title: Involvement of doublesex and mab-3-related transcription factors in human female germ cell development demonstrated by xenograft and interference RNA strategies.
abstract: We identified three doublesex and mab-3-related transcription factors (DMRT) that were sexually differentially expressed in human fetal gonads and present in the ovaries at the time of meiotic initiation. These were also identified in murine embryonic female germ cells. Among these, we focused on DMRTA2 (DMRT5), whose function is unknown in the developing gonads, and clarified its role in human female fetal germ cells, using an original xenograft model. Early human fetal ovaries (8-11 weeks post-fertilization) were grafted into nude mice. Grafted ovaries developed normally, with no apparent overt changes, when compared with ungrafted ovaries at equivalent developmental stages. Appropriate germ cell density, mitotic/meiotic transition, markers of meiotic progression and follicle formation were evident. Four weeks after grafting, mice were treated with siRNA, specifically targeting human DMRTA2 mRNA. DMRTA2 inhibition triggered an increase in undifferentiated FUT4-positive germ cells and a decrease in the percentage of meiotic γH2AX-positive germ cells, when compared with mice that were injected with control siRNA. Interestingly, the expression of markers associated with pre-meiotic germ cell differentiation was also impaired, as was the expression of DMRTB1 (DMRT6) and DMRTC2 (DMRT7). This study reveals, for the first time, the requirement of DMRTA2 for normal human female embryonic germ cell development. DMRTA2 appears to be necessary for proper differentiation of oogonia, prior to entry into meiosis, in the human species. Additionally, we developed a new model of organ xenografting, coupled with RNA interference, which provides a useful tool for genetic investigations of human germline development.
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France AP-HP, University Paris-Sud, Reproductive Biology Unit, Clamart F-92140, France.
authors.forename: Marine
authors.initials: M
authors.lastname: Poulain
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France AP-HP, University Paris-Sud, Reproductive Biology Unit, Clamart F-92140, France.
authors.forename: Nelly
authors.initials: N
authors.lastname: Frydman
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Tourpin
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Muczynski
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: Vincent
authors.initials: V
authors.lastname: Mucsynski
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: Benoit
authors.initials: B
authors.lastname: Souquet
authors.affiliation: AP-HP, University Paris-Sud, Department of Obstetrics and Gynecology, Clamart F-92140, France.
authors.forename: Alexandra
authors.initials: A
authors.lastname: Benachi
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: René
authors.initials: R
authors.lastname: Habert
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France.
authors.forename: Virginie
authors.initials: V
authors.lastname: Rouiller-Fabre
authors.affiliation: University Paris Diderot, Sorbonne Paris Cité, Laboratory of Development of the Gonads, Unit of Stem Cells and Radiation, UMR 967, Fontenay aux Roses F-92265, France CEA, DSV, iRCM, SCSR, LDG, Fontenay aux Roses F-92265, France INSERM, Unité 967, Fontenay aux Roses F-92265, France University Paris-Sud, UMR 967, Fontenay aux Roses F-92265, France gabriel.livera@cea.fr.
authors.forename: Gabriel
authors.initials: G
authors.lastname: Livera
mesh_terms: D000818:Animals; D002454:Cell Differentiation / Q000235:genetics; D053487:DEAD-box RNA Helicases / Q000378:metabolism; D005260:Female; D005647:Fucosyltransferases / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D006657:Histones / Q000378:metabolism*; D006801:Humans; D016256:Lewis X Antigen / Q000378:metabolism*; D051379:Mice; D016513:Mice, SCID; D010053:Ovary / Q000254:growth & development / Q000637:transplantation*; D010063:Ovum / Q000166:cytology* / Q000254:growth & development; D034622:RNA Interference; D034741:RNA, Small Interfering; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics*; D014183:Transplantation, Heterologous
chemical_list: C000598461:DMRTA2 protein, human; C468783:H2AX protein, human; D006657:Histones; D016256:Lewis X Antigen; D034741:RNA, Small Interfering; D014157:Transcription Factors; C066693:FUT4 protein, human; D005647:Fucosyltransferases; C498288:DDX4 protein, human; D053487:DEAD-box RNA Helicases
keywords: DMRT; human germ line; ovary development; siRNA; xenograft