Role of CEACAM1 isoforms in an in vivo model of mammary morphogenesis:: mutational analysis of the cytoplasmic domain of CEACAM1-4S reveals key residues involved in lumen formation
Role of CEACAM1 isoforms in an in vivo model of mammary morphogenesis:: mutational analysis of the cytoplasmic domain of CEACAM1-4S reveals key residues involved in lumen formation
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DOI：
10.1038/sj.onc.1210577
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发表时间：
2007-12-06
期刊：
影响因子：
8
通讯作者：
Shively, J. E.
中科院分区：
文献类型：
--
作者：
Yokoyama, S.;Chen, C-J;Shively, J. E.
CEACAM1（癌胚抗原相关细胞粘附分子 1）是一种在上皮细胞中表达的 I 型跨膜糖蛋白，具有三个或四个胞外结构域 (ECD) 以及长或短的胞质结构域亚型。我们之前已经证明，四个胞外结构域，即短胞质结构域亚型 CEACAM1-4S，在乳腺形态发生的体外模型中的管腔形成中发挥着重要作用。在这项研究中，我们用 CEACAM1 的长或短胞质结构域亚型转染 MCF-7 细胞，并在 NOD/SCID 小鼠的人源化小鼠乳腺脂肪垫中培养细胞。在该体内模型中，只有长胞质结构域亚型 CEACAM1-4L 形成有腔的腺体。在揭示短胞质结构域中关键 Thr 和 Ser 残基磷酸化的其他研究的基础上，我们将磷酸化模拟突变（例如，Thr 或 Ser 变为 Asp）或无效突变（Thr 或 Ser 变为 Ala）到 CEACAM1-4S 的胞质结构域中，并在体内模型中对其进行测试。 Thr或Ser突变为Asp或双突变体Thr+Ser突变为Asp，但无效突变体除外，诱导含有中央腔的凋亡细胞的腺体形成。此外，CEACAM1-4S的磷酸化模拟突变体诱导β1-整合素的下调、β2-整合素的过度表达、抑制粘着斑激酶（pTyr-397）的磷酸化，并导致以波形蛋白、α-平滑肌肌动蛋白和β2-整合素的表达为特征的肌成纤维细胞分化，以及丰富的细胞外基质的产生。
CEACAM1 (carcinoembryonic antigen-related cell adhesion molecule 1) is a type I transmembrane glycoprotein expressed in epithelial cells with three or four extracellular domains (ECDs) and either long or short cytoplasmic domain isoforms. We have previously shown that the four extracellular domains, short cytoplasmic domain isoform, CEACAM1-4S, plays an essential role in lumen formation in an in vitro model of mammary morphogenesis. In this study, we transfected MCF-7 cells with either the long or short cytoplasmic domain isoforms of CEACAM1, and grew the cells in humanized mammary mouse fat pads in NOD/SCID mice. In this in vivo model, only the long cytoplasmic domain isoform, CEACAM1-4L, formed glands with lumen. On the basis of other studies that revealed phosphorylation of key Thr and Ser residues in the short cytoplasmic domain, we introduced phosphorylation mimic ( for example, Thr or Ser to Asp) or null ( Thr or Ser to Ala) mutations into the cytoplasmic domain of CEACAM1-4S and tested them in the in vivo model. Mutation of either Thr or Ser to Asp or the double mutant Thr+Ser to Asp, but not the null mutants, induced gland formation with a central lumen-containing apoptotic cells. Moreover, the phosphorylation mimic mutants of CEACAM1-4S induced downregulation of beta 1-integrin, overexpression of beta 2-integrin, inhibited phosphorylation of focal adhesion kinase (pTyr-397) and resulted in myo.broblast differentiation as characterized by expression of vimentin, a-smooth muscle actin and beta 2-integrin, as well as the production of abundant extracellular matrix.