A new dataset of pheromone and pheromone-gene structures from the ciliate, Euplotes crassus

Abstract

Like many other organisms, ciliates communicate and interact socially via diffusible chemical signals, named pheromones, that are functionally associated with a genetic mating-type mechanism of cell self/not-self recognition. In Euplotes species, pheromones form species-specific families of small, globular, and disulfide-rich proteins folding into exclusively helical secondary structures. Each is specified by one of a series of high-multiple alleles that are inherited in Mendelian fashion with relationships of co-dominance at the so-called mat genetic locus of the cell transcriptionally inert micronuclear genome, and expressed in the transcriptionally active macronuclear genome as individual DNA molecules in which the central coding region is flanked by 5’-leader and 3’-trailer noncoding regions ending with C ₄A ₄/T ₄G ₄ telomeric repeats. In E. crassus, a cosmopolitan marine species with a long tradition in the study of ciliate mating systems and breeding patterns, oligonucleotides specific to amino acid sequences of pheromones E c-1 and E c-α were previously used to clone and sequence a first set of four structurally distinct macronuclear ( mac) pheromone coding genes, mac-ec-α, mac-ec-1, mac-ec-2 and mac-ec-3, from two interbreeding strains, L-2D and POR-73. The use of these oligonucleotides in PCR amplifications of macronuclear DNA preparations from three other E. crassus interbreeding strains, ES10, Fava4 and MN4, has now resulted in the characterization of a second set of eight new pheromone coding genes, mac-ec-β, mac-ec-γ, mac-ec-δ, mac-ec-ε, mac-ec- µ, mac-ec-4, mac-ec-5 and mac-ec-6. Multiple alignment between previously and newly determined pheromone-gene sequences reinforces the concept that the E. crassus pheromone-gene family includes two sub-families, which likely reflect a duplication of the micronuclear mat gene locus and represent an apomorphic trait of the E. crassus clade. Members of one sub-family (each identified with a Greek letter) show a 500-bp 5’-leader noncoding region rich in AGGA/AGGGA repetitions, and encode 56-amino acid pheromones with eight conserved Cys residues. Members of the other sub-family (each identified with an Arabic numeral) show an 800-bp 5’-leader noncoding region without AGGA/AGGGA repetitions, and encode 45-amino acid pheromones with ten conserved Cys residues.