PMEPA1, a transforming growth factor-beta-induced marker of terminal colonocyte differentiation whose expression is maintained in primary and metastatic colon cancer.

To identify potential effectors of transforming growth factor (TGF)-beta-mediated suppression of colon cancer, we used GeneChip expression microarrays to identify TGF-beta-induced genes in VACO 330, a nontransformed TGF-beta-sensitive cell line derived from a human adenomatous colon polyp. PMEPA1 was identified as a gene highly up-regulated by TGF-beta treatment of VACO 330. Northern blot analysis confirmed TGF-beta induction of PMEPA1 in VACO 330, as well as a panel of three other TGF-beta-sensitive colon cell lines. PMEPA1 induction could be detected as early as 2 h after TGF-beta treatment and was not inhibited by pretreatment of cells with cycloheximide, suggesting that PMEPA1 is a direct target of TGF-beta signaling. Wild-type PMEPA1 and an alternative splice variant lacking the putative transmembrane domain were encoded by the PMEPA1 locus and were shown by epitope tagging to encode proteins with differing subcellular localization. Both variants were found to be expressed in normal colonic epithelium, and both were shown to be induced by TGF-beta. Consistent with TGF-beta playing a role in terminal differentiation of colonocytes, in situ hybridization of normal colonic epithelium localized PMEPA1 expression to nonproliferating, terminally differentiated epithelium located at the top of colonic crypts. Intriguingly, in situ hybridization and Northern blot analysis showed that the expression of PMEPA1 was well maintained both in colon cancer primary tumors and in colon cancer liver metastases. PMEPA1 is thus a novel TGF-beta-induced marker of a differentiated crypt cell population. Moreover, as PMEPA1 expression is maintained, presumptively in a TGF-beta-independent manner after malignant transformation and metastasis, it demonstrates that even late colon cancers retain a strong capacity to execute many steps of the normal colonic differentiation program.

为了确定转化生长因子（TGF）-β介导的结肠癌抑制的潜在效应因子，我们使用基因芯片表达微阵列来鉴定VACO 330中TGF -β诱导的基因，VACO 330是一种源自人腺瘤性结肠息肉的未转化的TGF -β敏感细胞系。PMEPA1被鉴定为在VACO 330中经TGF -β处理后高度上调的基因。Northern杂交分析证实了在VACO 330以及另外三种TGF -β敏感的结肠细胞系中TGF -β对PMEPA1的诱导作用。早在TGF -β处理后2小时就可检测到PMEPA1的诱导，并且用环己酰亚胺预处理细胞不会抑制这种诱导，这表明PMEPA1是TGF -β信号传导的直接靶标。野生型PMEPA1以及一种缺失假定跨膜结构域的可变剪接变体由PMEPA1基因座编码，并且通过表位标记显示它们编码具有不同亚细胞定位的蛋白质。这两种变体都在正常结肠上皮中表达，并且都显示可由TGF -β诱导。与TGF -β在结肠细胞终末分化中起作用一致，对正常结肠上皮的原位杂交将PMEPA1的表达定位在位于结肠隐窝顶部的非增殖性、终末分化的上皮中。有趣的是，原位杂交和Northern杂交分析表明，PMEPA1的表达在结肠癌原发肿瘤和结肠癌肝转移中都保持良好。因此，PMEPA1是一种新的由TGF -β诱导的分化隐窝细胞群的标志物。此外，由于在恶性转化和转移后PMEPA1的表达（推测是以一种不依赖TGF -β的方式）得以维持，这表明即使是晚期结肠癌也保留了执行正常结肠分化程序许多步骤的强大能力。