Search

The transcription factors cAMP-responsive element binding protein (CREB) and cAMP-responsive element modulator (CREM) regulate gene transcription in response to elevated cAMP levels. The Crem isoform inducible cAMP early repressor (Icer) is transcribed by the internal promoter P2 as a critical regulator of multiple cellular processes. Here, we describe a novel inducible Crem isoform, sma//lcer (sm/cer regulated by a newly identified promoter (P6). Ch revealed binding of CREB to P6 in human and mouse myocardium. P6 activity was induced by constitutive active CREB or stimulation of adenylyl cyclase. In mice, smlcer mRNA was ubiquitously expressed and transiently induced by β-adrenoceptor stimulation e.g., in heart and lung. SmICER repressed both basal a cAMP-induced activities of P6 and P2 promoters. Stimulation of adenylyl cyclase induced P2 and P6 in cell type-specific manner. Alternative translational star sites resulted in three different smICER proteins linked to increased apoptosis sensitivity. In conclusion, the Crem gene provides two distinct and mutually controlled mechanisms of a cAMP-dependent induction of transcriptional repressors. Our results suggest not only that smICER is a novel regulator of cAM mediated gene regulation, but also emphasize that biological effects that have been ascribed solely to ICER, should be revised with regard to smICER. [ABSTRACT FROM AUTHOR]

The activity of protein phosphatase 2A (PP2A) is determined by the expression and localization of the regulatory B-subunits. PP2A-B56α is the dominant isoform of the B'-family in the heart. Its role in regulating the cardiac response to β-adrenergic stimulation is not yet fully understood. We therefore generated mice deficient in B56α to test the functional cardiac effects in response to catecholamine administration versus corresponding WT mice. We found the decrease in basal PP2A activity in hearts of KO mice was accompanied by a counterregulatory increase in the expression of B0 subunits (β and γ) and higher phosphorylation of sarcoplasmic reticulum Ca2+ regulatory and myofilament proteins. The higher phosphorylation levels were associated with enhanced intraventricular pressure and relaxation in catheterized KO mice. In contrast, at the cellular level, we detected depressed Ca2+ transient and sarcomere shortening parameters in KO mice at basal conditions. Consistently, the peak amplitude of the L-type Ca2+ current was reduced and the inactivation kinetics of ICaL were prolonged in KO cardiomyocytes. However, we show β-adrenergic stimulation resulted in a comparable peak amplitude of Ca2+ transients and myocellular contraction between KO and WT cardiomyocytes. Therefore, we propose higher isoprenaline-induced Ca2+ spark frequencies might facilitate the normalized Ca2+ signaling in KO cardiomyocytes. In addition, the application of isoprenaline was associated with unchanged L-type Ca2+ current parameters between both groups. Our data suggest an important influence of PP2A-B56α on the regulation of Ca2+ signaling and contractility in response to β-adrenergic stimulation in the myocardium. [ABSTRACT FROM AUTHOR]

Protein phosphatase 2A (PP2A) is a family of multifunctional serine/threonine phosphatases consisting of a catalytic C, a structural A, and a regulatory B subunit. The substrate and therefore the functional specificity of PP2A are determined by the assembly of the enzyme complex with the appropriate regulatory B subunit families, namely B55, B56, PR72, or PR93/ PR110. It has been suggested that additional levels of regulating PP2A function may result from the phosphorylation of B56 isoforms. In this study, we identified a novel phosphorylation site at Ser41 of B56α. This phosphoamino acid residue was efficiently phosphorylated in vitro by PKCα. We detected a 7-fold higher phosphorylation of B56α in failing human hearts compared with nonfailing hearts. Purified PP2A dimeric holoenzyme (subunits C and A) was able to dephosphorylate PKCα-phosphorylated B56α. The potency of B56α for PP2A inhibition was markedly increased by PKCα phosphorylation. PP2A activity was also reduced in HEK293 cells transfected with a B56α mutant, where serine 41 was replaced by aspartic acid, which mimics phosphorylation. More evidence for a functional role of PKCα-dependent phosphorylation of B56α was derived from Fluo-4 fluorescence measurements in phenylephrine-stimulated Flp293 cells. The endoplasmic reticulum Ca2+ release was increased by 23% by expression of the pseudophosphorylated form compared with wild-type B56α. Taken together, our results suggest that PKCα can modify PP2A activity by phosphorylation of B56α at Ser41. This interplay between PKCα and PP2A represents a new mechanism to regulate important cellular functions like cellular Ca2+ homeostasis. [ABSTRACT FROM AUTHOR]

Sphingosine 1-phosphate (S1P) is a lysosphingolipid with anti-atherogenic properties, but mechanisms underlying its effects remain unclear. We here investigated atherosclerosis development in cholesterol-rich diet-fed LDL receptor-deficient mice with high or low overexpression levels of S1P receptor type 1 (S1P1) in macrophages. S1P1-overexpressing macrophages showed increased activity of transcription factors PU.1, IRF8, and LXR and were skewed towards a M2-distinct phenotype characterized by enhanced production of IL-10, IL-1RA, and IL-5, increased ATP-binding cassette transporter A1- and G1-dependent cholesterol efflux, increased expression of MerTK and efferocytosis, and reduced apoptosis due to elevated Bcl6 and MafB. A similar macrophage phenotype was observed in mice administered S1P1-selective agonist KRP203. Mechanistically, the enhanced PU.1, IRF8, and LXR activity in S1P1-overexpressing macrophages led to down-regulation of the cAMP-dependent protein kinase A and activation of the signaling cascade encompassing protein kinases Akt and mTOR complex 1 (mTORC1) as well as the late endosomal/lysosomal adaptor MAPK and mTOR activator 1 (Lamtor-1). Atherosclerotic lesions in aortic roots and brachiocephalic arteries were profoundly or moderately reduced in mice with high and low S1P1 overexpression in macrophages, respectively. We conclude that S1P1 signaling polarizes macrophages towards an anti-atherogenic functional phenotype and countervails the development of atherosclerosis in mice.

The activity of protein phosphatase 2A (PP2A) is determined by the expression and localization of the regulatory B-subunits. PP2A-B56α is the dominant isoform of the B'-family in the heart. Its role in regulating the cardiac response to β-adrenergic stimulation is not yet fully understood. We therefore generated mice deficient in B56α to test the functional cardiac effects in response to catecholamine administration versus corresponding WT mice. We found the decrease in basal PP2A activity in hearts of KO mice was accompanied by a counter-regulatory increase in the expression of B' subunits (β and γ) and higher phosphorylation of sarcoplasmic reticulum Ca 2+ regulatory and myofilament proteins. The higher phosphorylation levels were associated with enhanced intraventricular pressure and relaxation in catheterized KO mice. In contrast, at the cellular level, we detected depressed Ca 2+ transient and sarcomere shortening parameters in KO mice at basal conditions. Consistently, the peak amplitude of the L-type Ca 2+ current was reduced and the inactivation kinetics of I CaL were prolonged in KO cardiomyocytes. However, we show β-adrenergic stimulation resulted in a comparable peak amplitude of Ca 2+ transients and myocellular contraction between KO and WT cardiomyocytes. Therefore, we propose higher isoprenaline-induced Ca 2+ spark frequencies might facilitate the normalized Ca 2+ signaling in KO cardiomyocytes. In addition, the application of isoprenaline was associated with unchanged L-type Ca 2+ current parameters between both groups. Our data suggest an important influence of PP2A-B56α on the regulation of Ca 2+ signaling and contractility in response to β-adrenergic stimulation in the myocardium., Competing Interests: Conflict of interest The authors declare that they have no conflicts of interest with the contents of this article., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)