Changing expression and subcellular distribution of karyopherins during murine oogenesis

Mammalian oocyte growth and development is driven by a strict program of gene expression that relies on the timely presence of transcriptional regulators via nuclear pores. By targeting specific cargos for nucleo-cytoplasmic transport, karyopherin (KPN) proteins are key to the relocation of essential transcription factors and chromatin-remodelling factors into and out of the nucleus. Using multiple complementary techniques, here we establish that KPNA genes and proteins are dynamically expressed and relocalised throughout mouse oogenesis and folliculogenesis. Of the KPNAs examined (Kpna1, Kpna2, Kpna3, Kpna4, Kpna6, Kpna7, Kpnb1, Ipo5 and Xpo1), all were expressed in the embryonic ovary with up-regulation of protein levels concomitant with meiotic entry for KPNA2, accompanied by the redistribution of the cellular localisation of KPNA2 and XPO1. In contrast, postnatal folliculogenesis revealed significant up-regulation of Kpna1, Kpna2, Kpna4, Kpna6 and Ipo5 and down-regulation of Kpnb1, Kpna7 and Xpo1 at the primordial to primary follicle transition. KPNAs exhibited different localisation patterns in both oocytes and granulosa cells during folliculogenesis, with three KPNAs – KPNA1, KPNA2 and IPO5 – displaying marked enrichment in the nucleus by antral follicle stage. Remarkably, varied subcellular expression profiles were also identified in isolated pre-ovulatory oocytes with KPNAs KPNA2, KPNB1 and IPO5 detected in the cytoplasm and at the nuclear rim and XPO1 in cytoplasmic aggregates. Intriguingly, meiotic spindle staining was also observed for KPNB1 and XPO1 in meiosis II eggs, implying roles for KPNAs outside of nucleo-cytoplasmic transport. Thus, we propose that KPNAs, by targeting specific cargoes, are likely to be key regulators of oocyte development.

Microanatomical Study of Embryonic Gonadal Development in Japanese Quail (Coturnix japonica)

Gonadal development of quail embryos was examined histologically using histological and histochemical methods. In the present study, quail embryos were studied at various stages of incubation period based on phases of gonadogenesis. Germ cell migration was observed on day 3-4 but gonadal differentiation and gonadal function were observed on day 6–8 and day 11–14, respectively. During germ cell migration, quail primordial germ cells (qPGCs) were successfully detected in both left and right genital ridges as well as the dorsal mesentery by lectin histochemistry. Unexpectedly, qPGCs-like cells were found next to the neural tube by Mallory-AZAN stain. During gonadal differentiation, embryonic sex can be distinguished histologically since day 8 of incubation. Embryonic testis exhibited a thin cortex, whereas embryonic ovary exhibited a thick cortex. Testicular cord formation was found in the medulla of embryonic testes while the lacunae and fat-laden cells were found in the medulla of embryonic ovary during gonadal function. This is the first report on a comparison of phases of gonadogenesis and histochemical study of quail embryonic gonads in both sexes.

Functional differentiation of chick gonads following depletion of primordial germ cells

The endocrine capacity of embryonic chick gonads depleted in germ cells was compared to that of controls to determine whether the somatic elements of germ-cell-depleted gonad$ will undergo normal functional sex differentiation. Primordial germ cells were removed from early embryos, Hamburger and Hamilton stages 7-11, by excision of the anterior germinal crescent. Embryos were sacrificed at 14 days of incubation, and their gonads were analysed for functional differentiation by: (1) electron microscopy to detect ultrastructural cellular morphology characteristic of steroid-secreting cells; (2) growth in cell culture to detect development of characteristic cell morphologies; (3) radioimmunoassay of cell-culture media to detect androgens and oestrogens (androstenedione and oestradiol 17 β) secreted by gonadal cells; and (4) measurement of steroid levels produced by cultures treated with human chorionic gonadotropin (hCG) to detect the ability of gonadal cells to increase steroid production in response to gonadotropin stimulation. As a bioassay of gonadal endocrine activity, a gross morphological analysis was performed on the genital ducts, the development of which is ovary-dependent in females and testis-dependent in males. This study demonstrated that both male and female embryonic gonads exhibit normal functional differentiation following a significant reduction in the number of primordial germ cells. These results confirm and extend our previous finding that morphological differentiation of sterile embryonic gonads is normal (McCarrey & Abbott, 1978). It is concluded frofn the present study that a normal complement of germ cells is not essential to either morphological or functional sex differentiation of the somatic elements of the embryonic ovary or testis, thus arguing against any inductive role for the germ cells in this process.