NSF Award
0091504
Prior to implantation of the mammalian embryo, a sequence of changes takes place in the mother's uterus in response to the female sex hormones estrogen and progesterone. The actions of estrogen and progesterone are necessary for stimulating the maternal environment to become receptive for the implanting embryo. Estrogen and progesterone elicit changes in the uterus by binding to specific receptor proteins, thereby altering the rates of transcription (synthesis) of target genes. Although many of the target genes in the mammalian uterus have not been identified, evidence indicates that several of the candidates are involved in the control of cell proliferation and differentiation. Mammalian cells use a family of proteins called cyclin and cyclin dependent kinases to regulate cell proliferation. Cell proliferation involves an orderly sequence of events in which the cellular contents are duplicated and then the cell divides in two. This cycle of duplication and division is known as the cell cycle. Progression of cells through the cell cycle involves the temporal and sequential activation of cyclin genes, as well as the inactivation of a series of crucial checkpoints. Cellular differentiation requires cessation of cell cycle transit, cell cycle exit, and selection of the differentiation pathway. The integration of positive and negative regulators, and the means by which they control cell cycle progression, are the major challenges for understanding the sequence of molecular events linking cell cycle transit with cellular differentiation. Elucidating these hormone-dependent cell cycle regulatory interactions in the maternal cells of the uterus is the central objective of this research project. The first goal of this research is to determine the temporal and spatial expression patterns of the key cell cycle regulators during hormone-induced stromal cell proliferation in the rat uterus. Time course analysis of the appearance and cell-specific distribution of the key regulatory proteins will be mapped using immunohistochemical techniques. Transit through the cell cycle will be assessed in these same rat uterine samples by measuring the entry of cells into DNA replication (duplication) and mitosis (chromosomal division). This information will be extended by studying the formation of active cyclin-cyclin dependent kinase complexes in cultured uterine stromal cells stimulated to synchronously re-enter the cell cycle by estrogen and progesterone. The function of cell cycle inhibitory proteins on cell cycle regulation will be determined by assessing the temporal appearance and changes in the amount of inhibitory proteins in hormonally stimulated stromal cells in both the uterus and in culture. These hormone-sensitive changes in cell cycle activators and inhibitors will identify the mechanistic actions of estrogen and progesterone on the formation and activation of protein complexes required to control cell cycle transit. With this information in hand it will be possible to investigate the mechanisms by which differentiation signal transducers, acting in conjunction with estrogen and progesterone, alter the progression of stromal cells through the cell cycle and re-direct cellular transit into the differentiation pathway. Completion of this study is expected to increase the current knowledge about estrogen and progesterone action in target cells. The results are also expected to provide important, new information about the endocrine control of cell proliferation and differentiation in the mammalian uterus, thereby impacting both domestic animal and wildlife reproduction.
