College of Biotechnology, Jiangsu University of Science and Technology, Zhenjiang 212018, China.
Jiangsu Province Key Laboratory of Animal Breeding and Molecular Design, College of Animal Science and Technology, Yangzhou University, Yangzhou 225009, China.
Int J Mol Sci. 2023 Aug 7;24(15):12516. doi: 10.3390/ijms241512516.
The role of gga-miR-31 in chicken germ cell differentiation and spermatogenesis is of significant importance. The transcriptional properties of gga-miR-31 are crucial in establishing the foundation for the formation of chicken spermatogonia stem cells and spermatogenesis. In this study, a series of recombinant vectors including varying lengths of the gga-miR-31 promoter were predicted and constructed. Through the utilization of the dual luciferase reporting system, the upstream -2180~0 bp region of gga-miR-31 was identified as its promoter region. Furthermore, it was predicted and confirmed that the activity of the gga-miR-31 promoter is increased by retinoic acid (RA). The binding of RA to the gga-miR-31 and Stra8 promoter regions was found to be competitive. Through the deletion of C-jun binding sites and the manipulation of C-jun expression levels, it was determined that C-jun inhibits the activity of the gga-miR-31 promoter. Furthermore, the combined treatment of C-jun and RA demonstrated that the positive regulatory effect of RA on the gga-miR-31 promoter is attenuated in the presence of high levels of C-jun. Overall, this study establishes a foundation for further investigation into the regulatory mechanisms of gga-miR-31 action, and provides a new avenue for inducing chicken embryonic stem cells (ESC) to differentiate into spermatogonial stem cells (SSC), and sperm formation.
gga-miR-31 在鸡生殖细胞分化和精子发生中的作用非常重要。gga-miR-31 的转录特性对于建立鸡精原干细胞和精子发生的基础至关重要。在本研究中，预测并构建了一系列包含不同长度 gga-miR-31 启动子的重组载体。通过双荧光素酶报告系统的利用，确定了 gga-miR-31 的上游-2180~0 bp 区域为其启动子区域。此外，预测并证实了视黄酸（RA）可增加 gga-miR-31 启动子的活性。发现 RA 与 gga-miR-31 和 Stra8 启动子区域的结合是竞争性的。通过删除 C-jun 结合位点和操纵 C-jun 表达水平，确定 C-jun 抑制 gga-miR-31 启动子的活性。此外，C-jun 和 RA 的联合处理表明，在高水平 C-jun 存在的情况下，RA 对 gga-miR-31 启动子的正向调节作用减弱。总之，本研究为进一步研究 gga-miR-31 作用的调控机制奠定了基础，并为诱导鸡胚胎干细胞（ESC）分化为精原干细胞（SSC）和精子形成提供了新的途径。
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