Modulation of distinct isoforms of L-type calcium channels by Gq-coupled receptors in Xenopus oocytes: Antagonistic effects of Gβγ and protein kinase C

L-type voltage dependent Ca (2+) channels (L-VDCCs; Cav 1.2) are crucial in cardiovascular physiology. In heart and smooth muscle, hormones and transmitters operating via Gq enhance L-VDCC currents via essential protein kinase C (PKC) involvement. Heterologous reconstitution studies in Xenopus oocytes suggested that PKC and Gq-coupled receptors increased L-VDCC currents only in cardiac long N-terminus (NT) isoforms of {alpha} 1C, whereas known smooth muscle short-NT isoforms were inhibited by PKC and Gq activators. We report a novel regulation of the long-NT {alpha} 1C isoform by G{beta}{gamma}. G{beta}{gamma} inhibited whereas a G{beta}{gamma} scavenger protein augmented the Gq- but not phorbol ester - mediated enhancement of channel activity, suggesting that G{beta}{gamma} acts upstream from PKC. In vitro binding experiments reveal binding of both G{beta}{gamma} and PKC to {alpha} 1C-NT. However, PKC modulation was not altered by mutations of multiple potential phosphorylation cites in the NT, and was attenuated by a mutation of C-terminally located serine S1928. The insertion of exon 9a in intracelular loop 1 rendered the short-NT {alpha} 1C sensitive to PKC stimulation and to G{beta}{gamma} scavenging. Our results suggest a complex antagonistic interplay between Gq-activated PKC and G{beta}{gamma} in regulation of L-VDCC, in which multiple cytosolic segments of {alpha} 1C are involved

Protein kinase C enhances plasma membrane expression of cardiac L-type calcium channel, Ca(V)1.2

L-type-voltage-dependent Ca(2+) channels (L-VDCCs; CaV1.2, α(1C)), crucial in cardiovascular physiology and pathology, are modulated via activation of G-protein-coupled receptors and subsequently protein kinase C (PKC). Despite extensive study, key aspects of the mechanisms leading to PKC-induced Ca(2+) current increase are unresolved. A notable residue, Ser1928, located in the distal C-terminus (dCT) of α(1C) was shown to be phosphorylated by PKC. Ca(V)1.2 undergoes posttranslational modifications yielding full-length and proteolytically cleaved CT-truncated forms. We have previously shown that, in Xenopus oocytes, activation of PKC enhances α(1C) macroscopic currents. This increase depended on the isoform of α1C expressed. Only isoforms containing the cardiac, long N-terminus (L-NT), were upregulated by PKC. Ser1928 was also crucial for the full effect of PKC. Here we report that, in Xenopus oocytes, following PKC activation the amount of α(1C) protein expressed in the plasma membrane (PM) increases within minutes. The increase in PM content is greater with full-length α(1C) than in dCT-truncated α(1C), and requires Ser1928. The same was observed in HL-1 cells, a mouse atrium cell line natively expressing cardiac α(1C), which undergoes the proteolytic cleavage of the dCT, thus providing a native setting for exploring the effects of PKC in cardiomyocytes. Interestingly, activation of PKC preferentially increased the PM levels of full-length, L-NT α(1C). Our findings suggest that part of PKC regulation of Ca(V)1.2 in the heart involves changes in channel's cellular fate. The mechanism of this PKC regulation appears to involve the C-terminus of α(1C), possibly corroborating the previously proposed role of NT-CT interactions within α(1C)

Comorbidities and Lack of Blood Transfusion May Negatively Affect Maternal Outcomes of Women with Obstetric Hemorrhage Treated with NASG

The Non-Pneumatic Anti-Shock Garment (NASG) is a first-aid device to reduce mortality from severe obstetric hemorrhage, the leading cause of maternal mortality globally. We sought to evaluate patient characteristics associated with mortality among a cohor

Expression levels of RGS7 and RGS4 proteins determine the mode of regulation of the G protein-activated K(+) channel and control regulation of RGS7 by G beta 5

Regulators of G protein signaling RGS4 and RGS7 accelerate the kinetics of K(+) channels (GIRKs) in the Xenopus oocyte system. Here, via quantitative analysis of RGS expression, we reveal biphasic effects of RGSs on GIRK regulation. At low concentrations, RGS4 inhibited basal GIRK activity, but stimulated it at high concentrations. RGS7, which is associated with the G protein subunit G beta 5, is regulated by G beta 5 by two distinct mechanisms. First, G beta 5 augments RGS7 activity, and second, it increases its expression. These dual effects resolve previous controversies regarding RGS4 and RGS7 function and indicate that they modulate signaling by mechanisms supplementary to their GTPase-activating protein activity