Epigenetic silencing of CREB3L1 by DNA methylation is associated with high-grade metastatic breast cancers with poor prognosis and is prevalent in triple negative breast cancers.
Epigenetic silencing of CREB3L1 by DNA methylation is associated with high-grade metastatic breast cancers with poor prognosis and is prevalent in triple negative breast cancers.
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DOI：
10.1186/s13058-016-0672-x
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发表时间：
2016-01-25
期刊：
影响因子：
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通讯作者：
Anderson DH
中科院分区：
文献类型：
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作者：
Ward AK;Mellor P;Smith SE;Kendall S;Just NA;Vizeacoumar FS;Sarker S;Phillips Z;Alvi R;Saxena A;Vizeacoumar FJ;Carlsen SA;Anderson DH
CREB3L1（cAMP 反应元件结合蛋白 3 样蛋白 1）是未折叠蛋白反应的成员，最近被确定为乳腺癌和膀胱癌的转移抑制因子。使用实时定量 PCR (qPCR) 和免疫印迹来确定组蛋白脱乙酰化和 DNA 甲基化抑制剂对乳腺癌细胞系中 CREB3L1 表达的影响。对乳腺癌细胞系和肿瘤样本进行类似分析，并使用亚硫酸氢钠转化和 DNA 测序测定 CREB3L1 基因甲基化。使用免疫组织化学来确定细胞核与细胞质 CREB3L1 蛋白。除了 CREB3L1 mRNA 表达与预后之外，还进行大型乳腺癌数据库分析以检查 CREB3L1 基因甲基化与 mRNA 表达之间的关系。这项研究表明，先前在高度转移性乳腺癌细胞系中观察到的 CREB3L1 低表达部分是由表观遗传沉默引起的。用 DNA 甲基转移酶和组蛋白脱乙酰酶抑制剂处理几种具有低 CREB3L1 表达的高度转移性乳腺癌细胞系，诱导 CREB3L1 mRNA 和蛋白质的表达。在人类乳腺肿瘤中，CREB3L1 mRNA 表达在低度和中度肿瘤中上调，最常见的是管腔和 HER2 扩增亚型。相比之下，CREB3L1 表达在高级别肿瘤中受到抑制，其缺失最常与三阴性乳腺癌 (TNBC) 相关。重要的是，肿瘤数据库的生物信息学分析支持了这些发现，CREB3L1 基因的甲基化与 TNBC 相关，并且与 CREB3L1 mRNA 表达呈强负相关。 CREB3L1 mRNA 表达降低与肿瘤分级增加和无进展生存期缩短相关。免疫组织化学分析显示，低级别乳腺肿瘤经常具有细胞核 CREB3L1 蛋白，而高级别乳腺肿瘤中 CREB3L1 位于细胞质，这表明差异定位也可能调节 CREB3L1 抑制转移的有效性。我们的数据进一步强化了 CREB3L1 作为乳腺癌转移抑制因子的作用，并证明表观遗传沉默是 CREB3L1 表达缺失的主要调节因子。我们还强调，CREB3L1 表达在许多癌症类型中经常发生改变，表明它可能在癌症进展和转移中发挥更广泛的作用。本文的在线版本 (doi:10.1186/s13058-016-0672-x) 包含补充材料，可供授权用户使用。
CREB3L1 (cAMP-responsive element-binding protein 3-like protein 1), a member of the unfolded protein response, has recently been identified as a metastasis suppressor in both breast and bladder cancer. Quantitative real time PCR (qPCR) and immunoblotting were used to determine the impact of histone deacetylation and DNA methylation inhibitors on CREB3L1 expression in breast cancer cell lines. Breast cancer cell lines and tumor samples were analyzed similarly, and CREB3L1 gene methylation was determined using sodium bisulfite conversion and DNA sequencing. Immunohistochemistry was used to determine nuclear versus cytoplasmic CREB3L1 protein. Large breast cancer database analyses were carried out to examine relationships between CREB3L1 gene methylation and mRNA expression in addition to CREB3L1 mRNA expression and prognosis. This study demonstrates that the low CREB3L1 expression previously seen in highly metastatic breast cancer cell lines is caused in part by epigenetic silencing. Treatment of several highly metastatic breast cancer cell lines that had low CREB3L1 expression with DNA methyltransferase and histone deacetylase inhibitors induced expression of CREB3L1, both mRNA and protein. In human breast tumors, CREB3L1 mRNA expression was upregulated in low and medium-grade tumors, most frequently of the luminal and HER2 amplified subtypes. In contrast, CREB3L1 expression was repressed in high-grade tumors, and its loss was most frequently associated with triple negative breast cancers (TNBCs). Importantly, bioinformatics analyses of tumor databases support these findings, with methylation of the CREB3L1 gene associated with TNBCs, and strongly negatively correlated with CREB3L1 mRNA expression. Decreased CREB3L1 mRNA expression was associated with increased tumor grade and reduced progression-free survival. An immunohistochemistry analysis revealed that low-grade breast tumors frequently had nuclear CREB3L1 protein, in contrast to the high-grade breast tumors in which CREB3L1 was cytoplasmic, suggesting that differential localization may also regulate CREB3L1 effectiveness in metastasis suppression. Our data further strengthens the role for CREB3L1 as a metastasis suppressor in breast cancer and demonstrates that epigenetic silencing is a major regulator of the loss of CREB3L1 expression. We also highlight that CREB3L1 expression is frequently altered in many cancer types suggesting that it could have a broader role in cancer progression and metastasis. The online version of this article (doi:10.1186/s13058-016-0672-x) contains supplementary material, which is available to authorized users.