Title: Atf3 Expression in the Corpus Luteum: Possible Role in Luteal Regression Authors
|Mao, Dagan -|
|Hou, Xiaoying -|
|Talbot, Heather -|
|Cupp, Andrea -|
|Davis, John -|
Submitted to: Molecular Endocrinology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: October 23, 2013
Publication Date: December 1, 2013
Citation: Mao, D., Hou, X., Talbot, H., Cushman, R., Cupp, A., Davis, J.S. 2013. ATF3 expression in the corpus luteum: possible role in luteal regression. Molecular Endocrinology. 27(12):2066-2079. Interpretive Summary: In mammalian females the ovarian corpus luteum (CL) secretes progesterone, a steroid hormone important in the regulation of normal reproductive cycles and the maintenance of pregnancy. In cattle, it has been known for more than 40 years that prostaglandin F2 alpha (PGF) is the hormone secreted by the uterus in the absence of pregnancy to regress the corpus luteum and initiate a new estrous cycle, making PGF a major component of almost all protocols to synchronize estrus and induce ovulation. However, the efficacy of PGF in regressing the CL is influenced by stage of the estrous cycle because the bovine CL is less responsive to PGF before day 5 of the estrous cycle. Therefore, a better understanding of the molecular pathways activated by PGF in the bovine CL could aid in improving the efficacy of PGF and decrease the number of times that a beef cow needs to be handled in protocols to synchronize estrus. In the current project, we demonstrated that Activating Transcription Factor 3 (ATF3) gene expression increased in the mid-cycle bovine CL within 1 hour after treatment with the standard dose of PGF used to synchronize estrus. The large luteal cells that are responsible for basal progesterone secretion were the site of initial expression of ATF3 in response to PGF. Expression of ATF3 in the small luteal cells only occurred in response to the immunological stimuli that are known to be responsible for cell death and structural degradation of the CL. In conclusion, the action of PGF in large luteal cells is associated with the rapid induction of ATF3, which may contribute to the reduction in steroid synthesis during CL regression. The function of ATF3 in the bovine corpus luteum before day 5 of the estrous cycle remains to be investigated.
Technical Abstract: The present study investigated the induction and possible role of activating transcription factor 3 (ATF3) in the corpus luteum. Postpubertal cattle were treated at midcycle with prostaglandin F2alpha(PGF) for 0–4 hours. Luteal tissue was processed for immunohistochemistry, in situ hybridization, and isolation of protein and RNA. Ovaries were also collected from midluteal phase and first-trimester pregnant cows. Luteal cells were prepared and sorted by centrifugal elutriation to obtain purified small (SLCs) and large luteal cells (LLCs). Real-time PCR and in situ hybridization showed that ATF3 mRNA increased within 1 hour of PGF treatment in vivo. Western blot and immunohistochemistry demonstrated that ATF3 protein was expressed in the nuclei of LLC within 1 hour and was maintained for at least 4 hours. PGF treatment in vitro increased ATF3 expression only in LLC, whereas TNF induced ATF3 in both SLCs and LLCs. PGF stimulated concentration- and time-dependent increases in ATF3 and phosphorylation of MAPKs in LLCs. Combinations of MAPK inhibitors suppressed ATF3 expression in LLCs. Adenoviral-mediated expression of ATF3 inhibited LH-stimulated cAMP response element reporter luciferase activity and progesterone production in LLCs and SLCs but did not alter cell viability or change the expression or activity of key regulators of progesterone synthesis. In conclusion, the action of PGF in LLCs is associated with the rapid activation of stress-activated protein kinases and the induction of ATF3, which may contribute to the reduction in steroid synthesis during luteal regression. ATF3 appears to affect gonadotropin-stimulated progesterone secretion at a step or steps downstream of PKA signaling and before cholesterol conversion to progesterone.