose 2006) [1].

(Aversa 2013) [17]. Glycogen synthase kinase GSK-3β, a downstream target of CaMKII.

(Huang 2014) [13]. …This metabolic control is exerted via inhibitory phosphorylation of the caspase-2 prodomain by activated Ca2+/calmodulin-dependent protein kinase II (CaMKII).

Kinetics

(Gaertner 2004) [18] 

Interestingly, the rank order of CaM binding affinity (γ > β > δ > α) does not directly correlate with the rank order of their CaM dependence for autophosphorylation (β > γ > δ > α ). Simulations utilizing this data revealed that the measured differences in CaM binding affinities play a minor role in the autophosphorylation of the enzyme, which is largely dictated by the rate of autophosphorylation for each isoform.

Dependence of CaM-kinase II activity on CaM

Dependence of CaM-kinase II autophosphorylation on CaM

Ca2+/CaM-stimulated CaMKII activity is mildly (~1.2–1.3 fold) further increased by additional T286 autophosphorylation, but that this autophosphorylation is not required for the major part of the stimulated activity. (Coultrap 2012) [19]

References

1. Rose AJ, Kiens B, and Richter EA. Ca2+-calmodulin-dependent protein kinase expression and signalling in skeletal muscle during exercise. J Physiol. 2006 Aug 1;574(Pt 3):889-903. DOI:10.1113/jphysiol.2006.111757 |
2. Murgia M, Toniolo L, Nagaraj N, Ciciliot S, Vindigni V, Schiaffino S, Reggiani C, and Mann M. Single Muscle Fiber Proteomics Reveals Fiber-Type-Specific Features of Human Muscle Aging. Cell Rep. 2017 Jun 13;19(11):2396-2409. DOI:10.1016/j.celrep.2017.05.054 |
3. Hudmon A and Schulman H. Structure-function of the multifunctional Ca2+/calmodulin-dependent protein kinase II. Biochem J. 2002 Jun 15;364(Pt 3):593-611. DOI:10.1042/BJ20020228 |
4. Soderling TR and Stull JT. Structure and regulation of calcium/calmodulin-dependent protein kinases. Chem Rev. 2001 Aug;101(8):2341-52. DOI:10.1021/cr0002386 |
5. Colbran RJ. Inactivation of Ca2+/calmodulin-dependent protein kinase II by basal autophosphorylation. J Biol Chem. 1993 Apr 5;268(10):7163-70.
6. Brocke L, Chiang LW, Wagner PD, and Schulman H. Functional implications of the subunit composition of neuronal CaM kinase II. J Biol Chem. 1999 Aug 6;274(32):22713-22. DOI:10.1074/jbc.274.32.22713 |
7. Rosenberg OS, Deindl S, Sung RJ, Nairn AC, and Kuriyan J. Structure of the autoinhibited kinase domain of CaMKII and SAXS analysis of the holoenzyme. Cell. 2005 Dec 2;123(5):849-60. DOI:10.1016/j.cell.2005.10.029 |
8. Bhattacharyya M, Stratton MM, Going CC, McSpadden ED, Huang Y, Susa AC, Elleman A, Cao YM, Pappireddi N, Burkhardt P, Gee CL, Barros T, Schulman H, Williams ER, and Kuriyan J. Molecular mechanism of activation-triggered subunit exchange in Ca(2+)/calmodulin-dependent protein kinase II. Elife. 2016 Mar 7;5. DOI:10.7554/eLife.13405 |
9. Chin ER. Role of Ca2+/calmodulin-dependent kinases in skeletal muscle plasticity. J Appl Physiol (1985). 2005 Aug;99(2):414-23. DOI:10.1152/japplphysiol.00015.2005 |
10. Sacchetto R, Bovo E, Donella-Deana A, and Damiani E. Glycogen- and PP1c-targeting subunit GM is phosphorylated at Ser48 by sarcoplasmic reticulum-bound Ca2+-calmodulin protein kinase in rabbit fast twitch skeletal muscle. J Biol Chem. 2005 Feb 25;280(8):7147-55. DOI:10.1074/jbc.M413574200 |
11. Cho Y, Tachibana S, Hazen BC, Moresco JJ, Yates JR 3rd, Kok B, Saez E, Ross RS, Russell AP, and Kralli A. Perm1 regulates CaMKII activation and shapes skeletal muscle responses to endurance exercise training. Mol Metab. 2019 May;23:88-97. DOI:10.1016/j.molmet.2019.02.009 |
12. Morales-Alamo D, Ponce-González JG, Guadalupe-Grau A, Rodríguez-García L, Santana A, Cusso R, Guerrero M, Dorado C, Guerra B, and Calbet JA. Critical role for free radicals on sprint exercise-induced CaMKII and AMPKα phosphorylation in human skeletal muscle. J Appl Physiol (1985). 2013 Mar 1;114(5):566-77. DOI:10.1152/japplphysiol.01246.2012 |
13. Huang B, Yang CS, Wojton J, Huang NJ, Chen C, Soderblom EJ, Zhang L, and Kornbluth S. Metabolic control of Ca2+/calmodulin-dependent protein kinase II (CaMKII)-mediated caspase-2 suppression by the B55β/protein phosphatase 2A (PP2A). J Biol Chem. 2014 Dec 26;289(52):35882-90. DOI:10.1074/jbc.M114.585844 |
14. Flück M, Waxham MN, Hamilton MT, and Booth FW. Skeletal muscle Ca(2+)-independent kinase activity increases during either hypertrophy or running. J Appl Physiol (1985). 2000 Jan;88(1):352-8. DOI:10.1152/jappl.2000.88.1.352 |
15. Flück M, Booth FW, and Waxham MN. Skeletal muscle CaMKII enriches in nuclei and phosphorylates myogenic factor SRF at multiple sites. Biochem Biophys Res Commun. 2000 Apr 13;270(2):488-94. DOI:10.1006/bbrc.2000.2457 |
16. Sacchetto R, Bovo E, Salviati L, Damiani E, and Margreth A. Glycogen synthase binds to sarcoplasmic reticulum and is phosphorylated by CaMKII in fast-twitch skeletal muscle. Arch Biochem Biophys. 2007 Mar 1;459(1):115-21. DOI:10.1016/j.abb.2006.11.004 |
17. Aversa Z, Alamdari N, Castillero E, Muscaritoli M, Rossi Fanelli F, and Hasselgren PO. CaMKII activity is reduced in skeletal muscle during sepsis. J Cell Biochem. 2013 Jun;114(6):1294-305. DOI:10.1002/jcb.24469 |
18. Gaertner TR, Kolodziej SJ, Wang D, Kobayashi R, Koomen JM, Stoops JK, and Waxham MN. Comparative analyses of the three-dimensional structures and enzymatic properties of alpha, beta, gamma and delta isoforms of Ca2+-calmodulin-dependent protein kinase II. J Biol Chem. 2004 Mar 26;279(13):12484-94. DOI:10.1074/jbc.M313597200 |
19. Coultrap SJ, Barcomb K, and Bayer KU. A significant but rather mild contribution of T286 autophosphorylation to Ca2+/CaM-stimulated CaMKII activity. PLoS One. 2012;7(5):e37176. DOI:10.1371/journal.pone.0037176 |
20. Coultrap SJ, Zaegel V, and Bayer KU. CaMKII isoforms differ in their specific requirements for regulation by nitric oxide. FEBS Lett. 2014 Dec 20;588(24):4672-6. DOI:10.1016/j.febslet.2014.10.039 |