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Development
2007 Oct 01;13419:3549-63. doi: 10.1242/dev.006569.
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The Oct4 homologue PouV and Nanog regulate pluripotency in chicken embryonic stem cells.
Lavial F
,
Acloque H
,
Bertocchini F
,
Macleod DJ
,
Boast S
,
Bachelard E
,
Montillet G
,
Thenot S
,
Sang HM
,
Stern CD
,
Samarut J
,
Pain B
.
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Embryonic stem cells (ESC) have been isolated from pregastrulation mammalian embryos. The maintenance of their pluripotency and ability to self-renew has been shown to be governed by the transcription factors Oct4 (Pou5f1) and Nanog. Oct4 appears to control cell-fate decisions of ESC in vitro and the choice between embryonic and trophectoderm cell fates in vivo. In non-mammalian vertebrates, the existence and functions of these factors are still under debate, although the identification of the zebrafish pou2 (spg; pou5f1) and Xenopus Pou91 (XlPou91) genes, which have important roles in maintaining uncommitted putative stem cell populations during early development, has suggested that these factors have common functions in all vertebrates. Using chicken ESC (cESC), which display similar properties of pluripotency and long-term self-renewal to mammalian ESC, we demonstrated the existence of an avian homologue of Oct4 that we call chicken PouV (cPouV). We established that cPouV and the chicken Nanog gene are required for the maintenance of pluripotency and self-renewal of cESC. These findings show that the mechanisms by which Oct4 and Nanog regulate pluripotency and self-renewal are not exclusive to mammals.
Fig. 3. cPouV expression during chick embryo development. (A-J) Whole-mount in situ hybridisation to cPouV transcripts. Transcripts are detected in the area pellucida and area opaca of the epiblast in pre-streak embryos (A, stage XI; B, stage XIII), and in the hypoblast in a salt-and-pepper manner (B′). At stage XIV (C), the expression is very strong in the area pellucida of the epiblast, especially where the streak is forming (C′). Transcripts are expressed in the ingressing mesoderm at stages 2-3 (D,D′). As the primitive streak elongates and the embryo grows, expression is still detected in the ectoderm and mesoderm (E, stage 3+; F,F′, stage 4+; G, stage 5). At stage 7 (H) and 8 (I), cPouV mRNA is detected in the forming neural tube and in the underlying mesoderm, but is absent from the endoderm (data not shown). At stage 9 (J), cPouV mRNA is expressed in neural tissue (nt) and presumptive migrating germ cells (gc). B′,C′,D′,F′ are transverse sections of the embryos in B,C,D,F, respectively. ao, area opaca; ep, epiblast; gc, germ cells; hy, hypoblast; m, mesoderm; np, neural plate; nt, neural tube; ps, primitive streak.
Fig. 4. cNanog expression during chick embryo development. (A-J) Whole-mount in situ hybridisation to cNanog transcripts. Nanog transcripts are localised in the epiblast of the area pellucida and area opaca of the pre-streak embryo (A, stage XI; B, stage XII), but not in the hypoblast (B′). From stage XIV, cNanog mRNA disappears from the posterior area pellucida (C,C′) and from the growing primitive streak (D, stage 3; E,E′, stage 3+; F, stage 4+). cNanog transcripts are downregulated in the epiblast from stage 4+ (F), and are confined anteriorly in a crescent region in the epiblast (G, stage 5+). At stage 6 (H), expression is restricted to the neural plate and the neural tube (I, stage 7; J, stage 8). cNanog is also expressed in scattered cells in the germinal crescent from stage 4 (arrowhead in I and H′). B′ is a longitudinal section of the embryo in B, anterior at the right; C′,E′,H′ are transverse sections of embryos in C,E,H, respectively. ep, epiblast; gd, gonad; ms, mesonephros; np, neural plate; nt, neural tube; ps, primitive streak.
Fig. 5. cPouV and cNanog are expressed in germ cells during later embryonic development. (A-H) At stage 33, cPouV mRNA is detected in the developing gonad, which is attached to the mesonephros (A,B), in the germ cells (D), as detected by co-localisation (F) of cPouV (D) both with Cvh (chicken Vasa, E) expression and with SSEA-1-positive cells (G) revealed on adjacent sections counterstained by Hoechst and cPouV probe (H, arrows). (B) Section of gonad shown in A. (C) Bright field of the stage 33 gonad used for in situ hybridisation (D,E,F). (I-M) At stage 33, cNanog is highly expressed in gonads and in mesonophros tubules (I) and gonad (I,J), especially in germ cells (K), as revealed by SSEA-1 staining (L) on the same cells that express cNanog in adjacent sections (M, arrow) counterstained by Hoechst (L). (K) Section of dissected gonad from the urogenital tract (J). ms, mesonephros; gd, gonads. Scale bars: 15 μm.
