Int. J. Dev. Biol. 56: 93 - 102 (2012)
doi: 10.1387/ijdb.113434ns
© UBC Press

Comprehensive gene expression analyses in pluripotent stem cells of a planarian, Dugesia japonica

Norito Shibata*,1, Tetsutaro Hayashi2, Ryutaro Fukumura3, Junsuke Fujii4, Tomomi Kudome-Takamatsu2, Osamu Nishimura1, Syozo Sano4, Fuyan Son3, Nobuko Suzuki3, Ryoko Araki3, Masumi Abe3 and Kiyokazu Agata*,4

1Global COE Program: Evolution and Biodiversity, Grad. School of Science, Kyoto Univ. 2Center for Dev. Biol., RIKEN, 3Transcriptome Research Center, NIRS and 4Dept. Biophysics, Grad. School of Science, Kyoto Univ., Japan

ABSTRACT The neoblasts are the only somatic stem cells in planarians possessing pluripotency, and can give rise to all types of cells, including germline cells. Recently, accumulated knowledge about the transcriptome and expression dynamics of various pluripotent somatic stem cells has provided important opportunities to understand not only fundamental mechanisms of pluripotency, but also stemness across species at the molecular level. The neoblasts can easily be eliminated by radiation. Also, by using fluorescence activated cell sorting (FACS), we can purify and collect many neoblasts, enabling identification of neoblast-related genes by comparison of the gene expression level among intact and X-ray-irradiated animals, and purified neoblasts. In order to find such genes, here we employed the high coverage expression profiling (HiCEP) method, which enables us to observe and compare genome-wide gene expression le-vels between different samples without advance sequence information, in the planarian D. japonica as a model organism of pluripotent stem cell research. We compared expression levels of ~17,000 peaks corresponding to independent genes among different samples, and ob-tained 102 peaks as candidates. Expression analysis of genes identified from those peaks by in situ hybridization revealed that at least 42 genes were expressed in the neoblasts and in neoblast-related cells that had a different distribution pattern in the body than neoblasts. Also, single-cell PCR analysis of those genes revealed heterogeneous expression of some genes in the neoblast population. Thus, using multidimensional gene expression analyses, we were able to obtain a valuable data set of neoblast-related genes and their expression patterns.

Keywords:

planarian, pluripotent stem cell, HiCEP, comprehensive gene expression analysis

*Corresponding author e-mail: agata@mdb.biophys.kyoto-u.ac.jp