Center for Research in Reproductive Physiology, Department of Obstetrics, Gynecology, and Reproduction Services, Magee Women's Research Institute, University of Pittsburgh, Pittsburgh, Pennsylvania 15213, USA.
Biol Reprod. 2011 Nov;85(5):965-76. doi: 10.1095/biolreprod.111.093013. Epub 2011 Jul 6.
Within the testis, each Sertoli cell can support a finite number of developing germ cells. During development, the cessation of Sertoli cell proliferation and the onset of differentiation establish the final number of Sertoli cells and, thus, the total number of sperm that can be produced. The upstream stimulatory factors 1 and 2 (USF1 and USF2, respectively) differentially regulate numerous Sertoli cell genes during differentiation. To identify genes that are activated by USF proteins during differentiation, studies were conducted in Sertoli cells isolated from 5- and 11-day-old rats, representing proliferating and differentiating cells, respectively. Usf1 mRNA and USF1 protein levels were increased between 5 and 11 days after birth. In vitro studies revealed that USF1 and USF2 DNA-binding activity also increased at 11 days for the promoters of four potential target genes, Fshr, Gata4, Nr5a1, and Shbg. Chromatin immunoprecipitation assays confirmed that USF recruitment increased in vivo between 5 and 11 days after birth at the Fshr, Gata4, and Nr5a1 promoters. Expression of Nr5a1 and Shbg, but not of Fshr or Gata4, mRNAs was elevated in 11-day-old Sertoli cells compared with 5-day-old Sertoli cells. Transient transfection of USF1 and USF2 expression vectors up-regulated Nr5a1 and Shbg promoter activity. RNA interference assays demonstrated that USF1 and USF2 contribute to Nr5a1 and Shbg expression in differentiating cells. Together, these data indicate that increased USF levels induce the expression of Nr5a1 and Shbg during the differentiation of Sertoli cells, whereas Fshr and Gata4 expression is not altered by USF proteins during differentiation.
在睾丸中，每个支持细胞可以支持有限数量的发育中的生殖细胞。在发育过程中，支持细胞增殖的停止和分化的开始确定了支持细胞的最终数量，因此也确定了可以产生的精子总数。上游刺激因子 1 和 2（分别为 USF1 和 USF2）在分化过程中差异调节许多支持细胞基因。为了鉴定在分化过程中由 USF 蛋白激活的基因，在分别代表增殖和分化细胞的 5 天和 11 天龄大鼠的支持细胞中进行了研究。出生后 5 至 11 天，Usf1 mRNA 和 USF1 蛋白水平增加。体外研究表明，USF1 和 USF2 的 DNA 结合活性也在 11 天增加，针对四个潜在靶基因 Fshr、Gata4、Nr5a1 和 Shbg 的启动子。染色质免疫沉淀测定证实，出生后 5 至 11 天，Fshr、Gata4 和 Nr5a1 启动子中 USF 的募集增加。与 5 天龄的支持细胞相比，11 天龄的支持细胞中 Nr5a1 和 Shbg mRNA 的表达升高，但 Fshr 或 Gata4 mRNA 的表达没有升高。瞬时转染 USF1 和 USF2 表达载体上调了 Nr5a1 和 Shbg 启动子活性。RNA 干扰测定表明，USF1 和 USF2 有助于分化细胞中 Nr5a1 和 Shbg 的表达。总之，这些数据表明，USF 水平的升高诱导支持细胞分化过程中 Nr5a1 和 Shbg 的表达，而 Fshr 和 Gata4 的表达不受 USF 蛋白在分化过程中的影响。
