Current Pharmaceutical Design, 2012, 18, 000-000 1 1381-6128/12 $58.00+.00 © 2012 Bentham Science Publishers The Control of Cell Cycle in Mouse Primordial Germ Cells: Old and New Players Massimo De Felici* and Donatella Farini Department of Public Health and Cell Biology, Section of Histology and Embryology, University of Rome "Tor Vergata", Rome Abstract: The cell cycle of primordial germ cells (PGCs), the embryonic precursors of gametes, is characterized by a mitotic phase common to both sexes and a mitotic-meiotic switch in the female. In the present work, we will review the results obtained in the last dec- ade by studies aimed to clarify intrinsic and extrinsic regulatory signals of such processes, with particular reference to mouse PGCs. Be- sides providing a better understanding of how the gamete population is established in mammals, information about the players controlling the PGC cycle will be useful to clarify other intriguing aspects of germ cell biology such as the origin of germ cell tumours and the mechanisms allowing the maintenance of totipotency in the germ line. Keywords: Primordial germ cells, cell cycle, gametogenesis, embryonic germ cells, meiosis. INTRODUCTION In mammals, gametogenesis begins in the pre-gastrulating em- bryo, when the germ line is first specified in the epiblast and then determined as primordial germ cells (PGCs) in the extraembryonic mesoderm at the basis of allantois ([1] and references therein). From there, PGCs move through the forming hindgut and dorsal mesentery to colonize the developing gonadal ridges (GRs) (for a review, see [2]). In the mouse embryo, the number of PGCs in- creases by active proliferation from about 45 at 7.25 days post coi- tum (dpc), when they are at the basis of the allantois, to about 20,000 at 13.5 dpc, after their arrival in the GRs [3]. In the testis, PGCs entering mitotic arrest at G1/G0 phase at 13.5-14.5 dpc are termed prospermatogonia or gonocytes. In the ovary, at around the same time, PGCs, in some species now termed oogonia, undergo a switch between mitosis and meiosis and enter the prophase of meiosis I as primary oocytes [4]. In the female, the period of PGC/oogonia proliferation is particularly crucial since it determines the total number of oocytes to be present in an ovary. In fact, it is still widely accepted that after all PGCs/oogonia have entered meiosis, no new oocytes can be formed. On the contrary, the oocyte number is fated to progressively decrease over time leading to re- productive senescence and, specifically in humans, to the meno- pause. In both sexes, defects in PGC proliferation are likely to be at the origin of certain forms of sterility or sub fertility and to the for- mation of germ cell tumours (germinoma) within and outside the gonads. At the same time, deregulation of the mitotic cell cycle of PGCs is one of the factors involved in their in vitro transformation into the pluripotent cell lines termed embryonic germ (EG) cells (for a review, see [5]). GENERAL FEATURES OF PGC PROLIFERATION Proliferation of PGCs in the mouse embryo, from the time of their specification (around 7.25 dpc) to mitotic arrest or meiotic switch (around 13.5 dpc), was until recently generally believed to occur at a fairly constant rate of approximately 16 hr per cycle. This increases the PGC number from 40-50 to about 20,000 cells/embryo, with wide strain variation [3, 6]. Recent observations indicate, however, that between 8.0 to 9.0 dpc, the majority of PGCs are arrested at the G2 phase of the cell cycle [7]. Taking into account these results, we must reconsider that the period of active mouse PGC proliferation lasts a little more than 4 days, from 9.0 to 13.5 dpc, with a mean duration of the cell cycle of about 14 hr. *Address correspondence to this author at the Dipartimento di Sanità Pub- blica e Biologia Cellulare, Università di Roma "Tor Vergata", Via G. Mont- pellier 1, 00173 Roma, Italy; Tel: --39-06-7259 6174; Fax: --39-06-7259 6172; E-mail: defelici@uniroma2.it An intriguing characteristic of mouse PGCs, evidenced in early in vitro studies, is that they are apparently able to regulate the duration of their proliferation period by some autonomous mechanisms. This notion was mainly based on the observation that PGCs in culture stop proliferating at a time corresponding to the in vivo period (be- tween 12.5-14.5 dpc), independently of the influence of the somatic cell monolayer. For instance, PGCs obtained from 8.5 dpc embryos proliferate for four days while 10.5 dpc PGCs proliferate for two days only (for a review, see [4]). How PGCs are able to measure time is not known. It is likely that they do not use mechanisms based on cell division counting or shortening of telomeres [8-10], mechanisms used by other cell types [11-13]. Some evidence exists that embryonic cells generally posses an hourglass type of memory (for a review, see [11, 12]). In this model a specific event activates intracellular molecular pathways at a certain time resulting in a progressive increase or decrease in the amount of cell cycle proteins to a threshold level. Such type of timer comprises at least two func- tional modules: (1) a timing factor, which is activated in the cell by an input stimulus and that measures onwards time and (2) an effec- tor component, which stops cell proliferation and possibly initiates differentiation when the timing module indicates it is time. The input turning the hourglass in PGCs might possibly be ascribed to the action of one of the several growth factors known to be able to stimulate their proliferation (see next section). These factors should be present around 9.5 dpc in the hindgut, where PGCs reside at the beginning of their main proliferation period. The effector/s stopping mitotic cell cycle and initiating germ cell sex differentiation (enter- ing into meiosis in female or mitotic arrest in male), might be the mitotic and/or meiotic inhibiting substance (MIS) produced in the fetal gonads, but still not fully identified (see the review by [14], and below). Recent studies on the expression of cell-cycle genes and microRNAs in PGCs during development [15-17], on the ac- tion of the doublesex and mab-3 related transcription factor 1 (DMRT1, [18]) and of the RNA binding protein dead end homolog 1 (DND1, [19]) on PGC proliferation, appear to give some clues about possible cell cycle players that can function as the timing component of the hourglass (see the last sections of the present review). Finally, the kinetics of PGC proliferation has been proposed to be inherently stochastic. In such a model the decision to progress into cell cycle or arrest occurs in each cell as a consequence of a casual event whose probability is high or low in function of the development stage [8]. Stochastic models, in which casual oscilla- tions of cell cycle molecules gradually increase the probability of mitotic arrest, have been proposed [20]. These models are based on the notion that two oscillators can drive the cell cycle, one acting by degrading cyclins and the other by increasing the level of inhibitors