The heart arrhythmia-linked D130G calmodulin mutation causes premature inhibitory autophosphorylation of CaMKII

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The Ca2+/calmodulin (CaM)-dependent kinase II (CaMKII) is well known for transmitting Ca2+-signals, which leads to a multitude of physiological responses. Its functionality is believed to involve CaMKII holoenzyme dynamics where trans-autophosphorylation of the crucial phosphorylation site, T286 occurs. Phosphorylation of this site does not occur when stimulated exclusively with the arrhythmia associated D130G mutant form of CaM in vitro. Here, we present evidence that the loss-of-CaMKII function correlates with premature phosphorylation of its inhibitory phosphosite T306 in CaMKIIα and T307 in CaMKIIδ as this site was up to 20-fold more phosphorylated in the presence of D130G CaM compared to wildtype CaM. Indeed, changing this phosphosite to a non-phosphorylatable alanine reversed the inhibitory effect of D130G both in vitro and in live cell experiments. In addition, several phosphosites with so far undescribed functions directing the Ca2+-sensitivity of the CaMKII sensor were also affected by the presence of the D130G mutation implicating a role of several additional autophosphosites (besides T286 and T306/T307) so far not known to regulate CaMKII Ca2+ sensitivity. Furthermore, we show that introducing a D130G mutation in the CALM2 gene of the P19CL6 pluripotent mouse embryonic carcinoma cell line using CRISPR/Cas9 decreased the spontaneous beat frequency compared to wildtype cells when differentiated into cardiomyocytes supporting an alteration of cardiomyocyte physiology caused by this point mutation. In conclusion, our observations shed for the first time light on how the D130G CaM mutation interferes with the function of CaMKII and how it affects the beating frequency of cardiomyocyte-like cells.

Original languageEnglish
Article number119119
JournalBiochimica et Biophysica Acta - Molecular Cell Research
Issue number12
Number of pages9
Publication statusPublished - 2021

Bibliographical note

Publisher Copyright:
© 2021 The Author(s)

    Research areas

  • Calcium, Calmodulin, CaMKII, Cardiomyocyte, CRISPR/Cas9, Heart failure, Phosphorylation

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