Division of Geriatrics and Palliative Medicine, Jacobs School of Medicine and Biomedical Sciences, University at Buffalo, Buffalo, New York.
Veterans Affairs Western New York Healthcare System Research Service, Buffalo, New York.
J Cell Biochem. 2019 Aug;120(8):12382-12392. doi: 10.1002/jcb.28504. Epub 2019 Feb 28.
Cathepsin K (CTSK) is a secreted protease that plays an essential role in osteoclastic bone resorption and osteoporotic bone loss. We have previously shown that activator protein 1 (AP-1) stimulates CTSK promoter activity and that proximal nuclear factor of activated T cells cytoplasmic 1 (NFATc1)-binding sites play a major role in the stimulation of CTSK gene expression by receptor activator of NFκB ligand (RANKL). In the present study, we have extended these observations and further dissected the effects of transcription factors involved in the regulation of CTSK gene expression. Our aim was to investigate the cooperative interplay among transcription factors AP-1, microphthalmia-associated transcription factor (Mitf), and NFATc1, and the consequent regulatory effects on CTSK transcription. Experiments were carried out in RAW 264.7 cells, which can be readily differentiated to osteoclasts upon RANKL stimulation. Our data show that AP-1, Mitf, and NFATc1 are capable of independently stimulating CTSK promoter activity. A combination of any two factors further enhances CTSK promoter activity, with the combination of AP-1 (c-fos/c-jun) and NFATc1 inducing the largest increase. We further identify a synergistic effect when all three factors cooperate intimately at the proximal promoter region, yielding maximal transcriptional upregulation of the CTSK promoter. RANKL induces temporal localization of AP-1 and NFATc1 to the CTSK promoter. These results suggest that the interaction of multiple transcription factors mediate a maximal response to RANKL-induced CTSK gene expression.
组织蛋白酶 K (CTSK) 是一种分泌性蛋白酶,在破骨细胞骨吸收和骨质疏松性骨丢失中发挥重要作用。我们之前已经表明,激活蛋白 1 (AP-1) 刺激 CTSK 启动子活性,并且近端核因子活化 T 细胞胞浆 1 (NFATc1) 结合位点在核因子κB 配体 (RANKL) 刺激 CTSK 基因表达中起主要作用。在本研究中,我们扩展了这些观察结果,并进一步剖析了参与 CTSK 基因表达调控的转录因子的作用。我们的目的是研究参与 CTSK 转录调控的转录因子 AP-1、小眼畸形相关转录因子 (Mitf) 和 NFATc1 之间的协同相互作用,以及对 CTSK 转录的相应调节作用。实验在 RAW 264.7 细胞中进行,这些细胞在受到 RANKL 刺激后很容易分化为破骨细胞。我们的数据表明,AP-1、Mitf 和 NFATc1 能够独立地刺激 CTSK 启动子活性。两种因子的组合进一步增强了 CTSK 启动子活性,而 AP-1 (c-fos/c-jun) 和 NFATc1 的组合诱导的增加最大。当所有三个因子在近端启动子区域密切合作时,我们进一步鉴定出协同作用,从而使 CTSK 启动子的转录最大程度地上调。RANKL 诱导 AP-1 和 NFATc1 在 CTSK 启动子上的时间定位。这些结果表明,多种转录因子的相互作用介导了对 RANKL 诱导的 CTSK 基因表达的最大反应。