Article

Early lineage specification of long-lived germline precursors in the colonial ascidian Botryllus schlosseri

Biology Department, Center for Developmental Biology, and Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA.
Development (Impact Factor: 6.27). 10/2009; 136(20):3485-94. DOI: 10.1242/dev.037754
Source: PubMed

ABSTRACT In many taxa, germline precursors segregate from somatic lineages during embryonic development and are irreversibly committed to gametogenesis. However, in animals that can propagate asexually, germline precursors can originate in adults. Botryllus schlosseri is a colonial ascidian that grows by asexual reproduction, and on a weekly basis regenerates all somatic and germline tissues. Embryonic development in solitary ascidians is the classic example of determinative specification, and we are interested in both the origins and the persistence of stem cells responsible for asexual development in colonial ascidians. In this study, we characterized vasa as a putative marker of germline precursors. We found that maternally deposited vasa mRNA segregates early in development to a posterior lineage of cells, suggesting that germline formation is determinative in colonial ascidians. In adults, vasa expression was observed in the gonads, as well as in a population of mobile cells scattered throughout the open circulatory system, consistent with previous transplantation/reconstitution results. vasa expression was dynamic during asexual development in both fertile and infertile adults, and was also enriched in a population of stem cells. Germline precursors in juveniles could contribute to gamete formation immediately upon transplantation into fertile adults, thus vasa expression is correlated with the potential for gamete formation, which suggests that it is a marker for embryonically specified, long-lived germline progenitors. Transient vasa knockdown did not have obvious effects on germline or somatic development in adult colonies, although it did result in a profound heterochrony, suggesting that vasa might play a homeostatic role in asexual development.

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    • "Arrows point to testes and arrowheads point to eggs. Panels A and B are modified from Brown et al. 2009, Laird et al. 2005 and Laird and De Tomaso 2005. secondary bud(s). "
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    ABSTRACT: Gonad differentiation is an essential function for all sexually reproducing species, and many aspects of these developmental processes are highly conserved among the metazoa. The colonial ascidian, Botryllus schlosseri is a chordate model organism which offers two unique traits that can be utilized to characterize the genes underlying germline development: a colonial life history and variable fertility. These properties allow individual genotypes to be isolated at different stages of fertility and gene expression can be characterized comprehensively. Here we characterized the transcriptome of both fertile and infertile colonies throughout blastogenesis (asexual development) using differential expression analysis. We identified genes (as few as 7 and as many as 647) regulating fertility in Botryllus at each stage of blastogenesis. Several of these genes appear to drive gonad maturation, as they are expressed by follicle cells surrounding both testis and oocyte precursors. Spatial and temporal expression of differentially expressed genes was analyzed by in situ hybridization, confirming expression in developing gonads. We have identified several genes expressed in developing and mature gonads in B. schlosseri. Analysis of genes upregulated in fertile animals suggests a high level of conservation of the mechanisms regulating fertility between basal chordates and vertebrates.
    BMC Genomics 12/2014; 15(1):1183. DOI:10.1186/1471-2164-15-1183 · 4.04 Impact Factor
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    • "Our FISH protocol also reliably detected gene expression in germline and germline support cells. The B. schlosseri vasa gene has been previously described (Brown et al., 2009; Rosner et al., 2009), and we identified a homolog of B. primigenus Piwi (piwi; E-value 5 8.0 3 10 2151 ). We found expression of these germline markers in clusters of primordial germ cells associated FIG. 3. Expression patterns of markers for developing structures in B. schlosseri. "
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    ABSTRACT: Botryllus schlosseri is a colonial ascidian with characteristics that make it an attractive model for studying immunology, stem cell biology, evolutionary biology, and regeneration. Transcriptome sequencing and the recent completion of a draft genome sequence for B. schlosseri have revealed a large number of genes, both with and without vertebrate homologs, but analyzing the spatial and temporal expression of these genes in situ has remained a challenge. Here we report a robust protocol for in situ hybridization that enables the simultaneous detection of multiple transcripts in whole adult B. schlosseri using Tyramide Signal Amplification in conjunction with digoxigenin- and dinitrophenol-labeled RNA probes. Using this protocol we have identified a number of genes that can serve as markers for developing and mature structures in B. schlosseri, permitting analysis of phenotypes induced in loss-of-function experiments. © 2014 Wiley Periodicals, Inc.
    genesis 09/2014; 53(1). DOI:10.1002/dvg.22820 · 2.04 Impact Factor
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    • "Studies on self-and cross-fertilization declined in the 1970s but still offered cues regarding the successive investigations on the reproduction of B. schlosseri, primarily addressing the study of the morphology of the gonads and gametes (Burighel and Martinucci, 2000; Burighel et al., 1982; Manni et al., 1993, 1994; Sabbadin and Zaniolo, 1979), the success of fertilization in relation to population density and sperm release (Johnson and Yund, 2004; Phillippi et al., 2004; Stewart-Savage et al., 2001; Stewart-Savage and Yund, 1997), placentation (Zaniolo et al., 1987), germ cell recognition and migration (Ballarin et al., 2011; Brown et al., 2009; Rinkevich et al., 2013; Rosner et al., 2013; Sabbadin and Zaniolo, 1979). "
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    ABSTRACT: The colonial tunicate Botryllus schlosseri is a widespread filter-feeding ascidian that lives in shallow waters and is easily reared in aquaria. Its peculiar blastogenetic cycle, characterized by the presence of three blastogenetic generations (filtering adults, buds and budlets) and by recurrent generation changes, has resulted in over 60 years of studies aimed at understanding how sexual and asexual reproduction are coordinated and regulated in the colony. The possibility of using different methodological approaches, from classical genetics to cell transplantation, contributed to the development of this species as a valuable model organism for the study of a variety of biological processes. Here, we review the main studies detailing rearing, staging methods, reproduction and colony growth of this species, emphasizing the asymmetry in sexual and asexual reproduction potential, sexual reproduction in the field and the laboratory, and self- and cross-fertilization. These data, opportunely matched with recent tanscriptomic and genomic outcomes, can give a valuable help to the elucidation of some important steps in chordate evolution. © 2014 Wiley Periodicals, Inc.
    genesis 07/2014; 53(1). DOI:10.1002/dvg.22802 · 2.04 Impact Factor
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