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Germ cells are required for the successful propagation of sexually reproducing species. Understanding the mechanisms by which these cells are specified and how their totipotency is established and maintained has important biomedical and evolutionary implications. Freshwater planarians serve as fascinating models for studying these questions. They can regenerate germ cells from fragments of adult tissues that lack reproductive structures, suggesting that inductive signaling is involved in planarian germ cell specification. To study the development and regeneration of planarian germ cells, we have functionally characterized an ortholog of nanos, a gene required for germ cell development in diverse organisms, from Schmidtea mediterranea. In the hermaphroditic strain of this species, Smed-nanos mRNA is detected in developing, regenerating, and mature ovaries and testes. However, it is not detected in the vast majority of newly hatched planarians or in small tissue fragments that will ultimately regenerate germ cells, consistent with an epigenetic origin of germ cells. We show that Smed-nanos RNA interference (RNAi) results in failure to develop, regenerate, or maintain gonads in sexual planarians. Unexpectedly, Smed-nanos mRNA is also detected in presumptive testes primordia of asexual individuals that reproduce strictly by fission. These presumptive germ cells are lost after Smed-nanos RNAi, suggesting that asexual planarians specify germ cells, but their differentiation is blocked downstream of Smed-nanos function. Our results reveal a conserved function of nanos in germ cell development in planarians and suggest that these animals will serve as useful models for dissecting the molecular basis of epigenetic germ cell specification. epigenesis | germ cell specification | RNA interference | Schmidtea mediterranea | Platyhelminthes

Freshwater planarians have prodigious regenerative abilities that enable them to form complete organisms from tiny body fragments. This plasticity is also exhibited by the planarian germ cell lineage. Unlike many model organisms in which germ cells are specified by localized determinants, planarian germ cells appear to be specified epigenetically, arising postembryonically from stem cells. The planarian Schmidtea mediterranea is well suited for investigating the mechanisms underlying epigenetic germ cell specification. Two strains of S. mediterranea exist: a hermaphroditic strain that reproduces sexually and an asexual strain that reproduces by means of transverse fission. To date, expressed sequence tags (ESTs) have been generated only from the asexual strain. To develop molecular reagents for studying epigenetic germ cell specification, we have sequenced 27,161 ESTs from two developmental stages of the hermaphroditic strain of S. mediterranea; this collection of ESTs represents [approximately equal to]10,000 unique transcripts. BLAST analysis of the assembled ESTs showed that 66% share similarity to sequences in public databases. We annotated the assembled ESTs using Gene Ontology terms as well as conserved protein domains and organized them in a relational database. To validate experimentally the Gene Ontology annotations, we used whole-mount in situ hybridization to examine the expression patterns of transcripts assigned to the biological process 'reproduction.' Of the 53 genes in this category, 87% were expressed in the reproductive organs. In addition to its utility for studying germ cell development, this EST collection will be an important resource for annotating the planarian genome and studying this animal's amazing regenerative abilities. Gene Ontology | germ cells | planarians | Platyhelminthes

Germ cells are required for the successful propagation of sexually reproducing species. Understanding the mechanisms by which these cells are specified and how their totipotency is established and maintained has important biomedical and evolutionary implications. Freshwater planarians serve as fascinating models for studying these questions. They can regenerate germ cells from fragments of adult tissues that lack reproductive structures, suggesting that inductive signaling is involved in planarian germ cell specification. To study the development and regeneration of planarian germ cells, we have functionally characterized an ortholog of nanos, a gene required for germ cell development in diverse organisms, from Schmidtea mediterranea. In the hermaphroditic strain of this species, Smed-nanos mRNA is detected in developing, regenerating, and mature ovaries and testes. However, it is not detected in the vast majority of newly hatched planarians or in small tissue fragments that will ultimately regenerate germ cells, consistent with an epigenetic origin of germ cells. We show that Smed-nanos RNA interference (RNAi) results in failure to develop, regenerate, or maintain gonads in sexual planarians. Unexpectedly, Smed-nanos mRNA is also detected in presumptive testes primordia of asexual individuals that reproduce strictly by fission. These presumptive germ cells are lost after Smed-nanos RNAi, suggesting that asexual planarians specify germ cells, but their differentiation is blocked downstream of Smed-nanos function. Our results reveal a conserved function of nanos in germ cell development in planarians and suggest that these animals will serve as useful models for dissecting the molecular basis of epigenetic germ cell specification. [ABSTRACT FROM AUTHOR]