CaMKIIβ Association with the Actin Cytoskeleton Is Regulated by Alternative Splicing

Abstract

The Ca 2؉ /calmodulin (CaM)-dependent protein kinase II (CaMKII)␤ has morphogenic functions in neurons not shared by the ␣ isoform. CaMKII␤ contains three exons (v1, v3, and v4) not present in the CaMKII␣ gene, and two of these exons (v1 and v4) are subject to differential alternative splicing. We show here that CaMKII␤, but not ␣, mediated bundling of F-actin filaments in vitro. Most importantly, inclusion of exon v1 was required for CaMKII␤ association with the F-actin cytoskeleton within cells. CaMKII␤e, which is the dominant variant around birth and lacks exon v1 sequences, failed to associate with F-actin. By contrast, CaMKII␤, which instead lacks exon v4, associated with F-actin as full-length CaMKII␤. Previous studies with CaMKII␤ mutants have indicated a role of nonstimulated kinase activity in enhancing dendritic arborization. Here, we show that F-actin-targeted CaMKII␤, but not ␣, was able to phosphorylate actin in vitro even by nonstimulated basal activity in absence of Ca 2؉ /CaM. In rat pancreatic islets and in skeletal muscle, the actin-associated CaMKII␤ and ␤M were the predominant variants, respectively. Thus, cytoskeletal targeting may mediate functions of CaMKII␤ variants also outside the nervous system. INTRODUCTIONChanges in synaptic connectivity between neurons are widely thought to underlie higher brain functions such as learning and memory and are also important during development. Long-lasting changes in connectivity are often associated with morphological plasticity in structure or number of synaptic connections (for reviews, see Huntley et al., 2002;McGee and Bredt, 2003;Lamprecht and LeDoux, 2004;Segal, 2005). The ␣ isoform of Ca 2ϩ /calmodulin(CaM)-dependent protein kinase II (CaMKII) has been studied extensively because of its prominent role in regulating the strength of individual synaptic connections (for examples, see Malinow et al., 1989;Silva et al., 1992;Giese et al., 1998; for reviews, see Malenka and Nicoll, 1999;Lisman et al., 2002;Griffith, 2004). Much less is known about the other major brain isoform, CaMKII␤. However, CaMKII␤ has specific morphogenic functions in regulating dendritic arborization and synapse density not shared by the ␣ isoform (Fink et al., 2003). This isoform specificity is thought to be mediated by the specific binding of CaMKII␤, but not ␣ to F-actin (Shen et al., 1998;Fink et al., 2003).The four CaMKII isoforms encoded by different genes (␣, ␤, ␥, and ␦) are highly homologous to each other and can phosphorylate and regulate a variety of substrate proteins in response to Ca 2ϩ signals (for reviews, see Soderling et al., 2001;Hudmon and Schulman, 2002;Lisman et al., 2002;Colbran and Brown, 2004). The multimeric CaMKII holoenzymes are composed of 12 subunits (Kolodziej et al., 2000;Morris and Torok, 2001;Rosenberg et al., 2005;Rosenberg et al., 2006) of a single isoform or combinations of different isoforms (Bayer et al., 1998;Shen et al., 1998;Brocke et al., 1999;Lantsman and Tombes, 2005). All CaMKII isoforms contain an N-terminal kinase domain and a ...

The role of CaMKII as an F-actin-bundling protein crucial for maintenance of dendritic spine structure

“…Again, this activity was associated with CaMKII␤; homo-meric CaMKII␣ failed to show such activity. This assay, in conjunction with the cosedimentation assay, establishes that CaMKII is an F-actin-bundling protein, consistent with a recent report (20).…”

“…CaMKII accumulates on the dendritic spine upon neuronal stimulation (19,20,(29)(30)(31). Because CaMKII␣ does not bundle F-actin but still shows activity dependent accumulation, the accumulation is independent of F-actin binding (19).…”

“…Because CaMKII␣ does not bundle F-actin but still shows activity dependent accumulation, the accumulation is independent of F-actin binding (19). Instead, selfassociation of CaMKII (20,30) or binding with the NR2B subunit of NMDA receptors (31), densin-180 (32,33), and/or ␣-actinin (32-34) may play a role in such activity-dependent synaptic delivery of CaMKII.…”

“…This knowledge led us and others to speculate that CaMKII may serve not only as a signal transduction molecule but also as a structural element at synapses (2,14). In relation to this speculation, it has been demonstrated that CaMKII binds to F-actin through a specific sequence present in the ␤ subunit (16)(17)(18)(19)(20), but this interaction has been considered to anchor CaMKII to the postsynaptic sites. However, considering that CaMKII is an oligomeric protein composed of 12-14 subunits with rotational symmetry (2), it is possible that one oligomer can simultaneously bind to multiple F-actin, thereby acting as an F-actin-bundling protein at postsynaptic sites.…”

CaMKII Plays a Nonenzymatic Role in Hippocampal Synaptic Plasticity and Learning by Targeting CaMKII to Synapses

“…Mainly due to these different actin-binding properties, ␣CaMKII and ␤CaMKII were shown to have opposing effects on synaptic strength in cultured neurons (Thiagarajan et al, 2002). Interestingly, ␤CaMKII not only binds to actin, but is also capable of bundling actin in a kinase-independent manner (O'Leary et al, 2006;Okamoto et al, 2007;Sanabria et al, 2009). This nonenzymatic bundling feature is likely achieved by single CaMKII oligomers binding to multiple actin filaments.…”