Goomer R S, Holst B D, Wood I C, Jones F S, Edelman G M
Department of Neurobiology, Scripps Research Institute, La Jolla, CA 92037.
Proc Natl Acad Sci U S A. 1994 Aug 16;91(17):7985-9. doi: 10.1073/pnas.91.17.7985.
Previous studies have shown that in vitro expression of the neural cell adhesion molecule (N-CAM) can be regulated by the products of homeobox genes HoxB9, -B8, and -C6. N-CAM is a Ca(2+)-independent immunoglobulin-related CAM that plays an important role in neural development. In the present study, we investigated whether the liver cell adhesion molecule (L-CAM) a member of the Ca(2+)-dependent CAM family (cadherins) is also regulated by homeobox-containing genes. In transient cotransfection experiments of NIH 3T3 cells, we observed that both HoxD9 and liver-enriched POU-homeodomain transcription factor, HNF-1, activated chloramphenicol acetyltransferase gene reporter constructs containing the L-CAM promoter and an enhancer present in the second intron of the chicken L-CAM gene. Using electrophoretic mobility-shift assays, we found that components of cell extracts from NIH 3T3 cells transfected with HoxD9 bound to a small region of the L-CAM enhancer having a consensus sequence that is a putative binding site for HNF-1. Components of extracts from the chicken hepatoma cell line LMH that had been transfected with an HNF-1 expression vector also bound to this same site. In nuclear run-on experiments with nuclei from LMH cells that were transfected with expression vectors for HoxD9 or HNF-1, L-CAM RNA levels were increased 33-fold and 4-fold respectively. Using the same run-on procedure, it was confirmed that nuclei prepared from normal embryonic chicken liver cells expressed the RNAs for HoxD9, HNF-1, and L-CAM. Taken together with previous observations, these data raise the possibility that homeobox-containing genes will have a widespread role in the place-dependent expression of CAMs belonging both to immunoglobulin-related and to cadherin families.
先前的研究表明，神经细胞黏附分子（N-CAM）的体外表达可受同源框基因HoxB9、-B8和-C6产物的调控。N-CAM是一种不依赖Ca(2+)的免疫球蛋白相关细胞黏附分子，在神经发育中起重要作用。在本研究中，我们调查了肝细胞黏附分子（L-CAM），一种依赖Ca(2+)的细胞黏附分子家族（钙黏着蛋白）的成员，是否也受含同源框基因的调控。在NIH 3T3细胞的瞬时共转染实验中，我们观察到HoxD9和肝脏富集的POU-同源结构域转录因子HNF-1均激活了氯霉素乙酰转移酶基因报告构建体，该构建体含有L-CAM启动子和鸡L-CAM基因第二个内含子中的一个增强子。使用电泳迁移率变动分析，我们发现用HoxD9转染的NIH 3T3细胞提取物中的成分与L-CAM增强子的一个小区域结合，该区域具有一个共有序列，是HNF-1的一个假定结合位点。用HNF-1表达载体转染的鸡肝癌细胞系LMH提取物中的成分也与同一位点结合。在用HoxD9或HNF-1表达载体转染的LMH细胞核进行的核转录实验中，L-CAM RNA水平分别增加了33倍和4倍。使用相同的转录程序，证实从正常胚胎鸡肝细胞制备的细胞核表达HoxD9、HNF-1和L-CAM的RNA。结合先前的观察结果，这些数据增加了含同源框基因在免疫球蛋白相关和钙黏着蛋白家族的细胞黏附分子的位置依赖性表达中具有广泛作用的可能性。
