Cloning of porcine signal transducer and activator of transcription 3 cDNA and its expression in reproductive tissues

Abstract

The signal transducer and activator of transcription 3 (Stat3) protein is a member of the Stat family that has a variety of biological functions including cell growth, anti-apoptosis, and cell motility, depending on the cell type and stimulus. Recent studies have suggested that Stat3 plays an important role in embryo development. Although the Stat3 gene has been cloned in humans, mice, cow, and rats, its sequence in pigs is unknown. In the present study, the 2476 bp Stat3 cDNA was cloned using real time reverse transcriptase (RT)-PCR. Comparison of sequences across species revealed that the porcine Stat3 cDNA is 93 and 90% homologous to human and mouse respectively. To study the expression pattern of Stat3, RNA and protein were isolated from heart, lung, kidney, ovary, oviduct, and uterus tissues. RT-PCR and western blot indicated that Stat3 is expressed in all the tissues tested, and the level of expression is relatively high in tissues from the reproductive system. In addition, immunohistochemistry studies suggested that the Stat3 protein was present in the oocyte, granulosa, theca, and interstitial cells of the ovary, the mucosal folds in the oviduct, and both the epithelium and stromal layers in the endometrium. To study whether Stat3 is functional in responding to growth factor stimulation in the ovary, granulosa cells were isolated from large follicles (O3 mm) and cultured in the presence of epidermal growth factor (EGF; 10 ng/ml) for 5, 10, 15, 30, and 60 min, following which western blots were performed using an antibody against the phosphorylated Stat3. Phosphorylated Stat3 was upregulated following 5 min of EGF challenge and was sustained during the 15-min stimulation, and decreased back to the control level following 60-min stimulation. The translocation of phosphorylated Stat3 from cytoplasm to nucleus following stimulation of EGF was also detected via immunocytochemistry. Our data suggests that Stat3 may play a role in porcine ovarian function.

Identification of Sites of STAT3 Action in the Female Reproductive Tract through Conditional Gene Deletion

“…Hormones, growth factors and cytokines all contribute to the control of follicle activation and development [2], [5]. High levels of STAT3 and activated phospho-STAT3 have been detected in the granulosa cells, oocytes and theca cells [6], [7] of growing follicles. Abundant STAT3 has been demonstrated in oocytes at all stages of follicular development, although its cytoplasmic localisation has suggested the majority is usually not transcriptionally active in oocytes [6], [8].…”

“…In porcine granulosa cells STAT3 can be phosphorylated in response to EGF treatment [7] or leptin [13] in vitro. Active DNA-binding STAT3 was identified in the granulosa cells of immature follicles from hypophysectomised rats [14], [15].…”

ERK1/2-dependent gene expression in the bovine ovulating follicle

“…Similar to STAT3 , ADAMTS1 expression increased in ERK1/2 inhibited theca cells. Previous studies have shown that ADAMTS1 has been detected in bovine theca cells and is increased by 24 h post-GnRH 61 , while STAT3 has only yet been localized in theca cells of antral follicles in swine 62 . Also, expression of EGR1 and TNFAIP6 was not affected by the inhibitor treatment in theca cells.…”

Toward building the cow folliculome

“…4C. STAT3 is a gene important for oocyte and early embryo development but in pig granulosa cells it acts as an enhancer of EGF action (Wen et al, 2006). The same pattern goes for TP53 (tumor protein p53) which is predicted as activated in the 3 different routes mainly as a response to estradiol which was the natural LH trigger in the follicle.…”