凋亡核酸内切酶EndoG在人T细胞、B细胞和NK细胞中诱导可变剪接并抑制端粒酶催化亚基的活性

Induction of Alternative Splicing and Inhibition of Activity of Telomerase Catalytic Subunit by Apoptotic Endonuclease EndoG in Human T, B, and NK Cells.

作者信息

Zhdanov D D, Gladilina Yu A, Pokrovskaya M V, Aleksandrova S S, Grishin D V, Podobed O V, Sokolov N N

机构信息

Laboratory of Medical Biotechnology, V. N. Orekhovich Research Institute of Biomedical Chemistry, Moscow, Russia.

出版信息

Bull Exp Biol Med. 2018 Mar;164(4):478-482. doi: 10.1007/s10517-018-4016-y. Epub 2018 Mar 5.

DOI:10.1007/s10517-018-4016-y

PMID:29504103

Abstract

The effect of apoptotic endonuclease EndoG on alternative splicing of mRNA of human telomerase catalytic subunit hTERT (human telomerase reverse transcriptase) and telomerase activity in normal human lymphocytes were studied. Human CD4, CD8, B, and NK cells were transfected with a plasmid pEndoG-GFP containing EndoG gene or control plasmid pGFP. The levels of mRNA of EndoG or hTERT splicing variants were analyzed by real-time PCR. Protein content was assessed by Western blotting. Telomerase activity was measured by the telomere repeats amplification protocol. EndoG overexpression reduced the expression of active full-length hTERT and increased the expression of inactive splice variant. Shifted balance of hTERT splice variants in cells led to a significant decrease in telomerase activity within 72 h after transfection.

摘要

研究了凋亡核酸内切酶EndoG对正常人淋巴细胞中人端粒酶催化亚基hTERT（人端粒酶逆转录酶）mRNA可变剪接及端粒酶活性的影响。将含EndoG基因的质粒pEndoG-GFP或对照质粒pGFP转染人CD4、CD8、B和NK细胞。通过实时PCR分析EndoG或hTERT剪接变体的mRNA水平。通过蛋白质印迹法评估蛋白质含量。采用端粒重复序列扩增法测定端粒酶活性。EndoG过表达降低了活性全长hTERT的表达，并增加了无活性剪接变体的表达。细胞中hTERT剪接变体平衡的改变导致转染后72小时内端粒酶活性显著降低。