A Voltage-Gated H+ Channel Underlying pH Homeostasis in Calcifying Coccolithophores

Abstract

Marine coccolithophorid phytoplankton are major producers of biogenic calcite, playing a significant role in the global carbon cycle. Predicting the impacts of ocean acidification on coccolithophore calcification has received much recent attention and requires improved knowledge of cellular calcification mechanisms. Uniquely amongst calcifying organisms, coccolithophores produce calcified scales (coccoliths) in an intracellular compartment and secrete them to the cell surface, requiring large transcellular ionic fluxes to support calcification. In particular, intracellular calcite precipitation using HCO3 − as the substrate generates equimolar quantities of H+ that must be rapidly removed to prevent cytoplasmic acidification. We have used electrophysiological approaches to identify a plasma membrane voltage-gated H+ conductance in Coccolithus pelagicus ssp braarudii with remarkably similar biophysical and functional properties to those found in metazoans. We show that both C. pelagicus and Emiliania huxleyi possess homologues of metazoan Hv1 H+ channels, which function as voltage-gated H+ channels when expressed in heterologous systems. Homologues of the coccolithophore H+ channels were also identified in a diversity of eukaryotes, suggesting a wide range of cellular roles for the Hv1 class of proteins. Using single cell imaging, we demonstrate that the coccolithophore H+ conductance mediates rapid H+ efflux and plays an important role in pH homeostasis in calcifying cells. The results demonstrate a novel cellular role for voltage gated H+ channels and provide mechanistic insight into biomineralisation by establishing a direct link between pH homeostasis and calcification. As the coccolithophore H+ conductance is dependent on the trans-membrane H+ electrochemical gradient, this mechanism will be directly impacted by, and may underlie adaptation to, ocean acidification. The presence of this H+ efflux pathway suggests that there is no obligate use of H+ derived from calcification for intracellular CO2 generation. Furthermore, the presence of Hv1 class ion channels in a wide range of extant eukaryote groups indicates they evolved in an early common ancestor.

Voltage-gated proton channel in a dinoflagellate

“…His occupies the equivalent position in kH V 1, EhH V 1, and CpH V 1 (45). The work by Sakata et al (43) showed that the murine equivalent N → R mutant retained proton conductance.…”

Emiliania huxleyi increases calcification but not expression of calcification-related genes in long-term exposure to elevated temperature and p CO 2

“…The voltage-gated H þ channel investigated in [53] that was shown to be important in calcification either did not map to any of the sequences in the E. huxleyi 1516 reference transcriptome or had zero expression in our treatments ( The four transcripts examined by qPCR showed no significant differences in expression between treatments or generation points (figure 7). Expression levels of the examined genes showed a high level of variability in the RNA-seq and qPCR assays.…”

Biomineralization in perforate foraminifera

“…Compared to foraminifera, biomineralization in corals (Al-Horani et al, 2003;Sinclair and Risk, 2006;Venn et al, 2013), coccolithophores (Marsh, 2003;Taylor et al, 2011;Ziveri et al, 2012;Bach et al, 2013), gastropods (e.g. Nehrke and Nouet, 2011) and bivalves (Nudelman et al, 2006;Nehrke et al, 2012;Shi et al, 2013) are understood in greater detail.…”