A membrane-associated phosphoswitch in Rad controls adrenergic regulation of cardiac calcium channels

Arianne Papa, Pedro J. del Rivero Morfin, Bi Xing Chen, Lin Yang, Alexander N. Katchman, Sergey I. Zakharov, Guoxia Liu, Michael S. Bohnen, Vivian Zheng, Moshe Katz, Suraj Subramaniam, Joel A. Hirsch, Sharon Weiss, Nathan Dascal, Arthur Karlin, Geoffrey S. Pitt, Henry M. Colecraft, Manu Ben-Johny*, Steven O. Marx*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

1 Scopus citations

Abstract

The ability to fight or flee from a threat relies on an acute adrenergic surge that augments cardiac output, which is dependent on increased cardiac contractility and heart rate. This cardiac response depends on β-adrenergic–initiated reversal of the small RGK G protein Rad–mediated inhibition of voltage-gated calcium channels (CaV) acting through the Cavβ subunit. Here, we investigate how Rad couples phosphorylation to augmented Ca2+ influx and increased cardiac contraction. We show that reversal required phosphorylation of Ser272 and Ser300 within Rad’s polybasic, hydrophobic C-terminal domain (CTD). Phosphorylation of Ser25 and Ser38 in Rad’s N-terminal domain (NTD) alone was ineffective. Phosphorylation of Ser272 and Ser300 or the addition of 4 Asp residues to the CTD reduced Rad’s association with the negatively charged, cytoplasmic plasmalemmal surface and with CaVβ, even in the absence of CaVα, measured here by FRET. Addition of a posttranslationally prenylated CAAX motif to Rad’s C-terminus, which constitutively tethers Rad to the membrane, prevented the physiological and biochemical effects of both phosphorylation and Asp substitution. Thus, dissociation of Rad from the sarcolemma, and consequently from CaVβ, is sufficient for sympathetic upregulation of Ca2+ currents.

Original languageEnglish
Article numbere176943
JournalJournal of Clinical Investigation
Volume134
Issue number5
DOIs
StatePublished - 1 Mar 2024

Funding

FundersFunder number
National Institutes of HealthF31HL158232, P01 HL164319, R01 HL146149, R01 HL155377, R01 HL121253, R01 HL140934
National Institutes of Health
Office of the DirectorS10RR027050
Office of the Director
Washington University in St. Louis
United States-Israel Binational Science Foundation2021065, P30CA013696
United States-Israel Binational Science Foundation

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