Jablonski Elizabeth M, Webb Ashley N, McConnell Nisha A, Riley Marcus C, Hughes Francis M
Department of Biology, University of North Carolina, Charlotte, NC 28223, USA.
Am J Physiol Cell Physiol. 2004 Apr;286(4):C975-85. doi: 10.1152/ajpcell.00180.2003. Epub 2003 Nov 26.
Apoptosis is characterized by a conserved series of morphological events beginning with the apoptotic volume decrease (AVD). This study investigated a role for aquaporins (AQPs) during the AVD. Inhibition of AQPs blocked the AVD in ovarian granulosa cells undergoing growth factor withdrawal and blocked downstream apoptotic events such as cell shrinkage, changes in the mitochondrial membrane potential, DNA degradation, and caspase-3 activation. The effects of AQP inhibition on the AVD and DNA degradation were consistent in thymocytes and with two additional apoptotic signals, thapsigargin and C(6)-ceramide. Overexpression of AQP-1 in Chinese hamster ovary (CHO-AQP-1) cells enhanced their rate of apoptosis. The AVD is driven by loss of K(+) from the cell, and we hypothesize that after the AVD, AQPs become inactive, which halts further water loss and allows K(+) concentrations to decrease to levels necessary for apoptotic enzyme activation. Swelling assays on granulosa cells, thymocytes, and CHO-AQP-1 cells revealed that indeed, the shrunken (apoptotic) subpopulation has very low water permeability compared with the normal-sized (nonapoptotic) subpopulation. In thymocytes, AQP-1 is present and was shown to colocalize with the plasma membrane receptor tumor necrosis factor receptor-1 (TNF-R1) both before and after the AVD, which suggests that this protein is not proteolytically cleaved and remains on the cell membrane. Overall, these data indicate that AQP-mediated water loss is important for the AVD and downstream apoptotic events, that the water permeability of the plasma membrane can control the rate of apoptosis, and that inactivation after the AVD may help create the low K(+) concentration that is essential in apoptotic cells. Furthermore, inactivation of AQPs after the AVD does not appear to be through degradation or removal from the cell membrane.
细胞凋亡的特征是一系列保守的形态学事件，始于凋亡体积减小（AVD）。本研究调查了水通道蛋白（AQP）在AVD过程中的作用。抑制AQP可阻断经历生长因子撤除的卵巢颗粒细胞中的AVD，并阻断下游凋亡事件，如细胞收缩、线粒体膜电位变化、DNA降解和半胱天冬酶-3激活。AQP抑制对AVD和DNA降解的影响在胸腺细胞中以及与另外两种凋亡信号（毒胡萝卜素和C(6)-神经酰胺）的作用一致。在中国仓鼠卵巢（CHO-AQP-1）细胞中过表达AQP-1可提高其凋亡率。AVD是由细胞内K(+)的流失驱动的，我们推测在AVD之后，AQP变得无活性，这会阻止进一步的水分流失，并使K(+)浓度降低到凋亡酶激活所需的水平。对颗粒细胞、胸腺细胞和CHO-AQP-1细胞的肿胀试验表明，与正常大小（非凋亡）亚群相比，萎缩（凋亡）亚群的水通透性确实非常低。在胸腺细胞中，AQP-1存在，并且在AVD前后均显示与质膜受体肿瘤坏死因子受体-1（TNF-R1）共定位，这表明该蛋白没有被蛋白水解切割，而是保留在细胞膜上。总体而言，这些数据表明，AQP介导的水分流失对AVD和下游凋亡事件很重要，质膜的水通透性可以控制凋亡速率，并且AVD后失活可能有助于产生凋亡细胞中必不可少的低K(+)浓度。此外，AVD后AQP的失活似乎不是通过降解或从细胞膜上移除。
