Jenkins James, Papkovsky Dmitri B, Dmitriev Ruslan I
School of Biochemistry and Cell Biology, University College Cork, Western Road, Cork, Ireland.
School of Biochemistry and Cell Biology, University College Cork, Western Road, Cork, Ireland
Biochem J. 2016 Aug 15;473(16):2507-18. doi: 10.1042/BCJ20160477. Epub 2016 Jun 17.
The mammalian SPCA1 and SPCA2 ATPases localize in membranes of the secretory pathway and transport ions of Ca(2+) and Mn(2+) The role of tissue-specific SPCA2 isoform, highly expressed in lungs, mammary gland and gastrointestinal tract, is poorly understood. To elucidate the function of SPCA2, we studied human colon cancer HCT116 cells, grown under ambient and decreased O2 levels. We found that in contrast with other Ca(2+)-ATPase isoforms the expression of SPCA2 was up-regulated under hypoxia (3% O2), in both adherent (2D) and spheroid (3D) cultures. In spheroids, experiencing lowest O2 levels (30-50 μM, measured by phosphorescence lifetime imaging microscopy), we observed lower staining with reactive oxygen species (ROS)-specific fluorescent probe, which correlated with increased SPCA2. However, SPCA2 expression was up-regulated in cells exposed to reactive oxygen and nitrogen species donors, and when grown at higher density. We noticed that the culture exposed to hypoxia showed overall increase in S phase-positive cells and hypothesized that SPCA2 up-regulation under hypoxia can be linked to Mn(2+)-dependent cell cycle arrest. Consequently, we found that SPCA2-transfected cells display a higher number of cells entering S phase. Altogether, our results point at the important role of SPCA2 in regulation of cell cycle in cancer cells.
哺乳动物的SPCA1和SPCA2 ATP酶定位于分泌途径的膜中，并运输Ca(2+)和Mn(2+)离子。在肺、乳腺和胃肠道中高表达的组织特异性SPCA2同工型的作用尚不清楚。为了阐明SPCA2的功能，我们研究了在环境氧水平和降低的氧水平下培养的人结肠癌HCT116细胞。我们发现，与其他Ca(2+)-ATP酶同工型不同，在缺氧条件下（3% O2），SPCA2的表达在贴壁（二维）和球体（三维）培养中均上调。在球体中，经历最低氧水平（通过磷光寿命成像显微镜测量为30 - 50 μM）时，我们观察到活性氧（ROS）特异性荧光探针的染色较低，这与SPCA2增加相关。然而，在暴露于活性氧和氮供体的细胞中以及在更高密度下生长时，SPCA2表达上调。我们注意到暴露于缺氧的培养物中S期阳性细胞总体增加，并推测缺氧条件下SPCA2上调可能与Mn(2+)依赖性细胞周期停滞有关。因此，我们发现转染SPCA2的细胞进入S期的细胞数量更多。总之，我们的结果表明SPCA2在癌细胞细胞周期调控中起重要作用。
