IndraLab
Statements
"A stable interaction between ?(C)-camkii and the intracellular loop between domains 1 and 2 of na(v)1.5 was observed. This region was also phosphorylated by ?(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-?(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."
"Among the sites identified, only six were previously suggested to be the targets for specific kinases using in silico and/or in vitro analyses: S36 and S525 were attributed to the regulation by PKA; S484 and S664 were assigned to the serum- and glucocorticoid-inducible kinase 3 (SGK3); and S516 and S571 were ascribed to CaMKII (reviewed in Marionneau and Abriel, 2015). In marked contrast, several previously described phosphorylation sites were not detected in the present study, including the PKA-dependent S528, the CaMKII-associated T594, the PKC-dependent S1506, the adenosine monophosphate–activated protein kinase (AMPK)–dependent T101 (Liu et al., 2019), and the six Fyn-dependent tyrosines (Ahern et al., 2005; Iqbal et al., 2018).|The simplest interpretation of these findings is that these three phosphorylation clusters, at positions S457-S460, S483-T486, and S664-S671, are likely involved in regulating the basal and/or gating properties of native cardiac NaV1.5 channels. Conversely, the other phosphorylation sites, with lower stoichiometries, may play spatially or temporally distinct roles in the physiological or more pathophysiological regulation of channel expression or gating. | Remarkably, this MS analysis also revealed that the vast majority of identified phosphorylation sites (at least 26) are clustered, suggesting concomitant phosphorylation and roles in regulating channel expression and/or function. Unexpectedly, however, except for S664, S667, and S671, no apparent effects of phosphomimetic or phosphosilent mutations were observed on heterologously expressed (in HEK-293 cells) NaV1.5"