Abstract
Canonical TGF-β signals are transduced from the cell surface to the cytoplasm, and then translocated into the nucleus, a process that involves ligands (TGF-β1), receptors (TGFBR2/1), receptor-activated SMADs (SMAD2/3), and the common SMAD (SMAD4). Here we provide evidence that SMAD4, a core component of the canonical TGF-β signaling pathway, regulates the canonical TGF-β signaling pathway in porcine granulosa cells (GCs) through a feedback mechanism. Genome-wide analysis and qRT-PCR revealed that SMAD4 affected miRNA biogenesis in GCs. Interestingly, TGFBR2, the type II receptor of the canonical TGF-β signaling pathway, was downregulated in SMAD4-silenced GCs and found to be a common target of SMAD4-inhibited miRNAs. miR-425, the most significantly elevated miRNA in SMAD4-silenced GCs, mediated the SMAD4 feedback regulation of the TGF-β signaling pathway. This was accomplished through a direct interaction between the transcription factor SMAD4 and the miR-425 promoter, and a direct interaction between miR-425 and the TGFBR2 3′-UTR. Furthermore, miR-425 enhanced GC apoptosis by targeting TGFBR2 and the canonical TGF-β signaling pathway, which was rescued by SMAD4 and TGF-β1. Overall, our findings demonstrate that a positive feedback mechanism exists within the canonical TGF-β signaling pathway. This study also provides new insights into mechanism underlying the canonical TGF-β signaling pathway, which regulates GC function and follicular development.
Highlights
Canonical transforming growth factor (TGF)-β signals are transduced from the cell surface to the cytoplasm, and translocated into the nucleus, a process that involves ligands (TGF-β1), receptors (TGFBR2/1), receptor-activated SMADs (SMAD2/3), and the common SMAD (SMAD4)
Among the seven upregulated pre-miRNAs, six pre-miRNAs had similar expression patterns as mature miRNAs, except that mature miR-10b did not show a significant difference (Fig. 1e). These results indicate that SMAD4 is involved in the function of SMAD4-dependent miRNAs in granulosa cells (GCs)
We investigated the SMAD4 feedback mechanism regulating the canonical TGF-β signaling pathway. miR425 is a functional target of SMAD4 and a direct inhibitor of TGFBR2 in GCs, leading us to speculate that miR-425 might mediate the SMAD4 feedback regulation of the canonical TGF-β signaling pathway
Summary
Canonical TGF-β signals are transduced from the cell surface to the cytoplasm, and translocated into the nucleus, a process that involves ligands (TGF-β1), receptors (TGFBR2/1), receptor-activated SMADs (SMAD2/3), and the common SMAD (SMAD4). TGFBR2, the type II receptor of the canonical TGF-β signaling pathway, was downregulated in SMAD4-silenced GCs and found to be a common target of SMAD4-inhibited miRNAs. miR-425, the most significantly elevated miRNA in SMAD4-silenced GCs, mediated the SMAD4 feedback regulation of the TGF-β signaling pathway This was accomplished through a direct interaction between the transcription factor SMAD4 and the miR-425 promoter, and a direct interaction between miR-425 and the TGFBR2 3′-UTR. 1234567890():,; 1234567890():,; The transforming growth factor (TGF)-β signaling pathway is one of the most important signaling pathways It is expressed in all organisms but it is involved in the regulation of most cellular and molecular processes during development and disease[1,2]. We sough to understand the feedback mechanism in the canonical TGF-β signaling pathway
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