摘要
To explore the expression pattern of the TRIB1 gene in yak follicles and its effect on the steroidogenesis of granulosa cells (GCs). Here, 4–5 years old female yaks were treated as the subjects. Immunohistochemically assay found that TRIB1 protein was expressed in different developmental follicles. Among different cell types of follicles, including cumulus cells (CCs), granulosa cells (GCs) and theca cells (TCs), the TRIB1 protein was most abundant in GCs (P < 0.0001). In addition, we cloned the coding sequence (CDS) of the yak TRIB1 gene, which is 1119 bp, encoding 372 amino acids (AA). The amino acid sequence homology of TRIB1 is >80% to those of other species, except for zebrafish. To further explore the function of TRIB1 in steroidogenesis, the pcDNA3.1(+)-TRIB1 eukaryotic expression vector was constructed and then transfected into GCs. The data showed that overexpression of TRIB1 significantly reduced the progesterone (P4) secretion of granulosa cells measured by ELISA assay (P < 0.05), but not Estradiol (E2) secretion. Consistently, TRIB1 gain-of-function downregulated the mRNA levels of steroidogenesis related genes steroidogenic acute regulatory protein (StAR), cytochrome P450 family 11 subfamily A member 1 (CYP11A1) and 3β-hydroxysteroid dehydrogenase (3β-HSD) (P < 0.01), while cytochrome P450 family 17 subfamily A member 1 (CYP17A1) and cytochrome P450 family 19 subfamily A member 1 (CYP19A1) had no significant difference (P > 0.05). Interestingly, mito-tracker staining showed that mitochondrial number significantly decreased in TRIB1 overexpressed GCs (P < 0.01). Further, overexpression of TRIB1 inhibited mRNA levels of mitochondrial biogenesis related genes, including Mitochondrial transcription factor (TFAM) and Peroxisome proliferator-activated receptor alpha co-activator (PPARGC1A) (P < 0.05). Conclusively, this work indicates that TRIB1 inhibited progesterone synthesis of GCs might be involved in the reduction of the mitochondria number.
摘要译文
探索牛卵泡中Trib1基因的表达模式及其对颗粒细胞（GCS）类固醇生成的影响。在这里，4-5岁的女性牛被视为受试者。免疫组织化学分析发现，在不同的发育卵泡中表达了Trib1蛋白。在包括积云细胞（CC），颗粒细胞（GCS）和Theca细胞（TCS）在内的不同细胞类型中，Trib1蛋白在GC中最丰富（P <0.0001）。此外，我们克隆了Yak Trib1基因的编码序列（CD），即1119 bp，编码372个氨基酸（AA）。除斑马鱼外，Trib1的氨基酸序列同源性> 80％。为了进一步探索Trib1在类固醇生成中的功能，构建了PCDNA3.1（+） - Trib1真核生物表达载体，然后转染到GC中。数据表明，Trib1的过表达显着降低了通过ELISA分析测量的颗粒细胞的孕酮（P4）分泌（P <0.05），但不能显着降低雌二醇（E2）分泌。一致地，Trib1功能获得的功能下调了类固醇生成相关基因的mRNA水平类固醇生成急性调节蛋白（StAR），细胞色素P450家族11个下属A成员1（CYP11A1）和3β-羟基肌动物脱氢酶（3β-HSD）（3β-HSD）（3β-HSD）（P <0.01），而细胞色素p450家族17亚家族成员1（CYP17A1）和细胞色素P450家族19个子家族A成员1（CYP19A1）没有显着差异（P> 0.05）。有趣的是，线粒体跟踪器的染色表明，在过表达的GC中，线粒体数显着降低（p <0.01）。此外，TRIB1的过表达抑制了线粒体生物发生相关基因的mRNA水平，包括线粒体转录因子（TFAM）和过氧化物酶体增殖物激活受体α共激活剂（PPARGC1A）（PPARGC1A）（P <0.05）。结论性地，这项工作表明，部落1抑制GC的孕酮合成可能与线粒体数的减少有关。
Dan Zhao[a][b][c][d];Yiling Fan[a][b][c][d];Xianrong Xiong[a][b][c][d];Shi Yin[a][b][c][d];Wei Fu[a][b][c][d];Yan Ma[a][b][c][d];Yongqi Yue[a][b][c][d];Zhidong Zhao[e];Jian Li[a][b][c][d]PersonEnvelope;Yan Xiong[a][b][c][d]PersonEnvelope. Molecular characterization of TRIB1 gene and its role in regulation of steroidogenesis in bos grunniens granulosa cells[J]. Theriogenology, 2022,191: 1-9