Mammalian testis development is a complex and highly sophisticated process. To study the dynamic change of normal testis development at the transcriptional level, we investigated mouse testes at three postnatal ages: ...Mammalian testis development is a complex and highly sophisticated process. To study the dynamic change of normal testis development at the transcriptional level, we investigated mouse testes at three postnatal ages: 6 days postnatal, 4 weeks old, and 10 weeks old, representing infant (PN1), juvenile (PN2), and adult (PN3) stages, respectively. Using ultra high-throughput RNA sequencing (RNA-seq) technology, we obtained 211 million reads with a length of 35 bp. We identified 18837 genes that were expressed in mouse testes, and found that genes expressed at the highest level were involved in spermatogenesis. The gene expression pattern in PN1 was distinct from that in PN2 and PN3, which indicates that spermatogenesis has commenced in PN2. We analyzed a large number of genes related to spermatogenesis and somatic development of the testis, which play important roles at different developmental stages. We also found that the MAPK, Hedgehog, and Wnt signaling pathways were significantly involved at different developmental stages. These findings further our understanding of the molecular mechanisms that regulate testis development. Our study also demonstrates significant advantages of RNA-seq technology for studying transcriptome during development.展开更多
目的通过研究丝氨酸/精氨酸蛋白特异激酶2(serine/arginine-rich protein specific kinase 2,SRPK2)基因mRNA及其编码蛋白产物在小鼠睾丸组织中的表达特征,探讨该基因在精子发生过程中的作用及意义。方法分别采用半定量逆转录聚合酶链反...目的通过研究丝氨酸/精氨酸蛋白特异激酶2(serine/arginine-rich protein specific kinase 2,SRPK2)基因mRNA及其编码蛋白产物在小鼠睾丸组织中的表达特征,探讨该基因在精子发生过程中的作用及意义。方法分别采用半定量逆转录聚合酶链反应(RT-PCR)和蛋白免疫印迹杂交(Western blotting)分析该基因mRNA及蛋白产物在小鼠多种组织中的表达;利用实时定量PCR(real-time quantitative PCR)分析SRPK2 mRNA在不同发育阶段小鼠睾丸组织中的差异表达;应用免疫组织化学染色和间接免疫荧光技术观察SRPK2蛋白在小鼠曲精小管中的细胞定位和生精细胞内的亚细胞定位。结果半定量RT-PCR和Western blotting分析显示SRPK2 mRNA和蛋白在小鼠睾丸组织中均大量表达;实时定量PCR分析发现SRPK2 mRNA在5周及8周龄雄性小鼠睾丸组织中显著表达,具有明显的阶段特异性表达特征。免疫组织化学染色结果表明SRPK2蛋白阳性着色主要位于曲精小管中的长形精子细胞核;间接免疫荧光分析显示SRPK2蛋白定位于长形精子细胞核表面。结论 SRPK2基因在小鼠睾丸组织中大量表达,并且具有显著的阶段特异性表达特征和明确的细胞核定位,极有可能在小鼠精子发生的变态成形期参与mRNA前体分子的剪接过程,其作用机制值得进一步深入研究。展开更多
基金supported by the National Basic Research Program of China(2011CB944100,2011CB944101)the National Natural Science Foundation of China (90919024)+1 种基金the Special Foundation Work Program of Ministry of Science and Technology (2009FY120100)the National High Technology Research and Development Program of Ministry of Science and Technology of China (2012AA020409)
文摘Mammalian testis development is a complex and highly sophisticated process. To study the dynamic change of normal testis development at the transcriptional level, we investigated mouse testes at three postnatal ages: 6 days postnatal, 4 weeks old, and 10 weeks old, representing infant (PN1), juvenile (PN2), and adult (PN3) stages, respectively. Using ultra high-throughput RNA sequencing (RNA-seq) technology, we obtained 211 million reads with a length of 35 bp. We identified 18837 genes that were expressed in mouse testes, and found that genes expressed at the highest level were involved in spermatogenesis. The gene expression pattern in PN1 was distinct from that in PN2 and PN3, which indicates that spermatogenesis has commenced in PN2. We analyzed a large number of genes related to spermatogenesis and somatic development of the testis, which play important roles at different developmental stages. We also found that the MAPK, Hedgehog, and Wnt signaling pathways were significantly involved at different developmental stages. These findings further our understanding of the molecular mechanisms that regulate testis development. Our study also demonstrates significant advantages of RNA-seq technology for studying transcriptome during development.
