Effect of kinase-negativeEGFR gene on differentiation of embryonic germ cell line EG4

EG4 cells derived from primordial germ cells (PGCs) of 10.5 d post coitum 129/svJ mouse embryos can be used as a model system forin vitro differentiation study due to their pluripotential development ability. EG4 cell lines with stable expression of kinase-negativeEGFR cDNA, designated EG4-EGFR ^d , were generated by gene transfection. We found that: (i)EG4-EGFR ^d cells share the similar morphology and growing character with wildtype cells that can maintain undifferentiated state in long term culture. (ii) Treatment of EG4 cells with RA resulted in differentiation of adipocyte, while in mutant clones ofEG4-EGFR ^d , adipocytes were sparse or absent under the same condition, indicating the role of EGFR expressed during adipocyte development. (iii) Histological analysis showed that predominant tissues in teratocarcinomas derived fromEG4-EGFR ^d cells and wildtype cells are different. A large amount of undifferentiated cells was present in those coming from mutant cell clones. In addition some cardiac and skeletal muscles are prominently differentiated cell types. EG4 wildtype cells produced multiple differentiated cell types of three primary germ layers such as cartilage, epithelia and neural tube. These studies suggested that EGFR-dependent differentiation was inhibited in kinase-negative EG4 cells.