Function of novel gene, EAT that is early gene induced by all-trans retinoic acid in human EC cells

本研究的目的是分离维甲酸(RA)诱导人EC细胞NCR-G3分化的关键基因。我们分离到一个新的基因，EAT，它在分化的早期瞬时表达，与细胞死亡预防分子bcl2同源。通过对新基因EAT的分析，我们得到了以下结果。为了阐明EAT在正常组织和胎儿组织中的表达，我们用重组蛋白EAT制备了一种鼠单抗3A2。通过免疫组织化学研究，EAT在人类胎儿和成人的多种组织中都有表达，包括上皮细胞、造血细胞和内分泌细胞。在小鼠胚胎中，EAT在受精后迅速被诱导，在2-8-细胞期达到最高水平，然后在囊胚中下降到低于未受精卵的水平。2)我们确定了EAT是否能像报道的那样抑制化疗药物诱导的细胞凋亡。转EAT基因的细胞对CDDP或其他化疗药物表现出更高的抗药性。用顺铂和卡铂处理亲本细胞后，DNA片段化程度呈浓度依赖性。相比之下，用相同浓度的这些药物处理后，转EAT的细胞没有显示出DNA片段化。EAT转基因小鼠未出现CDDP引起的急性肾小管坏死，提示EAT可能通过其抗细胞凋亡功能在早期胚胎发育和后续细胞功能中发挥重要作用。

The aim of this study was to isolate the essential genes in retinoic acid (RA)-induced differentiation of human EC cells, NCR-G3 cell. We isolated a novel gene, EAT which transiently expressed in the early stage of differentiation and has homology with bcl-2, a cell death-preventing molecule. We obtained following results by analyzing this novel gene, EAT.To elucidate expression of EAT in normal and fetal tissue, a murine monoclonal antibody, 3A2 was generated by use of recombinant protein to EAT. By immnohistochemical studies, EAT expression was observed in a wide variety of human fetal and adult tissues, including those of epithelial, hematopoietic and endocrine cells. In murine embryos, EAT was induced rapidly after fertilization reaching maximum level at the from the 2 - to 8-cell stage and then decrease to below unfertilized egg levels in blastocyst.2) We determined whether the apoptotic cell death induced by chemotherapeutic agents could be inhibited by EAT as is reported with Bcl-2. Cells transfected with EAT showed higher resistance to CDDP or among other chemotherapeutic agents tested. DNA fragmentation of the parental cells following treatment with CDDP and carboplatin was observed in a concentration-dependent manner. In contrast, cells transfected with EAT did not show DNA fragmentation following treatment with the same concentration of these drugs. Acute tubular necrosis induced by CDDP administration did not occur in the EAT transgenic mice.These results suggest that EAT may play an important role in early embryogenesis and sequential cell function by its anti-apoptotic functions.