Figure 4.

Fast voltage-activated inward currents in the coccolithophore S. apsteinii. As with C. pelagicus (Taylor and Brownlee, 2003), the coccolithophore S. apsteinii exhibits fast-activating and inactivating voltage-activated currents that likely underpin membrane excitability. The native currents observed in S. apsteinii cells have remarkably similar characteristics to the SaEUKCATA1 channel expressed in HEK cells. A, Image on the left shows a micrograph of patch-clamp pipette forming a seal on a decalcified S. apsteinii cell in normal seawater. Scale bar = 10 μm. The right shows fast-activating and inactivating inward current evoked by a voltage command from a holding potential of –115 mV to –55 mV (left). A second voltage stimulus to –45 mV evokes a short train of fast inward currents showing the recovery from inactivation is very fast (left). The cell was 19 pF and the seal was 5 GΩ. B, Family of depolarizing pulses from a holding potential of –115 mV in steps of + 10 mV (left) and corresponding current–voltage curve from the same cell recording (right) showing peak inward current at ∼–55 mV. The cell was 13 pF and the seal was 2 GΩ. C, Characteristic steady-state inactivation of native S. apsteinii voltage-activated currents. A prepulse protocol was used to depolarize the cell from a holding potential of −115 mV in + 5 mV increments for 100 ms before the test depolarization to –55 mV (left). Normalized peak inward current in response to the test pulse was plotted against prepulse voltage and fitted using a Boltzmann curve yielding a half-inactivation voltage of –53.5 mV (right). Cell was the same as in A. D, Multiple sequence alignment of the four pore domains of coccolithophore 4D-Ca_v/Na_vs. Sequences from coccolithophore genomes (E. huxleyi CCMP 1516) or transcriptomes (E. huxleyi PLY379 and S. apsteinii) are shown aligned to Na⁺-selective Na_v1.1, and Ca²⁺-permeable Ca_v2.1 from humans. The coccolithophore sequences most closely resemble the EEEE motif found in metazoan Ca²⁺ channels and lack the conserved lysines found in Na⁺ channels. The 4D-Ca_v/Na_v sequence found within the S. apsteinii transcriptome represents a partial sequence, as it lacks domain I.