Cárdenas Cesar, Liberona José Luis, Molgó Jordi, Colasante Cesare, Mignery Gregory A, Jaimovich Enrique
Centro de Estudios Moleculares de la Célula, Instituto de Ciencias Biomédicas, Facultad de Medicina, Universidad de Chile, Independencia 1027, Santiago 7, Chile.
J Cell Sci. 2005 Jul 15;118(Pt 14):3131-40. doi: 10.1242/jcs.02446.
Several lines of evidence indicate that increases in nuclear Ca(2+) have specific biological effects that differ from those of cytosolic Ca(2+), suggesting that they occur independently. The mechanisms involved in controlling nuclear Ca(2+) signaling are both controversial and still poorly understood. Using hypotonic shock combined with mechanical disruption, we obtained and characterized a fraction of purified nuclei from cultured rat skeletal myotubes. Both immunoblot studies and radiolabeled inositol 1,4,5-trisphosphate [IP(3)] binding revealed an important concentration of IP(3) receptors in the nuclear fraction. Immunofluorescence and immunoelectron microscopy studies localized type-1 and type-3 IP(3) receptors in the nucleus with type-1 receptors preferentially localized in the inner nuclear membrane. Type-2 IP(3) receptor was confined to the sarcoplasmic reticulum. Isolated nuclei responded to IP(3) with rapid and transient Ca(2+) concentration elevations, which were inhibited by known blockers of IP(3) signals. Similar results were obtained with isolated nuclei from the 1B5 cell line, which does not express ryanodine receptors but releases nuclear Ca(2+) in an IP(3)-dependent manner. Nuclear Ca(2+) increases triggered by IP(3) evoked phosphorylation of cAMP response element binding protein with kinetics compatible with sequential activation. These results support the idea that Ca(2+) signals, mediated by nuclear IP(3) receptors in muscle cells, are part of a distinct Ca(2+) release component that originates in the nucleus and probably participates in gene regulation mediated by cAMP response element binding protein.
多条证据表明，核内Ca(2+)的增加具有与胞质Ca(2+)不同的特定生物学效应，这表明它们是独立发生的。控制核Ca(2+)信号传导的机制既存在争议，目前仍知之甚少。我们采用低渗休克结合机械破碎的方法，从培养的大鼠骨骼肌管中获得并鉴定了一部分纯化的细胞核。免疫印迹研究和放射性标记的肌醇1,4,5-三磷酸[IP(3)]结合实验均显示，核部分中存在重要浓度的IP(3)受体。免疫荧光和免疫电子显微镜研究将1型和3型IP(3)受体定位在细胞核中，其中1型受体优先定位于内核膜。2型IP(3)受体局限于肌浆网。分离的细胞核对IP(3)的反应是Ca(2+)浓度迅速短暂升高，这被已知的IP(3)信号阻滞剂所抑制。从1B5细胞系分离的细胞核也得到了类似结果，该细胞系不表达兰尼碱受体，但以IP(3)依赖的方式释放核Ca(2+)。IP(3)引发的核Ca(2+)增加诱发了cAMP反应元件结合蛋白的磷酸化，其动力学与顺序激活相一致。这些结果支持了这样一种观点，即肌肉细胞中由核IP(3)受体介导的Ca(2+)信号是一种独特的Ca(2+)释放成分的一部分，该成分起源于细胞核，可能参与由cAMP反应元件结合蛋白介导的基因调控。
