Localization and expression pattern of type I postplasmic mRNAs in embryos of the ascidian Halocynthia roretzi

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Abstract

The posterior-vegetal cytoplasm (PVC) of fertilized ascidian eggs plays important roles in embryo development. It has been reported that some maternal RNAs are localized to the PVC. We identified four novel type I postplasmic mRNAs that are localized to the PVC through the use of data from a cDNA project of maternal mRNAs in the eggs of Halocynthia roretzi (MAGEST database). The mRNAs are HrGLUT, HrPEN-1, and HrPEM-3, which show similarity to a glucose transporter, a g1-related protein, and Ciona pem-3, respectively; and HrPEN-2, with no similarity. Maternal mRNAs of all four genes were identically localized to the PVC after ooplasmic segregation. During cleavage, they were concentrated in the centrosome-attracting body (CAB) and were then segregated into the small blastomeres located at the posterior pole. This localization pattern is common to all known type I postplasmic mRNAs found so far. HrGLUT, HrPEN-1, and HrPEM-3 were expressed zygotically in various tissues later in embryogenesis: HrGLUT and HrPEM-3 in the mesenchyme and nervous system, and HrPEN-1 in the ectodermal cells.

Section snippets

Results and discussion

It has been suggested that maternal factors localized to the posterior-vegetal cytoplasm (PVC) in Halocynthia roretzi eggs play important roles in control of cleavage pattern, determination of the anterior–posterior axis, autonomous specification of muscle fate, and generation of differences in responsiveness to inductive signals in mesenchyme and notochord precursor blastomeres. These functions were revealed in micromanipulative experiments in which PVC was removed and transplanted (Nishida,

In situ hybridization

Whole-mount in situ hybridization was performed as described Miya et al., 1994, Miya et al., 1997. The HrGLUT and HrPEN-1 antisense RNA probes were synthesized using a full-length cDNA as template. The HrPEM-3 probe was transcribed from 1083–3690 bp fragment of HrPEM-3 cDNA. The HrPEN-2 probe was generated from 1522–4015 bp fragment of HrPEN-2 cDNA. Most embryos were then dehydrated in a graded series of ethanol and rendered transparent with a 1:2 mixture (v/v) of benzyl alcohol and benzyl

Acknowledgements

We thank T. Kawashima (Kyoto University) and Y. Kohara (National Institute of Genetics) for providing MAGEST plasmids. We also thank members of the Asamushi Marine Biological Station and the Otsuchi Marine Research Center for help in collecting live ascidian adults, and members of the Misaki Marine Biological Laboratory for help in maintaining them. This work was supported by the Research for the Future Program of the Japanese Society for the Promotion of Science, and by Grants-in-Aid from the

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