A study about the impaired mitochondrial membrane potential and nuclear body formation, and their relationship with the induction of cell senescence and cell death

利用荧光标记的S1-1异构体特异性抗体和PML异构体特异性抗体，对培养的人正常细胞（TIG3S）、HeLa细胞、各种正常大鼠组织和各种人肿瘤组织的核体与相应的正常组织进行检测。在年轻TIG3S细胞中，很少观察到S1-1核体，但PML2、4、5、6核体明显可见。进一步证实了S1-1和PML6核体的共定位。在HeLa细胞中几乎没有观察到S1-1核体。此外，S1-1和PML蛋白的胞质定位显著，PML2和PML6的核小体显著，PML4和PML5的核小体少见。我们在年轻的TIG3S和HeLa细胞分裂中证实了罕见的S1-1核体。此外，年轻TIG3S和HeLa细胞的S1-1p110蛋白水平非常低，相反，S1-1p130蛋白水平非常高。此外，HeLa细胞的S1-1p130水平比TIG3S细胞高3倍。PML和S1-1（RBM10）有许多被CK2和PKC磷酸化的位点，HeLa细胞的CK2活性明显高于正常细胞。因此，被CK2磷酸化（PML的S517）的PML同工型在蛋白酶体中降解。然而，S1-1p130 （RBM10变异体-1）即使在HeLa细胞中也非常稳定。给予PKC或CK2抑制剂的HeLa和TIG3S细胞显著发育S1-1核小体。这表明在活跃分裂的细胞中，CK2对S1-1核小体的磷酸化抑制了它们的组装。在存在PKC或CK2抑制剂而不存在CDK2抑制剂的情况下，HeLa或TIG3S细胞分裂受到严重损害，药物促进细胞死亡。我们的研究结果表明S1-1和PML核小体调节细胞生长并促进细胞死亡途径

With fluorescently labeled S1-1 isoform specific antibodies and PML isoform specific antibodies, we examined the nuclear bodies of cultured human normal cells (TIG3S), HeLa cells, various normal rat tissues, and various human tumor tissues with the corresponding normal tissues. In young TIG3S cells, S1-1 nuclear bodies were rarely observed, but PML2, 4, 5, 6 nuclear bodies were observed significantly. Furthermore, co-localization of S1-1 and PML6 nuclear bodies was confirmed. In HeLa cells, S1-1 nuclear bodies were hardly observed. In addition, the remarkable cytoplasmic localization of S1-1 and of PML protein, significant nuclear bodies of PML2 and PML6, and rare nuclear bodies of PML4 and PML5 were observed. We confirmed the rare S1-1 nuclear bodies in dividing young TIG3S and HeLa cells. In addition, the S1-1p110 protein levels of young TIG3S and HeLa cells were very low, on the contrary, the S1-1p130 protein was at very high levels. Furthermore, the S1-1p130 level of HeLa cells was 3-fold higher than TIG3S cells. PML and S1-1(RBM10) have many phosphorylation sites by CK2 and PKC, and HeLa cells have significantly higher activity of CK2 than normal cells. Thereby, the PML isoforms which are phosphorylated (S517 of PML) by CK2, degrade in the proteasomes. However, S1-1p130 (RBM10 variant-1) was very stable even in HeLa cells. HeLa and TIG3S cells, which administered with PKC or CK2 inhibitor, significantly developed the S1-1 nuclear bodies. This suggests that the assembly of S1-1 nuclear bodies was suppressed through their phosphorylation by CK2 in the actively dividing cells. In the presence of PKC or CK2 inhibitor, but not CDK2 inhibitor, cell division of HeLa or TIG3S cells was strongly impaired, and the drugs promoted cell death. Our results suggest that S1-1 and PML nuclear bodies regulate the cell growth and promote the cell death pathway