Evidence for the chimeric origin of a pheromone-coding gene in Euplotes raikovi

Ciliates, unique among eukaryotes, evolved two types of nucleus which are distinct in both structure and function. A diploid transcriptionally silent germline micronucleus (MIC) with an orthodox chromosomicstructure coexists in the same cytoplasm with a polyploid transcriptionally active somatic macronucleus (MAC) showing a unique sub-chromosomic structure. This structure is acquired in coincidence with every sexual event, when the ex-conjugant (or ex-autogamic) cell initiates a new life cycle developing a new MAC from a mitotic product of the synkaryon. In spirotrichous ciliates such as Euplotes, the chromosomes of this product undergo impressive phenomena of polytenization, fragmentation in thousands of DNA fragments known as ‘Macronuclear Destined Sequences’ (MDSs), and DNA elimination. Under the guide of noncoding RNA templates synthesized by the old MAC before being destroyed, theseMDSs are assembled into sub-chromosomic (‘gene-size’)DNA molecules which,amplified to thousands of copies, compose the new MAC.The way to a correct MDSassembly may be crossed by errors, with the consequent generation of functional chimeric genes which can stably be integrated and expressed in the MAC genome.One of these chimeric genes came to light by studying the genetic basis of the pheromone-mediated self/not-self recognition mechanism in E. raikovi. The genome of type I cells secreting pheromone Er-1 was found to contain two structurally distinct MAC Er-1 coding genes,both expressed via a mechanism of intron splicing responsible for the synthesis of the Er-1 soluble form and a membrane-bound Er-1 isoform functioning as autocrine pheromone receptor. The sequence of one gene resulted unmistakably homologous throughout its length with the sequences of other members of the E. raikovipheromone gene family. In contrast, the sequence of the second gene resulted unique at level of a 359-bp segment of the 5’ region destined to specify the cytoplasmic domain of the Er-1 membrane-bound isoform. By showing that this segment arises from a wrong assembly of a MDS destined to a 2417-bp gene with no relationship with the signaling pheromone system, we provide additional evidence that the generation of functionally active chimeric genesfrom not-programmed phenomena of somatic MDS recombination is an effective and MIC-independent source of gene variants in the ciliate MAC genome.