Characterization of a zinc finger protein ZAN75: Nuclear localization signal, transcriptional activator activity, and expression during neuronal differentiation of P19 cells

The ZAN75 cDNA was first identified in NIH 3T3 cells and codes for a DNA-bi nding protein with two zinc finger motifs, In this study, we characterized the nuclear localization signal of ZAN75, tested if ZAN75 regulates transcr iption, and examined its expression during embryonic development and neuron al differentiation of P19 mouse embryonal carcinoma cells. By examining the cellular localization of deletion mutants of ZAN75 fused to green fluoresc ence protein, ZAN75 was revealed to have a bipartite nuclear localization s ignal sequence upstream of the zinc finger domains. The N-terminal region o f ZAN75, when fused to the GALE DNA-binding domain, strongly activated tran scription. The expression of ZAN75 mRNA was found to be developmentally reg ulated, showing the highest expression in E11.5 embryos. In situ hybridizat ion experiments using E11.5 embryos showed a high expression of the transcr ipts in neuronal tissues such as brain and neural tube. The expression of Z AN75 was transiently increased at both the mRNA and the protein levels when P19 cells were treated with retinoic acid to induce neuronal differentiati on. Taken together, these results indicate that ZAN75 is a transcriptional activator with a bipartite nuclear localization signal and may play a role in neuronal differentiation.