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Germline cell fate decisions are primarily controlled at the post-transcriptional level with DEAD-box RNA helicases playing a crucial role in germline development. In this study, we report the identification of two DEAD-box vasa/PL10 orthologues (RlVlg and RlPL10) in a species complex of the genus Rana, characterized by hybridogenetic reproduction, an enigmatic process that involves the exclusion of an individual genome, and endoreduplication events. Both genes were expressed during the early stages of gametogenesis of R. ridibunda, R. lessonae, and their natural hybrid R. esculenta. RlVlg expression was germline specific. On the other hand, RlPL10 was also expressed in somatic tissues, although only at low levels. The two genes were expressed in different phases of mitotic and meiotic spermatogenetic divisions as demonstrated by immunostaining with an anti-H3 phosphohistone antibody. The data indicate that RlVlg and RlPL10 may represent useful markers for dissecting the molecular aspects of genome exclusion and endoreduplication of the hybridogenetic gametogenesis.

European water frog hybrids Rana esculenta (R. ridibundaxR. lessonae) reproduce hemiclonally, by hybridogenesis: in the germ line they exclude the genome of one parental species and produce haploid gametes with an unrecombined genome of the other parental species. In the widespread L-E population system, both sexes of hybrids (E) coexist with R. lessonae (L). They exclude the lessonae genome and produce ridibunda gametes. In the R-E system, hybrid males coexist with R. ridibunda (R); they exclude either their ridibunda or their lessonae genome and produce sperm with a lessonae or with a ridibunda genome or a mixture of both kinds of sperm. We examined 13 male offspring, 12 of which were from crosses between L-E system and R-E system frogs. All were somatically hybrid. With one exception, they excluded the lessonae genome in the germ line and subsequently endoreduplicated the ridibunda genome. Spermatogonial metaphases contained a haploid or a diploid number of ridibunda chromosomes, identified through in situ hybridization to a satellite DNA marker, and by spermatocyte I metaphases containing a haploid number of ridibunda bivalents. The exception, an F1 hybrid between L-E system R. lessonae and R-E system R. ridibunda, was not hybridogenetic, showed no genome exclusion, and evidenced a disturbed gametogenesis resulting from the combination of two heterospecific genomes. None of the hybridogenetic hybrids showed any cell lines excluding the ridibunda genome, the pattern most frequent in hybrids of the R-E system, unique to that system, and essential for its persistence. A particular combination of R-E system lessonae and R-E system ridibunda genomes seems necessary to induce the R-E system type of hemiclonal gametogenesis.

Y-box proteins are a highly conserved family of gene expression regulatory factors. During gametogenesis they may play a dual role, as both transcriptional activators of germ cell-specific genes and as translational repressors of stored maternal transcripts. We report the identification of RlYB2, a Y-box homolog gene specifically expressed in the germ cells of green frogs belonging to Rana esculenta complex, a model system characterized by a hybridogenetic gametogenesis. In developing germ cells of the hybrid R. esculenta, arisen by natural cross of the parental species R. ridibunda and R. lessonae, one set of the parental genomes is excluded and the remaining one, first endoreduplicates and then is transmitted to gametes. In situ hybridizations performed on gonadal tissues showed that the RlYB2 transcript was widely expressed in the ooplasm at early stages of oogenesis in both the parental species and hybrids. Interestingly, a hybridization signal, presumably related to RlYB2-like nascent transcripts, was observed in nuclei of stage II oocytes. The presence of RlYB2 mRNA during early oogenesis suggests that this gene may be involved in regulating the transcription and/or translation of maternal mRNAs in this special vertebrate model system. [ABSTRACT FROM AUTHOR]