Roseman University of Health Sciences, School of Pharmacy, Henderson, NV 89014, USA.
Biochem Cell Biol. 2021 Aug;99(4):508-518. doi: 10.1139/bcb-2020-0480. Epub 2021 Jan 28.
G protein coupled receptor kinase 5 (GRK5) is localized within the nucleus and moderates functions such as DNA transcription, in addition to its localization at the plasma membrane. In this report, we show that GRK5 modifies the nucleolar stress response activated by the DNA polymerase inhibitor, actinomycin D (ActD). We show an increased sensitivity to the apoptotic effects of ActD on cervical HeLa cells and the breast cancer cell line MDA MB 231 with reduced protein expression of GRK5. We also tested two types of breast cancer cells (MDA MB 231 and MCF7 cells) and found that the rate of response to ActD varied between them because they have innate differences in the protein expression of GRK5. We also found that GRK5 phosphorylates nucleophosmin (NPM1) at T199 before and during the early stages of ActD treatment. Phosphorylation at T199 increases the ability of NPM1 to interact with p14 in vitro, which may affect the protein expression levels of p14. We found that the expression levels of p14 were lower in the cells transfected with the control shRNA, but higher in cells transfected with GRK5 shRNA. Collectively, this suggests that GRK5 modifies the nucleolar stress response associated with ActD.
G 蛋白偶联受体激酶 5（GRK5）位于细胞核内，除了在质膜上定位外，还调节 DNA 转录等功能。在本报告中，我们表明 GRK5 修饰了由 DNA 聚合酶抑制剂放线菌素 D（ActD）激活的核仁应激反应。我们发现，GRK5 表达减少的宫颈 HeLa 细胞和乳腺癌 MDA MB 231 细胞对 ActD 的凋亡作用更加敏感。我们还测试了两种类型的乳腺癌细胞（MDA MB 231 和 MCF7 细胞），发现它们对 ActD 的反应率不同，因为它们在 GRK5 的蛋白表达上存在内在差异。我们还发现，GRK5 在 ActD 处理的早期之前和期间在 T199 处磷酸化核仁磷酸蛋白（NPM1）。T199 处的磷酸化增加了 NPM1 与 p14 在体外相互作用的能力，这可能会影响 p14 的蛋白表达水平。我们发现，用对照 shRNA 转染的细胞中 p14 的表达水平较低，但用 GRK5 shRNA 转染的细胞中 p14 的表达水平较高。总的来说，这表明 GRK5 修饰了与 ActD 相关的核仁应激反应。
