N-glycans modulate Kv1.5 gating but have no effect on Kv1.4 gating

Abstract

Nerve and muscle action potential repolarization are produced and modulated by the regulated expression and activity of several types of voltage-gated K(+) (K(v)) channels. Here, we show that sialylated N-glycans uniquely impact gating of a mammalian Shaker family K(v) channel isoform, K(v)1.5, but have no effect on gating of a second Shaker isoform, K(v)1.4. Each isoform contains one potential N-glycosylation site located along the S1-S2 linker; immunoblot analyses verified that K(v)1.4 and K(v)1.5 were N-glycosylated. The conductance-voltage (G-V) relationships and channel activation rates for two glycosylation-site deficient K(v)1.5 mutants, K(v)1.5(N290Q) and K(v)1.5(S292A), and for wild-type K(v)1.5 expressed under conditions of reduced sialylation, were each shifted linearly by a depolarizing approximately 18 mV compared to wild-type K(v)1.5 activation. External divalent cation screening experiments suggested that K(v)1.5 sialic acids contribute to an external surface potential that modulates K(v)1.5 activation. Channel availability was unaffected by changes in K(v)1.5 glycosylation or sialylation. The data indicate that sialic acid residues attached to N-glycans act through electrostatic mechanisms to modulate K(v)1.5 activation. The sialic acids fully account for effects of N-glycans on K(v)1.5 gating. Conversely, K(v)1.4 gating was unaffected by changes in channel sialylation or following mutagenesis to remove the N-glycosylation site. Each phenomenon is unique for K(v)1 channel isoforms, indicating that sialylated N-glycans modulate gating of homologous K(v)1 channels through isoform-specific mechanisms. Such modulation is relevant to changes in action potential repolarization that occur as ion channel expression and glycosylation are regulated.

Sialic Acids Attached to O-Glycans Modulate Voltage-gated Potassium Channel Gating

“…Whole cell current recordings were performed using pulse protocols, solutions, whole cell patch clamp techniques, and data analyses as described previously (13,14). All experiments were conducted at room temperature, ϳ22°C.…”

“…Several studies detailed the isoform-specific effects of N-glycans on K v (13)(14)(15)(16)(17)20) and voltage-gated Na ϩ (Na v ) channel functions (10 -12). Negatively charged sialic acids (SA) are typically the terminal residues of glycoprotein glycan structures and were shown to impact gating of various voltage-gated ion channels differentially (10 -17, 20, 21).…”

Potassium channels in pulmonary arterial hypertension

“…Protein post-translational modification (PTM) is a highly dynamic and crucial mechanism in which the functional properties of a protein are altered by the covalent addition of a chemical group or protein to its amino-acid residues. Independently of PAH, K V 1.5 has been shown to undergo diverse, reversible or irreversible PTM, including phosphorylation, S-acylation, palmitoylation, sumoylation, ubiquitination and glycosylation, which impact on ion channel expression, channel trafficking to the membrane, stability and activity [35,[70][71][72][73][74]. In response to oxidative stress, a sulfenic acid modification has been reported on a cysteine residue present in the C-terminal domain of K V 1.5 that triggers internalisation of K V 1.5, decreases current density, and diverts the channel from the recycling endosome towards degradation [75].…”

Neuronal glycosylation differentials in normal, injured and chondroitinase-treated environments

“…The altered expression of significant α2,6-and α2,3-STs post-injury did not correlate with staining intensities, and interestingly, expression of the α2,8-ST, relevant to PSA synthesis, was downregulated at all timepoints. Electrical signaling in neurons, skeletal muscle cells and cardiomycetes is modulated by the sialic acid content of particular isoforms of ion channels [35]. Altered or aberrant sialic acid expression could impact neuron polarization [35], which may be consistent with altered excitability of neurons post-injury [33].…”

“…Electrical signaling in neurons, skeletal muscle cells and cardiomycetes is modulated by the sialic acid content of particular isoforms of ion channels [35]. Altered or aberrant sialic acid expression could impact neuron polarization [35], which may be consistent with altered excitability of neurons post-injury [33]. Exposure of α(2,6)-linked sialic acid and binding to SNA-I has been observed on apoptotic and necrotic cells [36], and α(2,6)-sialylation has been identified as blocking binding to galectins, hence functioning as a biological 'off switch' [31].…”