College of Animal Sciences, Zhejiang University, 866 Yuhangtang Road, Hangzhou 310058, China.
Institute of Animal Husbandry and Veterinary Science, Zhejiang Academy of Agricultural Sciences, 145 Shiqiao Road, Hangzhou 310021, China.
Poult Sci. 2019 Aug 1;98(8):3257-3267. doi: 10.3382/ps/pez240.
Telomerase reverse transcriptase (TERT) is a catalytic subunit of telomerase that adds TTAGGG repeats to the 3'-overhang of telomeres. In the present study, we detected that the duck TERT (dTERT) gene was highly expressed in small intestine and kidney, followed by heart, leg muscle, spleen, pancreas, gonad, and liver at neonatal stage. From embryonic to neonatal stage, the highest dTERT mRNA in liver appeared at stage E19 (19 days at embryonic stage), while for the leg muscle the maximum expression occurred at E26. We also measured the relative telomerase activity (RTA) and relative telomere length (RTL) in the examined tissues and found that the changed tendency of RTA and RTL was not very consistent with that of TERT. In silico analysis revealed that there were three CpG islands (S1, S2, and S3) within the 5' regulatory region of the dTERT gene. Bisulfite sequencing PCR (BSP) assay showed that liver (D7, 7 days after birth) which expressed significantly lower dTERT mRNA had an obviously higher methylation level of S1 compared with small intestine (D7) or liver (E19). Quantitative real-time PCR analysis revealed that the expression of DNA methyltransferase DNMT1 in liver (D7) was significantly higher than that in small intestine (D7) or in liver (E19). In vitro, dTERT expression was upregulated and the methylation status of S1 decreased in both duck embryonic fibroblasts and small intestinal epithelial cells following treatment with the demethylation reagent, 5-aza-2'-deoxycytidine (5-aza-dC), further suggesting that dTERT is epigenetically regulated by DNA methylation. This work lays a solid foundation for further study of TERT function and regulation in avian species.
端粒酶逆转录酶（TERT）是端粒酶的催化亚基，它在端粒的 3'-突出端添加 TTAGGG 重复序列。在本研究中，我们检测到鸭 TERT（dTERT）基因在新生期小肠和肾脏中高度表达，其次是心脏、腿部肌肉、脾脏、胰腺、性腺和肝脏。从胚胎到新生期，肝脏中 dTERT mRNA 的表达在 E19 期（胚胎期第 19 天）最高，而腿部肌肉的最大表达出现在 E26 期。我们还测量了检查组织中的相对端粒酶活性（RTA）和相对端粒长度（RTL），发现 RTA 和 RTL 的变化趋势与 TERT 不太一致。计算机分析显示，dTERT 基因的 5'调控区有三个 CpG 岛（S1、S2 和 S3）。亚硫酸氢盐测序 PCR（BSP）检测显示，表达明显较低 dTERT mRNA 的肝脏（出生后第 7 天，D7）S1 的甲基化水平明显高于小肠（D7）或肝脏（E19）。定量实时 PCR 分析显示，肝脏（D7）中 DNA 甲基转移酶 DNMT1 的表达明显高于小肠（D7）或肝脏（E19）。体外，在用去甲基化试剂 5-氮杂-2'-脱氧胞苷（5-aza-dC）处理鸭胚胎成纤维细胞和小肠上皮细胞后，dTERT 的表达上调，S1 的甲基化状态降低，进一步表明 dTERT 受到 DNA 甲基化的表观遗传调控。这项工作为进一步研究禽类 TERT 功能和调控奠定了基础。
