Bioinformatic analysis of CaBP/calneuron proteins reveals a family of highly conserved vertebrate Ca2+-binding proteins

Abstract

Background: Ca2+-binding proteins are important for the transduction of Ca2+ signals into physiological outcomes. As in calmodulin many of the Ca2+-binding proteins bind Ca2+ through EF-hand motifs. Amongst the large number of EF-hand containing Ca2+-binding proteins are a subfamily expressed in neurons and retinal photoreceptors known as the CaBPs and the related calneuron proteins. These were suggested to be vertebrate specific but exactly which family members are expressed outside of mammalian species had not been examined.

Findings: We have carried out a bioinformatic analysis to determine when members of this family arose and the conserved aspects of the protein family. Sequences of human members of the family obtained from GenBank were used in Blast searches to identify corresponding proteins encoded in other species using searches of non-redundant proteins, genome sequences and mRNA sequences. Sequences were aligned and compared using ClustalW. Some families of Ca2+-binding proteins are known to show a progressive expansion in gene number as organisms increase in complexity. In contrast, the results for CaBPs and calneurons showed that a full complement of CaBPs and calneurons are present in the teleost fish Danio rerio and possibly in cartilaginous fish. These findings suggest that the entire family of genes may have arisen at the same time during vertebrate evolution. Certain members of the family (for example the short form of CaBP1 and calneuron 1) are highly conserved suggesting essential functional roles.

Conclusions: The findings support the designation of the calneurons as a distinct sub-family. While the gene number for CaBPs/calneurons does not increase, a distinctive evolutionary change in these proteins in vertebrates has been an increase in the number of splice variants present in mammals.

Figure 1

Alignment of the sequences of human calmodulin, CaBPs and calneuron proteins. Protein sequences were compiled and aligned using ClustalW. Residues that are identical in more than one protein are highlighted in red and residues with similar properties are highlighted in blue. The lines above the sequences indicate the position of the predicted 12 amino acid coordinating loop within the EF-hands The green bar beneath the sequence indicates the position of the linker region in the CaBPs that is extended in comparison to calmodulin.

Figure 2

Alignment of the sequences of zebrafish (Danio rerio) calmodulin, CaBPs and calneuron proteins. Protein sequences were compiled and aligned using ClustalW. Residues that are identical in more than one protein are highlighted in red and residues with similar properties are highlighted in blue.

Figure 3

Alignment of the sequences of the individual zebrafish and the corresponding human CaBPs and calneuron proteins. Protein sequences were compiled and aligned using ClustalW. Residues that are identical in more than one protein are highlighted in red and residues with similar properties are highlighted in blue. The position of the predicted EF hands in each protein is indicated.

Figure 4

Phylogenetic tree showing the relatedness of the human and zebrafish calmodulin, CaBPs and calneuron proteins. A consensus maximum likelihood tree based on the alignment of the CaBP and calneuron proteins was generated and rooted on human calmodulin. The percentage of 2000 bootstrap replicates supporting the topology is given at each node.

Figure 5

Relative sequence identity of individual domains with the human sequence. The percentage identity compared to human is shown for three indicated domains for a range of species. Where no symbol is present no orthologous protein or domain could be found from Blast searches. The following species are shown Danio rerio (Dan), Xenopus laevis (Xen), Gallus gallus (Gal), Taeniopygia guttata (Tae), Monodelphis domestica (Mon), Canis lupus familiaris (Can), Mus musculus (Mus), Rattus norvegicus (Rat), Bos taurus (Bos), Macaca mulatta (Mac) Pan troglodytes (Pan) and Homo sapiens (Hom).