413(Civh) are highly enriched in the postplasm (Fujimura and Takamura 2000 ;
Takamura et al. 2002b). Vasa encodes an ATP-dependent RNA-helicase, and it is
expressed by germ cells and primordial germ cells in most phyla studied to date
(Mochizuki et al. 2001 ; Extavour and Akam 2003 ). Vasa is therefore a reliable
marker for primordial germ cells in all animals.
Vasa-mRNA is distributed broadly in unfertilized eggs (Fig. 8.8), but starts to accu-
mulate at the posterior cortex of the one-cell stage embryo (Fig. 8.8). It is then gradu-
ally segregated into the posterior-most blastomeres at the 4, 8, 16, 32, and 64 cell
stages (Shirae-Kurabayashi et al. 2006 ). The postplasm-containing blastomeres at the
64 cell stage, termed the “B7.6 cells,” undergo an asymmetric cell division during
gastrulation, giving rise to two distinct daughter cells: B8.11 and B8.12. Most of the
postplasmic components are distributed into the “B8.11” cells, which are never incor-
porated into the gonad. The “B8.12” cells accumulate CiVH-protein, which formes
perinuclear granules (Fig. 8.8). This CiVH protein is produced by translation of inher-
ited maternal civh RNA, which is released from the CAB prior to asymmetric division
of the B7.6 cells. Although the postplasm in Ascidians is thought to act as the germ
plasm, it is also highly enriched in several factors essential for somatic cell develop-
ment. Most of the postplasmic RNA’s and the CAB are inherited by the B8.11 cells,
which become somatic cells. An ascidian-specific gene, pem-1 (posterior end-mark-1),
was originally identified in Ciona savignyi as a postplasmic RNA (Satou 1999 ). In
Ciona intestinalis, knockdown of Ci-pem-1 derepressed the transcription of zygotic
genes in the germ line blastomeres, showing that in ascidians, Ci-Pem-1 functions to
prevent somatic gene expression in the germ line. Ci-Pem-1 localizes to the nuclei of
germ line blastomeres and forms a complex with two Ciona intestinalis homologs of
the transcriptional co-repressor Groucho (Shirae-Kurabayashi et al. 2011 ).
The B8.12 cells then separate from the B.11 cells and divide to produce four
CiVH-positive cells at the late tailbud stage (Shirae-Kurabayashi et al. 2006 ). A study
investigating the location CiVH-positive cells from tailbud stage to juveniles in Ciona
showed that the cells expressing Vasa-protein in middle tailbud embryos are located
in the endodermal strand, where PGC will originate (Takamura et al. 2002b). At the
larva stage, these Vasa-positive cells are located in the posterior half of the tail. During
metamorphosis, these cells move into the trunk region and localize to the debris of the
larval tail (Fig. 8.8). Eight CiVH-positive cells then align between the intestine and
stomach (Fig. 8.8), and subsequently increase in number and form a mass next to the
pyloric gland. These cells later get incorporated into the gonad rudiment, which con-
tains numerous Vasa-positive cells (Fig. 8.8). The testis rudiment later separates from
the gonad rudiment, the remainder of which differentiates into the ovary. In an inter-
esting experiment, the authors observed that when the larval tail containing the
CiVH-positive cells was surgically removed, the resulting juveniles did not contain
any cells expressing Vasa-protein after metamorphosis, indicating that germ cells
normally originate from the CiVH-expressing cells in the larval endodermal strand.
However, even in such juveniles, normal germ cells did eventually appear at a later
stage. Since a weak signal for civh-mRNA was detected in the anterior and middle
parts of the endodermal strand at the tailbud stage in cells that do not express CiVH-
protein, and the anterior part of the tail was not removed in the experiment, it is pos-
sible that cells in the endodermal strand have the potential to become germ cells.
8 Mechanisms of Vertebrate Germ Cell Determination