[1]牛晓璐,许万福,李松玉,等.蛋白激酶D1在烟曲霉介导的NF-κB信号通路激活及转录活性中的作用[J].南方医科大学学报,2013,(03):351.
点击复制

蛋白激酶D1在烟曲霉介导的NF-κB信号通路激活及转录活性中的作用()
分享到:

《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2013年03期
页码:
351
栏目:
出版日期:
2013-03-15

文章信息/Info

Title:
Functional role of protein kinase D1 in Aspergillums fumigates-induced activation of nuclear factor-κB signal pathway and transcription
作者:
牛晓璐许万福李松玉柯志勇曾方银罗深秋邓凡
关键词:
蛋白激酶D1烟曲霉分生孢子NF-κB转录激活磷酸化
Keywords:
protein kinase D1 Aspergillus fumigatus conidia nuclear factor-κB transcriptional activity phosphorylation
摘要:
目的研究蛋白激酶D1(PKD1)在烟曲霉介导的NF-κB信号通路激活及转录活性中的作用,为进一步探讨PKD1在烟曲
霉感染引起的肺曲霉病中的作用及其机制奠定基础。方法首先将GFP、GFP-PKD1表达质粒分别转染人肺腺癌细胞A549和
HEK293细胞中,分别将灭活的烟曲霉分生孢子(1×105 CFU/ml)于不同时间处理上述两组细胞,Western blotting证实PKD1的
过表达,并检测PKD1的磷酸化活性;其次在A549细胞中分别转染GFP-PKD1、siRNA-PKD1并以灭活的烟曲霉分生孢子处理
细胞30 min,分别检测下游NF-κB通路相关信号分子的磷酸化活性;最后,将NF-κB-luc 荧光素酶报告基因及内参照报告质
粒海肾荧光素酶(pRL-SV40)共转染入表达GFP、GFP-PKD1的A549细胞中,两组细胞在有无烟曲霉分生孢子的作用下处理
24 h,收集细胞裂解液,进行双色荧光素酶活性检测。结果Western blotting证实PKD1的过表达时间依赖性地增强烟曲霉刺激
的A549细胞和HEK293 细胞中PKD1的磷酸化活性及NF-κB通路中IКB和p65(pS276)的磷酸化活性,反之,敲低PKD1的表
达则抑制烟曲霉介导的NF-κB通路中IКB和p65(pS276)的磷酸化活性。过表达PKD1明显增加NF-κB-luc荧光素酶的转录激
活活性。结论PKD1可能在烟曲霉刺激的NF-κB通路的激活及转录活性中起重要作用。
Abstract:
Objective To explore the functional role of protein kinase D1 (PKD1) in the activation of nuclear factor-κB (NF-κB)
signal pathway and NF-κB transcription mediated by Aspergillums fumigates. Methods A549 cells and HEK293 cells were
transfected with green fluorescence protein (GFP) or GFP-PKD1 followed by treatment with 1×105 CFU/ml Aspergillus fumigatus
conidia for different time lengths. The phosphorylation levels of PKD1, IКB and p65 (pS276) in the transfected cells were
measured by Western blotting. A549 cells were transfected with GFP-PKD1 or siRNA-PKD1, and the phosphorylation of IКB
and p65 (pS276) was examined. Finally, NF-κB-luc and renilla luciferase reporter pRL-SV40 were cotransfected into GFP- or
GFP-PKD1-transfected A549 cells before exposure of the cells to Aspergillus fumigatus conidia for 24 h, and NF-κB
transcriptional activity in the cells was determined using dual-luciferase reporter assay. Results Overexpression of PKD1
significantly increased Aspergillus fumigatus conidia-stimulated phosphorylation of PKD1, IКB and p65 (pS276), whereas PKD1
knockdown by siRNA-PKD1 suppressed IКB and p65 (pS276) phosphorylation. Dual luciferase assay demonstrated that PKD1
overexpression markedly enhanced Aspergillus fumigatus-induced NF-κB transcription in A549 cells. Conclusion PKD1 may
contribute to the activation of NF-κB signal pathway and NF-κB transcription induced by Aspergillums fumigates.
更新日期/Last Update: 1900-01-01