Pan Yong, Weinman Edward J, Dai Jia Le
Department of Molecular Pathology, The University of Texas MD Anderson Cancer Center, Fannin Street, Houston, Texas 77054, USA.
Breast Cancer Res. 2008;10(1):R5. doi: 10.1186/bcr1846. Epub 2008 Jan 11.
The gene encoding Na+/H+ exchanger regulatory factor 1 (NHERF1) is a putative tumor suppressor gene that harbors frequent loss of heterozygosity (LOH) and intragenic mutations in breast carcinoma. The exact biologic activity of NHERF1 in mammary glands, however, remains unclear. It was recently proposed that NHERF1 forms a ternary complex with platelet-derived growth factor receptor (PDGFR) and phosphatase and tensin homolog (PTEN), linking NHERF1 suppressor activity to PDGF-initiated phosphoinositide-3 kinase (PI3K)/PTEN signaling.
The effect of NHERF1 on the kinetics of PDGF-induced Akt activation was determined in cells with varied NHERF1 background. Levels of active Akt in mammary gland of NHERF1 knockout and wild-type mice were compared. We also examined how NHERF1 expression status affects cell sensitivity to PDGFR inhibitor. A plausible connection between NHERF1 and PTEN pathway was explored at the genetic level.
We showed that NHERF1, through its PDZ-I domain, interacts directly with the carboxyl-terminal tail of PTEN. Knocking down NHERF1 expression in Zr75.1 cells markedly delayed the turnover of PDGF-induced phospho-Akt. Conversely, NHERF1 over-expression in MCF10A cells led to accelerated phospho-Akt degradation. The slowed decay of phospho-Akt that resulted from NHERF1 loss was evident in mouse embryonic fibroblasts isolated from NHERF1 knockout mice. In agreement with this, mammary gland tissues from these mice exhibited markedly elevated phospho-Akt. The responses of breast cancer cells to PDGFR inhibition were also altered by changes in NHERF1 expression level. Zr75.1 cells with NHERF1 knockdown were more resistant to STI-571-induced apoptosis than parental cells. Similarly, over-expression of NHERF1 rendered MCF10A cells more sensitive to STI-571. NHERF1-induced apoptotic response relies on an intact PTEN pathway; over-expression of NHERF1 in MCF10A cells with PTEN knockdown did not affect STI-571 sensitivity. It was found that NHERF1 LOH-positive breast cancer cells had reduced NHERF1 expression. Interestingly, these cells more frequently had wild-type PTEN or PI3KCA gene than the LOH-negative lines.
Our data indicate that the interaction of NHERF1 with PTEN counterbalances PI3K/Akt oncogenic signaling and may affect how cells respond to PDGFR inhibition in breast cancer. The dependence of NHERF1 responses on PTEN and genetic segregation of NHERF1 and PTEN (or PI3KCA) alterations suggest that NHERF1 is an active component of the PTEN pathway. Collectively, our study indicates that the biologic activity of NHERF1 in mammary gland is related to PTEN signaling.
编码钠氢交换调节因子1（NHERF1）的基因是一种假定的肿瘤抑制基因，在乳腺癌中常发生杂合性缺失（LOH）和基因内突变。然而，NHERF1在乳腺中的确切生物学活性仍不清楚。最近有人提出，NHERF1与血小板衍生生长因子受体（PDGFR）和磷酸酶及张力蛋白同源物（PTEN）形成三元复合物，将NHERF1的抑制活性与PDGF启动的磷酸肌醇-3激酶（PI3K）/PTEN信号传导联系起来。
在具有不同NHERF1背景的细胞中测定NHERF1对PDGF诱导的Akt激活动力学的影响。比较NHERF1基因敲除小鼠和野生型小鼠乳腺中活性Akt的水平。我们还研究了NHERF1表达状态如何影响细胞对PDGFR抑制剂的敏感性。在基因水平上探索了NHERF1与PTEN途径之间可能的联系。
我们发现，NHERF1通过其PDZ-I结构域与PTEN的羧基末端尾巴直接相互作用。在Zr75.1细胞中敲低NHERF1表达显著延迟了PDGF诱导的磷酸化Akt的周转。相反，在MCF10A细胞中过表达NHERF1导致磷酸化Akt降解加速。从NHERF1基因敲除小鼠分离的小鼠胚胎成纤维细胞中，NHERF1缺失导致的磷酸化Akt衰变减慢很明显。与此一致，这些小鼠的乳腺组织中磷酸化Akt明显升高。NHERF1表达水平的变化也改变了乳腺癌细胞对PDGFR抑制的反应。NHERF1敲低的Zr75.1细胞比亲本细胞对STI-571诱导的凋亡更具抗性。同样，NHERF1的过表达使MCF10A细胞对STI-571更敏感。NHERF1诱导的凋亡反应依赖于完整的PTEN途径；在PTEN敲低的MCF10A细胞中过表达NHERF1不影响STI-571敏感性。发现NHERF1 LOH阳性乳腺癌细胞的NHERF1表达降低。有趣的是，与LOH阴性细胞系相比，这些细胞中野生型PTEN或PI3KCA基因更为常见。
我们的数据表明，NHERF1与PTEN的相互作用可平衡PI3K/Akt致癌信号传导，并可能影响乳腺癌细胞对PDGFR抑制的反应方式。NHERF1反应对PTEN的依赖性以及NHERF1与PTEN（或PI3KCA）改变的基因分离表明，NHERF1是PTEN途径的一个活性成分。总体而言，我们的研究表明NHERF1在乳腺中的生物学活性与PTEN信号传导有关。
