Agonist-triggered downregulation of b-adrenergic receptors (ARs) constitutes vital negative feedback to prevent cellular overexcitation. Here, we report a novel downregulation of b 2 AR signaling highly specific for Ca v 1.2. We find that b 2 -AR binding to Ca v 1.2 residues 1923-1942 is required for b-adrenergic regulation of Ca v 1.2. Despite the prominence of PKA-mediated phosphorylation of Ca v 1.2 S1928 within the newly identified b 2 AR binding site, its physiological function has so far escaped identification. We show that phosphorylation of S1928 displaces the b 2 AR from Ca v 1.2 upon b-adrenergic stimulation rendering Ca v 1.2 refractory for several minutes from further b-adrenergic stimulation. This effect is lost in S1928A knock-in mice. Although AMPARs are clustered at postsynaptic sites like Ca v 1.2, b 2 AR association with and regulation of AMPARs do not show such dissociation. Accordingly, displacement of the b 2 AR from Ca v 1.2 is a uniquely specific desensitization mechanism of Ca v 1.2 regulation by highly localized b 2 AR/cAMP/PKA/ S1928 signaling. The physiological implications of this mechanism are underscored by our finding that LTP induced by prolonged theta tetanus (PTT-LTP) depends on Ca v 1.2 and its regulation by channel-associated b 2 AR.

Postsynaptic density protein-95 (PSD-95) is a central element of the postsynaptic architecture of glutamatergic synapses. PSD-95 mediates postsynaptic localization of AMPA receptors and NMDA receptors and plays an important role in synaptic plasticity. PSD-95 is released from postsynaptic membranes in response to Ca 2+ influx via NMDA receptors. Here, we show that Ca 2+ /calmodulin (CaM) binds at the N-terminus of PSD-95. Our NMR structure reveals that both lobes of CaM collapse onto a helical structure of PSD-95 formed at its N-terminus (residues 1-16). This N-terminal capping of PSD-95 by CaM blocks palmitoylation of C3 and C5, which is required for postsynaptic PSD-95 targeting and the binding of CDKL5, a kinase important for synapse stability. CaM forms extensive hydrophobic contacts with Y12 of PSD-95. The PSD-95 mutant Y12E strongly impairs binding to CaM and Ca 2+ -induced release of PSD-95 from the postsynaptic membrane in dendritic spines. Our data indicate that CaM binding to PSD-95 serves to block palmitoylation of PSD-95, which in turn promotes Ca 2+ -induced dissociation of PSD-95 from the postsynaptic membrane.

The L-type Ca2+ channel Cav1.2 controls multiple functions throughout the body including heart rate and neuronal excitability. It is a key mediator of fight-or-flight stress responses triggered by a signaling pathway involving β-adrenergic receptors (βARs), cyclic adenosine monophosphate (cAMP), and protein kinase A (PKA). PKA readily phosphorylates Ser1928 in Cav1.2 in vitro and in vivo, including in rodents and humans. However, S1928A knock-in (KI) mice have normal PKA-mediated L-type channel regulation in the heart, indicating that Ser1928 is not required for regulation of cardiac Cav1.2 by PKA in this tissue. We report that augmentation of L-type currents by PKA in neurons was absent in S1928A KI mice. Furthermore, S1928A KI mice failed to induce long-term potentiation in response to prolonged theta-tetanus (PTT-LTP), a form of synaptic plasticity that requires Cav1.2 and enhancement of its activity by the β2-adrenergic receptor (β2AR)–cAMP–PKA cascade. Thus, there is an unexpected dichotomy in the control of Cav1.2 by PKA in cardiomyocytes and hippocampal neurons.

Many postsynaptic proteins undergo palmitoylation, the reversible attachment of the fatty acid palmitate to cysteine residues, which influences trafficking, localization, and protein interaction dynamics. Both palmitoylation by palmitoyl acyl transferases (PAT) and depalmitoylation by palmitoyl-protein thioesterases (PPT) is regulated in an activity-dependent, localized fashion. Recently, palmitoylation has received attention for its pivotal contribution to various forms of synaptic plasticity, the dynamic modulation of synaptic strength in response to neuronal activity. For instance, palmitoylation and depalmitoylation of the central postsynaptic scaffold protein postsynaptic density-95 (PSD-95) is important for synaptic plasticity. Here, we provide a comprehensive review of studies linking palmitoylation of postsynaptic proteins to synaptic plasticity.

Despite the central role PSD-95 plays in anchoring postsynaptic AMPARs, how PSD-95 itself is tethered to postsynaptic sites is not well understood. Here we show that the F-actin binding protein α-actinin binds to the very N terminus of PSD-95. Knockdown (KD) of α-actinin phenocopies KD of PSD-95. Mutating lysine at position 10 or lysine at position 11 of PSD-95 to glutamate, or glutamate at position 53 or glutamate and aspartate at positions 213 and 217 of α-actinin, respectively, to lysine impairs, in parallel, PSD-95 binding to α-actinin and postsynaptic localization of PSD-95 and AMPARs. These experiments identify α-actinin as a critical PSD-95 anchor tethering the AMPAR-PSD-95 complex to postsynaptic sites.