Mutations in the thyroid transcription factor gene NKX2-1 result in decreased expression of SFTPB and SFTPC.
Mutations in the thyroid transcription factor gene NKX2-1 result in decreased expression of SFTPB and SFTPC.
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DOI：
10.1038/pr.2018.30
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发表时间：
2018-09
影响因子：
3.6
通讯作者：
Hamvas A
中科院分区：
文献类型：
--
作者：
Attarian SJ;Leibel SL;Yang P;Alfano DN;Hackett BP;Cole FS;Hamvas A
NK2同源盒1 （NKX2-1）基因突变与婴儿和儿童肺部疾病有关。我们假设正常表面活性剂基因表达的破坏与这些突变有助于观察到的呼吸表型。为了评估转激活活性，将含有表面活性剂蛋白B或C （SFTPB或SFTPC）启动子的荧光素酶报告载体与NKX2-1质粒共转染，并测量荧光素酶活性。为了评估突变蛋白与靶DNA的结合，电泳迁移转移试验（EMSA）使用代表NKX2-1一致结合序列的寡核苷酸探针标记的核蛋白进行，然后进行凝胶电泳。利用实时荧光定量PCR技术评估野生型（WT）和突变型NKX2-1过表达对SFTPB和SFTPC的影响。观察到两种突变体的SFTPB启动子的反活化降低，而L197P突变体的SFTPC启动子的反活化降低。EMSA显示两种突变与NKX2-1一致结合序列的DNA结合减少。用NKX2-1表达载体转染A549细胞，与WT相比，突变蛋白对SFTPB和SFTPC表达的刺激减弱。这些dna结合域突变破坏表面活性剂蛋白基因的转录激活，可能是表面活性剂功能破坏和随后呼吸窘迫的生物学机制。
Mutations in the NK2 homeobox 1 (NKX2–1) gene are associated with lung disease in infants and children. We hypothesize that disruption of normal surfactant gene expression with these mutations contributes to the respiratory phenotypes observed. To assess transactivational activity, cotransfection of luciferase reporter vectors containing surfactant protein B or C (SFTPB or SFTPC) promoters with NKX2–1 plasmids was performed and luciferase activity was measured. To assess the binding of mutated proteins to target DNA, electrophoretic mobility shift assays (EMSA) were performed using nuclear protein labeled with oligonucleotide probes representing NKX2–1 consensus binding sequences followed by gel electrophoresis. The effect of overexpression of wild-type (WT) and mutant NKX2–1 on SFTPB and SFTPC was evaluated with quantitative real-time PCR. Decreased transactivation of the SFTPB promoter by both mutants and decreased transactivation of the SFTPC promoter by the L197P mutation was observed. EMSA demonstrated decreased DNA binding of both mutations to NKX2–1 consensus binding sequences. Transfection of A549 cells with NKX2–1 expression vectors demonstrated decreased stimulation of SFTPB and SFTPC expression by mutant proteins compared with that of WT. Disruption of transcriptional activation of surfactant protein genes by these DNA-binding domain mutations is a plausible biological mechanism for disruption of surfactant function and subsequent respiratory distress.
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影响因子：
4.8
作者：
DEFELICE, M;DAMANTE, G;DILAURO, R
通讯作者：
DILAURO, R
影响因子：
4.7
作者：
de Filippis, Tiziana;Marelli, Federica;Persani, Luca
通讯作者：
Persani, Luca
DOI：
10.1136/adc.2009.180448
发表时间：
2011-11-01
影响因子：
4.4
作者：
Kleinlein, Barbara;Griese, Matthias;Holzinger, Andreas
通讯作者：
Holzinger, Andreas
影响因子：
4.1
作者：
Christophe-Hobertus, C;Christophe, D
通讯作者：
Christophe, D
影响因子：
5.1
作者：
Doyle, DA;Gonzalez, I;Scavina, M
通讯作者：
Scavina, M