Expression of biomineralization‐related ion transport genes in Emiliania huxleyi

Abstract

Biomineralization in the marine phytoplankton Emiliania huxleyi is a stringently controlled intracellular process. The molecular basis of coccolith production is still relatively unknown although its importance in global biogeochemical cycles and varying sensitivity to increased pCO₂ levels has been well documented. This study looks into the role of several candidate Ca²⁺, H⁺ and inorganic carbon transport genes in E. huxleyi, using quantitative reverse transcriptase PCR. Differential gene expression analysis was investigated in two isogenic pairs of calcifying and non-calcifying strains of E. huxleyi and cultures grown at various Ca²⁺ concentrations to alter calcite production. We show that calcification correlated to the consistent upregulation of a putative HCO₃⁻ transporter belonging to the solute carrier 4 (SLC4) family, a Ca²⁺/H⁺ exchanger belonging to the CAX family of exchangers and a vacuolar H⁺-ATPase. We also show that the coccolith-associated protein, GPA is downregulated in calcifying cells. The data provide strong evidence that these genes play key roles in E. huxleyi biomineralization. Based on the gene expression data and the current literature a working model for biomineralization-related ion transport in coccolithophores is presented.

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

“…Quantitative real time PCR (qPCR) verification of expression in all chemostats across all three generation points was conducted for calcification-related transcripts, a gamma carbonic anhydrase, a cation/H þ exchanger, a putative bicarbonate transporter and proton channel, and normalized to the housekeeping gene actin using published primer sequences (table 2) [26]. RNA was extracted from two samples per chemostat per generation point (see electronic supplementary material, table S1) using Tri Reagent (Molecular Research Center, Inc.) according to the manufacturer's protocol and reverse transcribed into cDNA using Improm-II reverse transcriptase (Promega).…”

“…Samples for physiological parameters were taken after 215, 414 and 703 generations to examine responses to future ocean conditions. Genome-wide transcriptomic changes between present and future ocean conditions are discussed at the 215 generation point, and qPCR for some genes that have been previously identified as involved with calcification [26] was measured across all generation points.…”

“…Genes putatively related to calcification (e.g. calcium and inorganic carbon transport, H þ transport and carbonic anhydrases) have been identified via gene expression studies comparing calcifying and non-calcifying E. huxleyi cells [25][26][27][28][29], or in short-term experiments where calcification was regulated by limitation of ions needed for calcification (i.e. Ca 2þ , HCO 3 2 /CO 3 22 [26,30,31]).…”

Reconsidering the role of carbonate ion concentration in calcification by marine organisms

“…As this ion is also the form used in the final precipitation reaction (Eq. 1), calcium does not need to be chemically transformed while being transported from seawater to the site of calcification (Allemand et al, 2004;Bentov et al, 2009;Mackinder et al, 2011). This is in clear contrast to CO 2− 3 , where the relation between ion source from seawater and ion sink during crystallization is considerably more complex since CO 2− 3 is in constant exchange with HCO − 3 and CO 2 .…”

Mineralization and non-ideality: on nature’s foundry

“…For example, in simple planktons, biochemical pathways regulate seawater endocytosis and transportation to mineralization sites (de Nooijer et al 2009). Important members include ion transporters such as Ca 2+ ATPase pumps and anion exchangers (Fan et al 2007;Mackinder et al 2011) as well as enzymes such as carbonic anhydrase. The activities of these biomolecules condition the pre-nucleation and early mineralization stages, for instance by regulating supersaturation and pH.…”