Polyploidy or whole genome duplication (WGD) provides raw genetic materials for sequence and expression evolution of duplicate genes. However, the mode and tempo of expression divergence between WGD duplicate genes in closely related species and recurrent allopolyploids are poorly understood. Arabidopsis is a suitable system for testing the hypothesis that duplicate genes increase expression diversity and regulatory networks. In Arabidopsis, WGD occurred more than once before the split between Arabidopsis thaliana and Arabidopsis arenosa, and both natural and human-made allotetraploids are available. Comparative genomic hybridization analysis indicated that single-copy and duplicate genes after WGD were well preserved in A. thaliana and A. arenosa. Analysis of gene expression microarrays showed that duplicate genes generally had higher levels of expression divergence between two closely related species than single-copy genes. The proportion of the progenitors' duplicate genes that were nonadditively expressed in the resynthesized and natural allotetraploids was significantly higher than that of single-copy genes. Duplicate genes related to environmental stresses tended to be differentially expressed, and multicopy duplicate genes were likely to diverge expression between progenitors and in the allotetraploids. Compared with single-copy genes, duplicate genes tended to contain TATA boxes and less DNA methylation in the promoter regions, facilitating transcriptional regulation by binding transcription factors and/or cis- and trans-acting proteins. The data suggest an important role of WGD duplicate genes in modulating diverse and novel gene expression changes in response to external environmental cues and internal genetic turmoil such as recurrent polyploidy events.