Calmodulin modulates Akt activity in human breast cancer cell lines
Calmodulin modulates Akt activity in human breast cancer cell lines
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DOI：
10.1007/s10549-008-0097-z
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发表时间：
2009-06-01
影响因子：
3.8
通讯作者：
Johnson, Michael D.
中科院分区：
文献类型：
--
作者：
Coticchia, Christine M.;Revankar, Chetana M.;Johnson, Michael D.
生长因子诱导Akt的激活存在于大多数人乳腺癌细胞系中，导致多种细胞结果，包括抑制细胞凋亡和提高存活率。我们证明，在大多数人乳腺癌细胞系中，表皮生长因子(EGF)启动的Akt激活是由普遍存在的钙敏感分子钙调蛋白介导的。具体地说，在雌激素受体(ER)阴性而不是ER阳性的乳腺癌细胞中，用钙调蛋白拮抗剂W-7处理时，Akt的激活被取消。在c-Myc过表达的小鼠乳腺癌细胞中，针对所有三个钙调蛋白基因的siRNAs抑制钙调蛋白的表达，从而显著抑制EGF诱导的Akt激活。此外，瞬时表达结构性活性Akt(Myr-Akt)可以克服W-7介导的Akt激活抑制。这些结果证实了钙调蛋白参与Akt途径。在某些细胞中，EGF启动的Akt信号的钙调蛋白依赖性不能用钙调蛋白的表达水平来解释。此外，这不能用ER状态或激活来解释，因为去掉雌激素和去除ER并不能将ER阳性的、W-7不敏感的MCF-7细胞株转化为钙调素依赖的信号。然而，强制过表达表皮生长因子受体(EGFR)或ErbB2确实部分恢复了钙调蛋白依赖的EGF刺激的Akt激活。这与观察到的W-7敏感细胞倾向于雌激素不依赖并表达高水平的EGFR家族成员是一致的。为了解决钙调蛋白是如何调节Akt活性的，我们观察了荧光标记的Akt和钙调蛋白在MCF-7和SK-BR-3细胞中的定位。我们发现在EGF刺激后Akt和钙调蛋白都被转移到膜上，并且这种转移到相同的亚细胞室被钙调蛋白抑制剂W-7抑制。因此，钙调蛋白可能通过调节Akt的亚细胞位置来调节Akt的活性，并且是预后不良的ER阴性乳腺癌的新靶点。
Growth factor-induced activation of Akt occurs in the majority of human breast cancer cell lines resulting in a variety of cellular outcomes, including suppression of apoptosis and enhanced survival. We demonstrate that epidermal growth factor (EGF)-initiated activation of Akt is mediated by the ubiquitous calcium sensing molecule, calmodulin, in the majority of human breast cancer cell lines. Specifically, in estrogen receptor (ER)-negative, but not ER-positive, breast cancer cells, Akt activation is abolished by treatment with the calmodulin antagonist, W-7. Suppression of calmodulin expression by siRNAs against all three calmodulin genes in c-Myc-overexpressing mouse mammary carcinoma cells results in significant inhibition of EGF-induced Akt activation. Additionally, transient expression of constitutively active Akt (Myr-Akt) can overcome W-7-mediated suppression of Akt activation. These results confirm the involvement of calmodulin in the Akt pathway. The calmodulin independence of EGF-initiated Akt signaling in some cells was not explained by calmodulin expression level. Additionally, it was not explained by ER status or activation, since removal of estrogen and ablation of the ER did not convert the ER-positive, W-7 insensitive, MCF-7 cell line to calmodulin dependent signaling. However, forced overexpression of either epidermal growth factor receptor (EGFR) or ErbB2 did partially restore calmodulin dependent EGF-stimulated Akt activation. This is consistent with observation that W-7 sensitive cells tend to be estrogen independent and express high levels of EGFR family members. In an attempt to address how calmodulin is regulating Akt activity, we looked at localization of fluorescently tagged Akt and calmodulin in MCF-7 and SK-BR-3 cells. We found that both Akt and calmodulin translocate to the membrane after EGF-stimulation, and this translocation to the same sub-cellular compartment is inhibited by the calmodulin inhibitor W-7. Thus, calmodulin may be regulating Akt activity by modulating its sub-cellular location and is a novel target in the poor prognosis, ER-negative subset of breast cancers.