Tcfap2c regulation of primordial germ cell development
Date
2011
Authors
Guttormsen, Jillian Bosick, author
Winger, Quinton A., advisor
Bouma, Gerrit J., committee member
Anthony, Russell V., committee member
Garrity, Deborah, committee member
Journal Title
Journal ISSN
Volume Title
Abstract
The development of germ cells during embryonic development is driven by a complex expression pattern of genes. The transcription factor Tcfap2c is expressed in germ cells throughout development from specification to adult sperm and oocytes. Tcfap2c expression is first seen in primordial germ cells around embryonic day (E)6.75 and has been classified as a germ cell specification gene. This study implicates Tcfap2c as a potential key factor in germ cells during specification, proliferation, migration and differentiation. In order to investigate the role of Tcfap2c in germ cells, we utilized the Cre/loxP conditional gene mutation strategy. Cre/loxP allows us to overcome the early embryonic lethality that arises from loss of Tcfap2c in traditional knock-out mice by creating Tcfap2c null mutation in specifically-targeted tissues. We created Tcfap2c mutant mice using the epiblast-specific Sox2-Cre model. Mutant ovaries from this model failed to express both germ cell specific markers and meiotic markers at E12.5. Immunohistochemistry at E18.5 failed to detect the germ cell specific marker NOBOX or the meiotic protein SYCP3, which confirmed that Sox2-Cre, Tcfap2c mutant mice lacked germ cells at late embryonic stages. However, Sox2-Cre, Tcfap2c mutant mice die prior to or at birth preventing us from studying adult gonads from these mice. To this end we used tamoxifen inducible ERTM-Cre mice to create Tcfap2c mutation in adult animals. We assessed ERTM-Cre, Tcfap2c mutant animals for fertility and gametogenesis; surprisingly, fertility, spermatogenesis and oogenesis were not affected in Tcfap2c mutant gonads. These results show that Tcfap2c is not necessary for adult maturation of gonocytes to produce mature sperm and oocytes. However, Sox2-Cre, Tcfap2c mutants lack germ cells indicating that Tcfap2c is necessary during fetal germ cell differentiation. The Sox2-Cre model was limited because Tcfap2c was deleted in the entire embryo and the mutants died at birth. Prdm1-Cre was used to produce a mouse where Tcfap2c is only deleted in germ cells beginning around specification. Prdm1-Cre, Tcfap2c mutants initially specified germ cell-like cells at E7.5; however, by E8.5 the germ cell numbers were decreased and they had not initiated migration towards the genital ridges. By E9.5 few if any germ cells were observed in Prdm1-Cre, Tcfap2c mutants. At E12.5 no germ cells were seen in Prdm1-Cre, Tcfap2c mutant XX or XY gonads. Adult ovaries and testes from Prdm1-Cre, Tcfap2c mutant mice were noticeably smaller than littermate controls and showed no oogenesis or spermatogenesis. The Prdm1-Cre model showed that mutation of Tcfap2c results in loss of germ cells in embryos by E9.5 suggesting that Tcfap2c plays a role during germ cell specification, proliferation and migration. We identified Tcfap2c as an important factor during early germ cell development; however, Tcfap2c expression is observed in germ cells well past specification. We believe that Tcfap2c is present in germ cells during during fetal gonad differentiation because it plays a role in regulating the gene expression pathways necessary for this event. We show that Tcfap2c is expressed in germ cells during the period of fetal gonad differentiation. Gene expression analysis of gonads from E11.5-13.5 reveals Tcfap2c as the most highly expressed member of the Tcfap2 family member. Tcfap2c is a member of a transcription factor family that regulates gene expression by binding consensus sequences within target gene promoters. TCFAP2 binding sites are present in promoter regions of germ cell specific genes Cadherin1 (Cdh1) and Kit oncogene (Kit), as well as in the promoter regions of genes involved in regulating pluripotency High mobility group AT-hook 2 (Hmga2), Nanog homeobox (Nanog) and Lin28. Using chromatin immunoprecipitation we demonstrate that TCFAP2C binds the promoter regions of Cdh1, Kit, Hmga2, Nanog and Lin28. The interaction between TCFAP2C and the promoter regions of Cdh1, Kit, Hmga2, Nanog and Lin28 indicates that Tcfap2c likely plays a functional role in the regulation of these genes. These genes are necessary for germ cell survival, migration and pluripotency. In conclusion, our results provide a new understanding of the role of Tcfap2c during different stages of germ cell development from specification to differentiation.
Description
Rights Access
Subject
development
TCFAP2C
germ cells