Heterosis and polyploidy have an overwhelming influence on plant evolution. Recently, polyploid rice hybrids have been used to breed new rice varieties because they combine the advantages of both heterosis and polyploidy. In this study, we generated six rice lines: autotetraploid rice hybrids and their autotetraploid parents, diploid donors, and hybrids of the diploid donors. To investigate the molecular mechanism controlling the effects of both hybridization and polyploidization, we performed bisulfite and RNA sequencing on young panicles at the pollen meiosis stage to compare the DNA metabolomes and transcriptomes among the six rice lines. The hybrids lines were hypermethylated compared to their corresponding parents and the autotetraploid lines showed globally increased DNA methylation of their transposable elements compared to the diploid donors. The alteration in DNA methylation level corresponded to the differential gene expressions among the rice genotypes, suggesting that methylation changes induced by polyploidization and hybridization may affect gene expression. Groups of gene candidates were identified that may be associated with heterosis and polyploidy. Our results provide DNA information that can be used to investigate epigenetic modification during heterosis and polyploidy in rice.