Ca2+/calmodulin-dependent protein kinase II (CaMKII) regulates cardiac sodium channel NaV1.5 gating by multiple phosphorylation sites.

Published

Journal Article

The cardiac Na(+) channel Na(V)1.5 current (I(Na)) is critical to cardiac excitability, and altered I(Na) gating has been implicated in genetic and acquired arrhythmias. Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) is up-regulated in heart failure and has been shown to cause I(Na) gating changes that mimic those induced by a point mutation in humans that is associated with combined long QT and Brugada syndromes. We sought to identify the site(s) on Na(V)1.5 that mediate(s) the CaMKII-induced alterations in I(Na) gating. We analyzed both CaMKII binding and CaMKII-dependent phosphorylation of the intracellularly accessible regions of Na(V)1.5 using a series of GST fusion constructs, immobilized peptide arrays, and soluble peptides. A stable interaction between δ(C)-CaMKII and the intracellular loop between domains 1 and 2 of Na(V)1.5 was observed. This region was also phosphorylated by δ(C)-CaMKII, specifically at the Ser-516 and Thr-594 sites. Wild-type (WT) and phosphomutant hNa(V)1.5 were co-expressed with GFP-δ(C)-CaMKII in HEK293 cells, and I(Na) was recorded. As observed in myocytes, CaMKII shifted WT I(Na) availability to a more negative membrane potential and enhanced accumulation of I(Na) into an intermediate inactivated state, but these effects were abolished by mutating either of these sites to non-phosphorylatable Ala residues. Mutation of these sites to phosphomimetic Glu residues negatively shifted I(Na) availability without the need for CaMKII. CaMKII-dependent phosphorylation of Na(V)1.5 at multiple sites (including Thr-594 and Ser-516) appears to be required to evoke loss-of-function changes in gating that could contribute to acquired Brugada syndrome-like effects in heart failure.

Full Text

Duke Authors

Cited Authors

  • Ashpole, NM; Herren, AW; Ginsburg, KS; Brogan, JD; Johnson, DE; Cummins, TR; Bers, DM; Hudmon, A

Published Date

  • June 8, 2012

Published In

Volume / Issue

  • 287 / 24

Start / End Page

  • 19856 - 19869

PubMed ID

  • 22514276

Pubmed Central ID

  • 22514276

Electronic International Standard Serial Number (EISSN)

  • 1083-351X

Digital Object Identifier (DOI)

  • 10.1074/jbc.M111.322537

Language

  • eng

Conference Location

  • United States