Biology subjects

Man, K. N. M.

Publications and source records attributed to Man, K. N. M..

4 recordsLinked to original sources

bioRxiv2022

α1 adrenergic receptor - PKC - Pyk2 - Src signaling boosts L-type Ca2+ channel Cav1.2 activity and long-term potentiation in rodents

The cellular mechanisms mediating norepinephrine functions in brain to result in behaviors are unknown. We identified the L-type Ca2+ channel (LTCC) CaV1.2 as a principal target for Gq- coupled 1-adrenergic receptors (ARs). 1AR signaling increased LTCC activity in hippocampal neurons. This regulation required PKC-mediated activation of the tyrosine kinases Pyk2 and, downstream, Src. Pyk2 and Src were associated with CaV1.2. In model neuroendocrine PC12 cells, stimulation of PKC induced tyrosine phosphorylation of CaV1.2, a modification abrogated by inhibition of Pyk2 and Src. Upregulation of LTCC activity by 1AR and formation of a signaling complex with PKC, Pyk2, and Src suggests that CaV1.2 is a central conduit for signaling by norepinephrine. Indeed, a form of hippocampal LTP in young mice requires both the LTCC and 1AR stimulation. Inhibition of Pyk2 and Src blocked this LTP, indicating that enhancement of CaV1.2 activity via 1AR - Pyk2 - Src signaling regulates synaptic strength.

neuroscience↗

bioRxiv2022

Half-calcified Calmodulin Promotes Basal Activity and Inactivation of the Calcium Channel CaV1.2.

The L-type Ca2+ channel CaV1.2 controls gene expression, cardiac contraction, and neuronal activity. Calmodulin (CaM) governs CaV1.2 open probability (Po) and Ca2+-dependent inactivation (CDI) but the mechanisms remain unclear. We identified a half Ca2+-saturated CaM species (Ca2/CaM) with Ca2+ bound solely at the third and fourth EF-hands (EF3 and EF4) under resting Ca2+ concentrations (50-100 nM) that constitutively pre-associates with CaV1.2 to promote Po and CDI. We present an NMR structure of a complex between the CaV1.2 IQ motif (residues 1644-1665) and Ca2/CaM12, a calmodulin mutant in which Ca2+ binding to EF1 and EF2 is completely disabled. The CaM12 N-lobe does not interact with the IQ motif. The CaM12 C-lobe bound two Ca2+ ions and formed close contacts with IQ residues I1654 and Y1657. I1654A and Y1657D mutations impaired CaM binding, CDI, and Po, as did disabling Ca2+ binding to EF3 and EF4 in the CaM34 mutant when compared to wildtype CaM. Accordingly, a previously unappreciated Ca2/CaM species promotes CaV1.2 Po and CDI identifying Ca2/CaM as an important mediator of Ca signaling.

biochemistry↗

bioRxiv2019

β-blockers augment L-type Ca2+ channel activity by target-selective spatially restricted β2AR-cAMP-PKA signaling in neurons

G protein-coupled receptors (GPCRs) transduce pleiotropic intracellular signals in mammalian cells. Here, we report that some antagonists of {beta} adrenergic receptors ({beta}ARs) such as {beta}-blocker carvedilol and alprenolol activate {beta}2AR at nanomolar concentrations, which promote G protein signaling and cAMP/PKA activity without action of G protein receptor kinases (GRKs). The cAMP/PKA signal is restricted within the local plasma membrane domain, leading to selectively enhance PKA-dependent augment of endogenous L-type calcium channel (LTCC) activity but not AMPA receptor in hippocampal neurons. Moreover, we have engineered a mutant {beta}2AR that lacks serine 204 and 207 in the catecholamine binding pocket. This mutant can be preferentially activated by carvedilol but not the orthosteric agonist isoproterenol. Carvedilol activates the mutant {beta}2AR in hippocampal neurons augmenting LTCC activity through cAMP/PKA signaling. Together, our study identifies a mechanism by which {beta}-blocker-dependent activation of GPCRs at low ligand concentrations promotes local cAMP/PKA signaling to selectively target membrane downstream effectors such as LTCC in neurons.

cell biology↗

bioRxiv2019

Alpha-Actinin-1 promotes activity of the L-type Ca2+ Channel CaV1.2

The L-type Ca2+ channel CaV1.2 governs gene expression, cardiac contraction, and neuronal activity. Binding of -actinin to the IQ motif of CaV1.2 supports its surface localization and postsynaptic targeting in neurons. We report a bi-functional mechanism that restricts CaV1.2 activity to its target sites. We solved separate NMR structures of the IQ motif (residues 1646-1664) bound to -actinin-1 and to apo-calmodulin (apoCaM). The CaV1.2 K1647A and Y1649A mutations, which impair -actinin-1 but not apoCaM binding, but not the F1658A and K1662E mutations, which impair apoCaM but not -actinin-1 binding, decreased single channel open probability, gating charge movement, and its coupling to channel opening. Thus, -actinin recruits CaV1.2 to defined surface regions and simultaneously boosts its open probability so that CaV1.2 is mostly active when appropriately localized.

molecular biology↗