ABSTRACT
Restricted expression of a mouse Vasa homolog gene (Mvh) expression is first detected in primordial germ cells (PGCs) after colonization of the genital ridges. Subsequently, Mvh is maintained until postmeiotic germ cells are formed. Here, we demonstrate that male mice homozygous for a targeted mutation of Mvh exhibit a reproductive deficiency. Male homozygotes produce no sperm in the testes, where premeiotic germ cells cease differentiation by the zygotene stage and undergo apoptotic death. In addition, the proliferation of PGCs that colonize homozygous male gonads is significantly hampered, and OCT-3/4 expression appears to be reduced. These results indicate that the loss of Mvh function causes a deficiency in the proliferation and differentiation of mouse male germ cells.
Subject(s)
RNA Helicases/metabolism , Spermatozoa/physiology , Testis/physiology , Aging/physiology , Amino Acid Sequence , Animals , Cell Differentiation , Conserved Sequence , DEAD-box RNA Helicases , Drosophila/genetics , Drosophila/physiology , Drosophila Proteins , Embryonic and Fetal Development , Heterozygote , Homozygote , Male , Meiosis , Mice , Mice, Knockout , RNA Helicases/deficiency , RNA Helicases/genetics , Reverse Transcriptase Polymerase Chain Reaction , Spermatozoa/cytology , Stem Cells/cytology , Stem Cells/physiology , Testis/embryology , Testis/growth & developmentABSTRACT
To demonstrate the cellular and subcellular localization of mouse vasa homologue protein during germ cell development, specific antibody was raised against the full-length MVH protein. The immunohistochemical analyses demonstrated that MVH protein was exclusively expressed in primordial germ cells just after their colonization of embryonic gonads and in germ cells undergoing gametogenic processes until the post-meiotic stage in both males and females. The co-culture of EG cells with gonadal somatic cells indicated inductive MVH expression caused by an intercellular interaction with gonadal somatic cells. In adult testis, MVH protein was localized in the cytoplasm of spermatogenic cells, including chromatoid bodies in spermatids, known to be a perinuclear nuage structure which includes polar granules that contain VASA protein in Drosophila.
Subject(s)
Oogenesis/physiology , RNA Helicases/biosynthesis , Spermatogenesis/physiology , Animals , Female , Germ Cells/metabolism , Gonads , Intracellular Fluid/metabolism , Male , Mice , Mice, Inbred ICRABSTRACT
With the aim of identifying the gene(s) located downstream from SRY, we transfected an ES cell line with XX karyotype, TMA-18, with a Sry DNA construct and established cell lines, TS18-1 and TS18-2, where the transfected Sry was expressed in the functional linear mRNA form. Among the five potential SRY-target genes examined, i.e., MIS, SF1, P450arom, Sox9 and WT1, only the expression of WT1 was induced de novo by the unscheduled expression of Sry in the transfected cell lines. No clear indication of Sry-induced enhancement of Sox9 expression was obtained in the present series of experiments. Function of a yet unidentified gene(s) located on the Y chromosome might be needed for the up-regulation of Sox 9 expression which takes place during the development of male gonads. Quantitative RT-PCR analysis of the patterns of WT1 expression in developing fetal gonads revealed that although both male and female fetal gonads express WT1, male gonads invariably expressed WT1 mRNA at higher levels than female ones after the Sry expression. Immunohistochemical analysis of the male fetal gonads between 10.5 and 13.5 dpc demonstrated the presence of strong WT1 immunoreactivity in Sertoli cells of the primordial testes. Suggestions were made in the past indicating that both SF1 and WT1 proteins might be active in a common pathway upstream from Sry. Our results showed that WT1 is located downstream, rather than upstream from Sry and behaves independently from SF1. Analysis using an appropriate in vitro system will be essential to understand the molecular mechanisms of SRY action within cells.
