Biochemical Analysis of AKAP-Anchored PKA Signaling Complexes

Byrne, Dominic P.; Omar, Mitchell H.; Kennedy, Eileen J.; Eyers, Patrick A.; Scott, John D.

doi:10.1007/978-1-0716-2245-2_19

Cited by 4 publications

(2 citation statements)

References 85 publications

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“…The signal transduction and sensitize recycling of adrenaline receptors [9], regulation of the cardiac hypertrophy signal [10], and the coupling of excitation-contraction, relaxation, and intracellular calcium circulation [10][11][12] in cardiac cells are involved in ensuring the normal function and morphology of the heart. In arrhythmia, AKAP150 acts in two ways: it intensifies arrhythmia caused by calcium channel-related gene mutation [13], and it may have a therapeutic effect on arrhythmia caused by potassium channel-related gene mutation [14,15]. β1-adrenergic receptor (β-AR) stimulation in vitro and in vivo increased the expression and activity of matrix metalloproteinases 2 and 9 (MMP-2 and MMP-9) in cardiac myocytes [16]; it also induced cardiomyocyte apoptosis [7,17,18].…”

Section: Introductionmentioning

confidence: 99%

Metoprolol Mitigates Ischemic Heart Remodeling and Fibrosis by Increasing the Expression of AKAP5 in Ischemic Heart

Zhu

Wang

et al. 2022

Oxidative Medicine and Cellular Longevity

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The harm of heart failure mainly causes patients to develop dyspnea, fatigue, fluid retention, and other symptoms, which impair patients’ activity tolerance and lead to a dramatic decrease in patients’ quality of life. The purpose of this study was to verify whether metoprolol regulates AKAP5 expression and test the role of AKAP5 postinjury in mitigating cardiac infarction-associated tissue remodeling and fibrosis. Sprague-Dawley (SD) rats underwent coronary artery ligation (CAL), which was followed immediately with metoprolol daily. And western blot and coimmunoprecipitation experiments were performed to detect the expression of related proteins in the sham-operated group, model group, and drug-treated group. HW/BW ratio and cardiac expression of COL1 and COL3 were increased in rats following CAL compared with shams. Treatment with metoprolol postinjury was associated with a decrease in HW/BW ratio and COL1/COL3 expression compared to uncontrolled rats. CAL resulted in decreased cardiac AKAP5 expression compared to the control group, while metoprolol treatment restored levels compared to baseline shams. Cardiac expression levels of NFATc3/p-NFATc3 and GATA4 were modest at baseline and increased with injury, whereas metoprolol suppressed gene expression to below injury-associated changes. Immunoprecipitation indicated that AKAP5 could bind and regulate PP2B. In summary, we know that metoprolol alleviates ischemic cardiac remodeling and fibrosis, and the mechanism of alleviating remodeling may improve cardiac AKAP5 expression and AKAP5-PP2B interaction.

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Section: Introductionmentioning

confidence: 99%

Metoprolol Mitigates Ischemic Heart Remodeling and Fibrosis by Increasing the Expression of AKAP5 in Ischemic Heart

Zhu

Wang

et al. 2022

Oxidative Medicine and Cellular Longevity

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“…Forskolin/IBMX-mediated disassembly of SPHKAP complexes still occurred when PKA catalytic activity was blocked (Supplementary Fig. 7f–i ), suggesting that supraphysiological elevation of cAMP mediated by these agents 32 rather than PKA-dependent phosphorylation causes SPHKAP-RI complexes to decluster in neurons. Taken together, our data suggest that when RI is associated with an AKAP, such as SPHKAP, it can acquire physical properties distinct from previously described LLPS-derived RI condensates 28 .…”

Section: Resultsmentioning

confidence: 99%

Neuronal ER-plasma membrane junctions couple excitation to Ca2+-activated PKA signaling

Vierra,

Ribeiro-Silva,

Kirmiz

et al. 2023

Nat Commun

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Junctions between the endoplasmic reticulum (ER) and the plasma membrane (PM) are specialized membrane contacts ubiquitous in eukaryotic cells. Concentration of intracellular signaling machinery near ER-PM junctions allows these domains to serve critical roles in lipid and Ca2+ signaling and homeostasis. Subcellular compartmentalization of protein kinase A (PKA) signaling also regulates essential cellular functions, however, no specific association between PKA and ER-PM junctional domains is known. Here, we show that in brain neurons type I PKA is directed to Kv2.1 channel-dependent ER-PM junctional domains via SPHKAP, a type I PKA-specific anchoring protein. SPHKAP association with type I PKA regulatory subunit RI and ER-resident VAP proteins results in the concentration of type I PKA between stacked ER cisternae associated with ER-PM junctions. This ER-associated PKA signalosome enables reciprocal regulation between PKA and Ca2+ signaling machinery to support Ca2+ influx and excitation-transcription coupling. These data reveal that neuronal ER-PM junctions support a receptor-independent form of PKA signaling driven by membrane depolarization and intracellular Ca2+, allowing conversion of information encoded in electrical signals into biochemical changes universally recognized throughout the cell.

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Classification of Cushing's syndrome PKAc mutants based upon their ability to bind PKI

Omar

Kihiu

Byrne

et al. 2023

Biochemical Journal

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Cushing’s syndrome is an endocrine disorder caused by excess production of the stress hormone cortisol. Precision medicine strategies have identified single allele mutations within the PRKACA gene that drive adrenal Cushing’s syndrome. These mutations promote perturbations in the catalytic core of protein kinase A (PKAc) that impair autoinhibition by regulatory subunits and compartmentalization via recruitment into AKAP signaling islands. PKAcL205R is found in approximately 45% of patients, whereas PKAcE31V, PKAcW196R and L198insW and C199insV insertion mutants are less prevalent. Mass spectrometry, cellular, and biochemical data indicate that Cushing’s PKAc variants fall into two categories: those that interact with the heat-stable protein kinase inhibitor PKI, and those that do not. In vitro activity measurements show that wild-type PKAc and W196R activities are strongly inhibited by PKI (IC50 < 1 nM). In contrast, PKAcL205R activity is not blocked by the inhibitor. Immunofluorescent analyses show that the PKI-binding variants wild-type PKAc, E31V, and W196R are excluded from the nucleus and protected against proteolytic processing. Thermal stability measurements reveal that upon co-incubation with PKI and metal-bound nucleotide, the W196R variant tolerates melting temperatures 10ºC higher than PKAcL205. Structural modeling maps PKI-interfering mutations to a ~20 Å diameter area at the active site of the catalytic domain that interfaces with the pseudosubstrate of PKI. Thus, Cushing’s kinases are individually controlled, compartmentalized, and processed through their differential association with PKI.

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Biochemical Analysis of AKAP-Anchored PKA Signaling Complexes

Cited by 4 publications

References 85 publications

Metoprolol Mitigates Ischemic Heart Remodeling and Fibrosis by Increasing the Expression of AKAP5 in Ischemic Heart

Metoprolol Mitigates Ischemic Heart Remodeling and Fibrosis by Increasing the Expression of AKAP5 in Ischemic Heart

Neuronal ER-plasma membrane junctions couple excitation to Ca2+-activated PKA signaling

Classification of Cushing's syndrome PKAc mutants based upon their ability to bind PKI

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