Reciprocal activation within a kinase effector complex: A mechanism for the persistence of molecular memory

Synaptic connections in neuronal circuits change in response to neuronal activity patterns. This can induce a persistent change in the efficacy of synaptic transmission, a phenomenon known as synaptic plasticity. One form of plasticity, long-term potentiation (LTP) has been extensively studied as the cellular basis of memory. In LTP, the potentiated synaptic transmission persists along with structural changes in the synapses. Many studies have sought to identify the "memory molecule" or the "molecular engram". Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) is probably the most well-studied candidate for the memory molecule. However, consensus has not yet been reached on a very basic aspect: how CaMKII is regulated during LTP. Here, I propose a new model of CaMKII regulation: reciprocal activation within a kinase effector complex (RAKEC) that is made between CaMKII and its effector protein, which is mediated by a persistent interaction between CaMKII and a pseudosubstrate sequence on T-lymphoma invasion and metastasis protein 1 (Tiam1), resulting in reciprocal activation of these two molecules. Through the RAKEC mechanism, CaMKII can maintain memory as biochemical activity in a synapse-specific manner. In this review, the detailed mechanism of the RAKEC and its expansion for the maintenance of LTP is described

Molecular cloning and expression profile of Xenopus calcineurin A subunit11The nucleotide sequence of XCnA has been deposited in DDBJ/DMBL/GenBank DNA database under the accession number AB037146.

AbstractWe have cloned a cDNA encoding a catalytic subunit of calcineurin (CnA) expressed in Xenopus oocytes. The deduced amino acid sequence indicates 96.3% and 96.8% identities with the mouse and human CnAα isoforms, respectively. Xenopus CnA (XCnA) RNA and protein are expressed as maternal and throughout development. Recombinant XCnA protein interacted with calmodulin in the presence of Ca2+. Deletion of calmodulin binding domain and auto-inhibitory domain revealed calcium independent phosphatase activity, thereby showing that XCnA is likely to be modulated by both calmodulin and calcium

CaMKII binds both substrates and activators at the active site [preprint]

Ca2+/calmodulin dependent protein kinase II (CaMKII) is a signaling protein that is required for long-term memory formation. Ca2+/CaM activates CaMKII by binding to its regulatory segment, thereby freeing the substrate binding site. Despite having a large variety of interaction partners, the specificity of CaMKII interactions have not been structurally well-characterized. One exceptional feature of this kinase is that interaction with specific binding partners persistently activates CaMKII. To address the molecular details of this, we solved X-ray crystal structures of the CaMKII kinase domain bound to four different binding partners that modulate CaMKII activity in different ways. We show that all four partners bind in the same manner across the substrate binding site. We generated a sequence alignment based on our structural observations, which revealed conserved interactions. Using biochemistry and molecular dynamics simulations, we propose a mechanistic model that persistent CaMKII activity is facilitated by high affinity binding partners, which compete with the regulatory segment to allow substrate phosphorylation

Changing the size of dendritic spines

Interactions between an enzyme kinase, an ion channel and cytoskeletal proteins maintain the structure of synapses involved in memory formation

ERK5 phosphorylates Kv4.2 and inhibits inactivation of the A-type current in PC12 cells

Extracellular signal-regulated kinase 5 (ERK5) regulates diverse physiological responses such as proliferation, differentiation, and gene expression. Previously, we demonstrated that ERK5 is essential for neurite outgrowth and catecholamine biosynthesis in PC12 cells and sympathetic neurons. However, it remains unclear how ERK5 regulates the activity of ion channels, which are important for membrane excitability. Thus, we examined the effect of ERK5 on the ion channel activity in the PC12 cells that overexpress both ERK5 and the constitutively active MEK5 mutant. The gene and protein expression levels of voltage-dependent Ca²⁺ and K⁺ channels were determined by RT-qPCR or Western blotting. The A-type K⁺ current was recorded using the whole-cell patch clamp method. In these ERK5-activated cells, the gene expression levels of voltage-dependent L- and P/Q-type Ca²⁺ channels did not alter, but the N-type Ca²⁺ channel was slightly reduced. In contrast, those of Kv4.2 and Kv4.3, which are components of the A-type current, were significantly enhanced. Unexpectedly, the protein levels of Kv4.2 were not elevated by ERK5 activation, but the phosphorylation levels were increased by ERK5 activation. By electrophysiological analysis, the inactivation time constant of the A-type current was prolonged by ERK5 activation, without changes in the peak current. Taken together, ERK5 inhibits an inactivation of the A-type current by phosphorylation of Kv4.2, which may contribute to the neuronal differentiation process

Synthesis and Characterization of Cell-Permeable Oligonucleotides Bearing Reduction-Activated Protecting Groups on the Internucleotide Linkages

Cell-permeable oligodeoxyribonucleotides (ODNs) bearing reduction-activated protecting groups were synthesized as oligonucleotide pro-drugs. Although these oligonucleotides were amenable to solid-phase DNA synthesis and purification, the protecting group on their phosphodiester moiety could be readily cleaved by nitroreductase and NADH. Moreover, these compounds exhibited good nuclease resistance against 3′-exonuclease and endonuclease and good stability in human serum. Fluorescein-labeled ODNs modified with reduction-activated protecting groups showed better cellular uptake compared with that of naked ODNs

Structural and Molecular Remodeling of Dendritic Spine Substructures during Long-Term Potentiation

Synapses store information by long-lasting modifications of their structure and molecular composition, but the precise chronology of these changes has not been studied at single-synapse resolution in real time. Here we describe the spatiotemporal reorganization of postsynaptic substructures during long-term potentiation (LTP) at individual dendritic spines. Proteins translocated to the spine in four distinct patterns through three sequential phases. In the initial phase, the actin cytoskeleton was rapidly remodeled while active cofilin was massively transported to the spine. In the stabilization phase, cofilin formed a stable complex with F-actin, was persistently retained at the spine, and consolidated spine expansion. In contrast, the postsynaptic density (PSD) was independently remodeled, as PSD scaffolding proteins did not change their amount and localization until a late protein synthesis-dependent third phase. Our findings show how and when spine substructures are remodeled during LTP and explain why synaptic plasticity rules change over time.RIKEN Brain Science InstituteNational Institutes of Health (U.S.) (Grant R01DA17310)Human Frontier Science Program (Strasbourg, France)Paul and Anne Punzak Marcus Fun

Activity-Dependent Dendritic Arborization Mediated by CaM-Kinase I Activation and Enhanced CREB-Dependent Transcription of Wnt-2

Members of the Wnt signaling family are important mediators of numerous developmental events, including activity-dependent dendrite development, but the pathways regulating expression and secretion of Wnt in response to neuronal activity are poorly defined. Here, we identify an NMDA receptor-mediated, Ca 2+-dependent signaling pathway that couples neuronal activity to dendritic arborization through enhanced Wnt synthesis and secretion. Activity-dependent dendritic outgrowth and branching in cultured hippocampal neurons and slices is mediated through activation by CaM-dependent protein kinase kinase (CaMKK) of the membrane-associated γ isoform of CaMKI. Downstream effectors of CaMKI include the MAP-kinase pathway of Ras/MEK/ERK and the transcription factor CREB. A serial analysis of chromatin occupancy screen identified Wnt-2 as an activity-dependent CREB-responsive gene. Neuronal activity enhances CREB-dependent transcription of Wnt-2, and expression of Wnt-2 stimulates dendritic arborization. This novel signaling pathway contributes to dynamic remodeling of the dendritic architecture in response to neuronal activity during development

Activity-Dependent Synaptogenesis: Regulation by a CaM-Kinase Kinase/CaM-Kinase I/βPIX Signaling Complex

Neuronal activity augments maturation of mushroom-shaped spines to form excitatory synapses, thereby strengthening synaptic transmission. We have delineated a Ca2+-signaling pathway downstream of the NMDA receptor that stimulates calmodulin-dependent kinase kinase (CaMKK) and CaMKI to promote formation of spines and synapses in hippocampal neurons. CaMKK and CaMKI form a multiprotein signaling complex with the guanine nucleotide exchange factor (GEF) βPIX and GIT1 that is localized in spines. CaMKI-mediated phosphorylation of Ser516 in βPIX enhances its GEF activity, resulting in activation of Rac1, an established enhancer of spinogenesis. Suppression of CaMKK or CaMKI by pharmacological inhibitors, dominant-negative (dn) constructs and siRNAs, as well as expression of the βPIX Ser516Ala mutant, decreases spine formation and mEPSC frequency. Constitutively-active Pak1, a downstream effector of Rac1, rescues spine inhibition by dnCaMKI or βPIX S516A. This activity-dependent signaling pathway can promote synapse formation during neuronal development and in structural plasticity