Identification of a calmodulin-regulated Ca2+-ATPase in the endoplasmic reticulum
Identification of a calmodulin-regulated Ca2+-ATPase in the endoplasmic reticulum
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DOI：
10.1104/pp.119.4.1165
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发表时间：
1999-04-01
期刊：
影响因子：
7.4
通讯作者：
Harper, JF
中科院分区：
文献类型：
--
作者：
Hong, BM;Ichida, A;Harper, JF
钙调素依赖性Ca ~（2+）-ATP酶是近年来在植物中发现的一个独特的钙调素依赖性ATP酶亚家族。与动物中最密切相关的泵相反，质膜型Ca 2 +-ATP酶，这个新的亚家族的成员的区别在于位于N-末端而不是C-末端的钙调蛋白调节的自身抑制剂。此外，至少有一些亚型似乎存在于非质膜位置。为了开始描述它们的功能，我们研究了拟南芥Ca 2 +-ATP酶（Arabidopsis Ca 2 +-ATPase，isoform 2 protein）的亚细胞定位。在这里，我们提供的证据表明，ACA 2 p驻留在内质网（ER）。在有和没有Mg 2+的浮力密度蔗糖梯度中，ACA 2 p与ER膜标记物和典型的“ER型”Ca 2 +-ATP酶（ACA 3 p/ECA 1 p）共分级。为了观察其亚细胞定位，在其C末端用绿色荧光蛋白标记ACA 2 p（ACA 2-GFP p）并在转基因拟南芥中表达。我们收集荧光图像从活的根细胞使用共聚焦和计算光学切片显微镜。ACA 2-GFPp表现为荧光网，与ER定位一致。此外，我们还观察到成熟表皮细胞核周围有较强的荧光，这与ACA 2 p也可能在核膜中起作用的假设一致。ER的位置使ACA 2 p与植物或动物中鉴定的所有其他钙调素调节泵不同。
A unique subfamily of calmodulin-dependent Ca2+-ATPases was recently identified in plants. In contrast to the most closely related pumps in animals, plasma membrane-type Ca2+-ATPases, members of this new subfamily are distinguished by a calmodulin-regulated autoinhibitor located at the N-terminal instead of a C-terminal end. In addition, at least some isoforms appear to reside in non-plasma membrane locations. To begin delineating their functions, we investigated the subcellular localization of isoform ACA2p (Arabidopsis Ca2+-ATPase, isoform 2 protein) in Arabidopsis. Here we provide evidence that ACA2p resides in the endoplasmic reticulum (ER). In buoyant density sucrose gradients performed with and without Mg2+, ACA2p cofractionated with an ER membrane marker and a typical "ER-type" Ca2+-ATPase, ACA3p/ECA1p. To visualize its subcellular localization, ACA2p was tagged with a green fluorescence protein at its C terminus (ACA2-GFPp) and expressed in transgenic Arabidopsis. We collected fluorescence images from live root cells using confocal and computational optical-sectioning microscopy. ACA2-GFPp appeared as a fluorescent reticulum, consistent with an ER location. In addition, we observed strong fluorescence around the nuclei of mature epidermal cells, which is consistent with the hypothesis that ACA2p may also function in the nuclear envelope. An ER location makes ACA2p distinct from all other calmodulin-regulated pumps identified in plants or animals.