Ancestral Ca2+ signaling machinery in early animal and fungal evolution

Abstract

Animals and fungi diverged from a common unicellular ancestor of Opisthokonta, yet they exhibit significant differences in their components of Ca2+ signaling pathways. Many Ca2+ signaling molecules appear to be either animal-specific or fungal-specific, which is generally believed to result from lineage-specific adaptations to distinct physiological requirements. Here, by analyzing the genomic data from several close relatives of animals and fungi, we demonstrate that many components of animal and fungal Ca2+ signaling machineries are present in the apusozoan protist Thecamonas trahens, which belongs to the putative unicellular sister group to Opisthokonta. We also identify the conserved portion of Ca2+ signaling molecules in early evolution of animals and fungi following their divergence. Furthermore, our results reveal the lineage-specific expansion of Ca2+ channels and transporters in the unicellular ancestors of animals and in basal fungi. These findings provide novel insights into the evolution and regulation of Ca2+ signaling critical for animal and fungal biology.

FIG. 1.

Schematic representation of Ca²⁺ signaling machineries in animals and fungi. (A) Animal Ca²⁺ signaling machinery. Ca²⁺ concentration in animal cells is tightly regulated by various Ca²⁺ permeable channels, Ca²⁺ ATPases and exchangers at the plasma membrane and the intracellular compartments. (B) Fungal Ca²⁺ signaling machinery. Ca²⁺ homeostasis is controlled by a relatively simpler system in fungi. (C) The eukaryotic tree of life showing the phylogenetic relationship of species in early animal and fungal evolution in Opisthokonta (Ruiz-Trillo et al. 2007; Sebe-Pedros et al. 2010). The black arrows indicate the directions of Ca²⁺ fluxes across the membranes. Abbreviations used: CAX, Ca²⁺/H⁺ exchanger; CatSper, sperm-associated cation channel; CaV, voltage-gated Ca²⁺ channel; Cch1, Ca²⁺ channel protein 1; CNG, cyclic nucleotide–gated channel; IP₃R, inositol 1,4,5-trisphosphate receptor; LGC, ligand-gated Ca²⁺ channel; Letm1, leucine zipper-EF-hand containing transmembrane protein 1; MiCa, mitochondrial Ca²⁺ channel; Mid1, stretch-activated cation channel; NC(K)X, Na⁺/Ca²⁺ (K⁺-dependent) exchanger; Orai, Orai Ca²⁺ release–activated Ca²⁺ channel; PMC, vacuole Ca²⁺ ATPase; PMCA, plasma membrane Ca²⁺ ATPase; PMR, yeast P-type Ca²⁺ ATPase; RyR, ryanodine receptor; SERCA, sarco/ER Ca²⁺ ATPase; SPCA, secretory pathway Ca²⁺ ATPase; STIM, stromal interaction molecule; TPC, two-pore channel; TRP, transient receptor potential channel; YVC1, yeast vacuolar channel protein.

FIG. 2.

Phylogenetic relationship of CatSper α subunits. The tree constructed with the maximum likelihood approach shows the phylogenetic relationship of CatSper α subunits from the basal fungus Allomyces macrogynus (Ama), the apusozoan protist Thecamonas trahens (Ttr), the sponge Amphimedon queenslandica (Aqu), the amphioxus Branchiostoma floridae (Bfl), Homo sapiens (Hsa), and Mus musculus (Mus). The prokaryotic voltage-gated Na⁺ channel from Bacillus halodurans, NaChBac, was used as an out-group. Bootstrap values greater than 50 are shown at the nodes.

FIG. 3.

Identification of protist ryanodine receptors. (A) A maximum likelihood phylogenetic tree showing the relationships of protist and animal homologs of IP₃Rs and RyRs. (B) Sequence alignments of IP₃ binding regions in IP₃Rs and the corresponding regions in RyRs. (C) Sequence alignments of ryanodine binding sites in RyRs and the corresponding regions in IP₃Rs. The asterisks indicate the location of functionally important residues indentified in animal IP₃Rs and RyRs previously. A black bar separates the sequences of IP₃Rs and RyRs. Bootstrap values greater than 50 are shown at the nodes. Abbreviations used for species: Amphimedon queenslandica (Aqu), Caenorhabditis elegans (Cel), Capsaspora owczarzaki (Cow), Homo sapiens (Hsa), Monosiga brevicollis (Mbr), Paramecium tetraurelia (Pte), Salpingoeca rosetta (Sro), and Thecamonas trahens (Ttr).

FIG. 4.

Early emergence of NCKX exchanger predating NCX exchangers. (A) A maximum likelihood phylogenetic tree depicting the relationships of NCX and NCKX exchangers from representative species. The putative prokaryotic cation/Ca²⁺ exchanger protein from Escherichia coli, YRBG, was used as an out-group. (B) Sequence alignments of α2 repeat regions of NC(K)X exchangers. The signature motif in α2 repeats (Cai and Lytton 2004a) is underlined. The asterisk symbol indicates the location of the key aspartate residue essential for exchanger activity (Cai and Lytton 2004a). The filled circle indicates the location of the conserved aspartate residue in NCKXs, which is critical for K⁺-dependence of NCKX exchangers (Kang et al. 2005). Bootstrap values greater than 50 are shown at the nodes. Abbreviations used for species: Capsaspora owczarzaki (Cow), Escherichia coli (Eco), Monosiga brevicollis (Mbr), Mus musculus (Mus), Salpingoeca rosetta (Sro), and Thecamonas trahens (Ttr).

FIG. 5.

Ca²⁺ signaling machineries in Apusozoan protist, fungi, unicellular relatives of animals, and animals. The phylogenetic relationships of species are inferred from the Tree of Life project (http://www.tolweb.org/) and recent references (James et al. 2006; Ruiz-Trillo et al. 2007; Sebe-Pedros et al. 2010). The number of protein homologs derived from currently available genomic databases is shown, while in few instances, a black dot indicates the presence of protein homolog (s). Absence of a number and a black dot indicates that no homolog was indentified. For protein abbreviations, see figure 1.