MPF components and meiotic competence in growing pig oocytes

Abstract

Growing pig oocytes (< or = 90 microns in diameter) are unable to resume meiosis in vitro. The objective of our present experiments has been to identify the reasons for meiotic competence in these cells. By comparing histone H1 kinase activity in growing and fully grown oocytes we demonstrate that incompetence is associated with an inability to activate H1 kinase in growing oocytes. Immunoblotting was used to determine whether this kinase inactivity resulted from a lack of either p34cdc2 protein or B-type cyclin. The results established that each of these cell cycle molecules are present in comparable amounts in both growing and fully grown oocytes. In the third series of studies experiments were carried out in an attempt to induce p34cdc2 activation during growth. Treatment with okadaic acid, an inhibitor of phosphatase 1 and 2A known to stimulate and accelerate the transition into M-phase of the meiotic cycle in a number of different species, was able to induce p34cdc2 kinase activity and facilitated the G2- to M-phase in growing oocytes. We conclude that although growing oocytes in pigs have sufficient key cell cycle components for the G2 to M transition, they remain incapable of converting these components to active maturation-promoting factor (MPF) until growth is virtually completed. Inhibition of phosphatase 1 or 2A induces the formation of active MPF during growth by an as yet unidentified pathway.

Size of the donor follicle, but not stage of reproductive cycle or seasonality, influences meiotic competency of selected domestic dog oocytes

“…The incidence of Mil (15%-30%) in smaller size follicles was quite comparable to a host of other reports for this species (Nickson et al, 1993;Yamada et al, 1993;Otoi et al, 2001;Willingham-Rocky et al, 2003). At the same time, the positive impact of larger follicle size in the dog was analogous to findings in other mammals, including the goat (De Smedt et al, 1994;Crozet et al, 1995), pig (Christmann et al, 1994), cow (Lonergan et al, 1994;Pavlok et al, 1997), and horse (Goudet et al, 1997). In fact, the common denominator for all of these species is that oocytes have more capacity to complete nuclear maturation when the host follicle is >2 mm in diameter.…”

“…MPF activation is regulated by its subunit concentration, intracellular localization and the phosphorylation-dephosphorylation of specific residues (Norbury and Nurse, 1992;Mothk et al, 1998;Abrieu et al, 2001). It has been shown that meiotically incompetent oocytes (i.e., growing or fully grown oocytes obtained from small follicles) are incapable of activating MPF due to the lack of (1) one of the MPF subunits (Chesnel and Eppig, 1995;de Vantéry et al, 1996;Mitra and Schultz, 1996), or (2) upstream regulators driving the cell cycle (Christmann et al, 1994;Dedieu et al, 1998). Mitogen activated protein kinase (MAPK) also has been shown to be associated with events leading to nuclear maturation completion, including microtubule dynamics, spindle assembly, chromosomal condensation and maintaining oocytes at the Mil stage (Kotani and Yamashita, 2002;Sun et al, 2002).…”

“…Perhaps more significant is follicular size. In other mammals, oocyte meiotic and developmental competence is progressively acquired during follicular growth (Christmann et al, 1994;DeSmedtetal., 1994;Lonerganetal., 1994;Crozetetal., 1995;Goudet et al, 1997;Pavlok et al, 1997). For example, in the goat, oocytes from follicles of 0.5-0.8 mm in diameter are only able to begin meiotic resumption, whereas those from follicles of 1•1.8 mm are capable of meiotic progression to metaphase I (MI), while those from follicles >3 mm are able to reach Mil (De Smedt et al, 1994).…”

Calcium Release at Fertilization: Artificially Mimicking the Oocyte's Response to Sperm.

“…Phosphorylation of histone H1 by cellular extracts forms the basis of an assay used to determine the level of MPF activity during meiosis and mitosis [90]. Changes in the level of MPF activity have been found to be similar during maturation of mouse [96,97], rabbit [98], goat [99], cattle [100][101][102] and pig [103][104][105][106][107] oocytes. Numerous studies have found that the level of MPF activity is relatively low at the germinal vesicle stage and steadily increases as meiosis progresses to the first metaphase (M-I) stage.…”

Calcium and meiotic maturation of the mammalian oocyte

“…In support of this hypothesis, incompetent oocytes show very different patterns of both polypeptide synthesis (McGaughey et al, 1979) and protein phosphorylation (Bornslaeger et al, 1988) compared with their competent counterparts. Moreover, although some MPF proteins are present in meiotically incompetent growing pig oocytes, they are incapable of being activated until the oocyte is fully grown (Christmann et al, 1993).…”