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首页> 外文期刊>BMC research notes >CDK9 inhibition strategy defines distinct sets of target genes
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CDK9 inhibition strategy defines distinct sets of target genes

机译：CDK9抑制策略定义了不同的靶基因集

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Background CDK9 is the catalytic subunit of the Positive Transcription Elongation Factor b (P-TEFb), which phosphorylates the CTD of RNAPII and negative elongation factors enabling for productive elongation after initiation. CDK9 associates with T-type cyclins and cyclin K and its activity is tightly regulated in cells at different levels. CDK9 is also the catalytic subunit of TAK (Tat activating Kinase), essential for HIV1 replication. Because of CDK9′s potential as a therapeutic target in AIDS, cancer, inflammation, and cardiomyophathy it is important to understand the consequences of CDK9 inhibition. A previous gene expression profiling study performed with human glioblastoma T98G cells in which CDK9 activity was inhibited either with a dominant negative mutant form of CDK9 (dnCDK9) or the pharmacological inhibitor Flavopiridol unveiled striking differences in gene expression effects. In the present report we extended these studies by (1) using both immortalized normal human fibroblasts and primary human astrocytes, (2) eliminating potential experimental variability due to transduction methodology and (3) also modulating CDK9 activity with siRNA. Findings Striking differences in the effects on gene expression resulting from the strategy used to inhibit CDK9 activity (dnCDK9 or FVP) remain even when potential variability due to viral transduction is eliminated. siRNA mediated CDK9 knockdown in human fibroblasts and astrocytes efficiently reduced CDK9 expression and led to potent changes in gene expression that exhibit little correlation with the effects of dnCDK9 or FVP. Interestingly, HEXIM1 a validated CDK9 target gene, was found to be potently downregulated by dnCDK9, FVP and siCDK9, but the cluster of genes with expression profiles similar to HEXIM1 was small. Finally, cluster analysis of all treatments revealed higher correlation between treatments than cell type origin. Conclusion The nature of the strategy used to inhibit CDK9 profoundly affects the patterns of gene expression resulting from CDK9 inhibition. These results suggest multiple variables that affect outcome, including kinetics of inhibition, potency, off-target effects, and selectivity issues. This is particularly important when considering CDK9 as a potential target for therapeutic intervention.
机译：背景CDK9是正转录延伸因子b（P-TEFb）的催化亚基，其使RNAPII的CTD和负延伸因子磷酸化，从而能够在引发后进行有效的延伸。 CDK9与T型细胞周期蛋白和细胞周期蛋白K相关，并且其活性在不同水平的细胞中受到严格调节。 CDK9也是TAK（Tat活化激酶）的催化亚基，对于HIV1复制至关重要。由于CDK9在AIDS，癌症，炎症和心肌病中具有作为治疗靶标的潜力，因此了解CDK9抑制的后果非常重要。先前对人胶质母细胞瘤T98G细胞进行的基因表达谱研究表明，CDK9活性被显性负突变形式的CDK9（dnCDK9）或药理抑制剂Flavopiridol抑制，揭示了基因表达效果的显着差异。在本报告中，我们通过（1）使用永生化的正常人成纤维细胞和原代人星形胶质细胞扩展了这些研究，（2）消除了由于转导方法而引起的潜在实验变异，以及（3）还通过siRNA调节CDK9活性。结果即使消除了由于病毒转导引起的潜在变异性，用于抑制CDK9活性的策略（dnCDK9或FVP）对基因表达的影响仍然存在惊人的差异。 siRNA介导的人类成纤维细胞和星形胶质细胞中CDK9的敲除有效降低了CDK9的表达，并导致基因表达的强效变化，而这种变化与dnCDK9或FVP的作用几乎没有关系。有趣的是，发现HEXIM1是一种经过验证的CDK9靶基因，被dnCDK9，FVP和siCDK9强烈下调，但表达谱类似于HEXIM1的基因簇很小。最后，所有治疗的聚类分析显示，治疗之间的相关性高于细胞类型起源。结论抑制CDK9的策略性质深刻影响了CDK9抑制产生的基因表达模式。这些结果表明，多种变量会影响结果，包括抑制动力学，效价，脱靶效应和选择性问题。当将CDK9视为治疗干预的潜在靶标时，这一点尤其重要。

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