DNA damage is a serious threat to all living organisms and may be induced by environmental stressors. Previous studies have revealed that the tardigrade (Ramazzotius varieornatus) DNA damage suppressor protein Dsup has protective effects in human cells and tobacco. However, whether Dsup provides radiation damage protection more widely in crops is unclear. To explore the effects of Dsup in other crops, stable Dsup overexpression lines through Agrobacterium-mediated transformation were generated and their agronomic traits were deeply investigated. In this study, the overexpression of Dsup not only enhanced the DNA damage resistance at the seeds and seedlings stages, they also exhibited grain size enlargement and starch granule structure and cell size alteration by the scanning electron microscopy observation. Notably, the RNA-seq revealed that the Dsup plants increased radiation-related and abiotic stress-related gene expression in comparison to wild types, suggesting that Dsup is capable to coordinate normal growth and abiotic stress resistance in rice. Immunoprecipitation enrichment with liquid chromatography-tandem mass spectrometry (IP-LC-MS) assays uncovered 21 proteins preferably interacting with Dsup in plants, suggesting that Dsup binds to transcription and translation related proteins to regulate the homeostasis between DNA protection and plant development. In conclusion, our data provide a detailed agronomic analysis of Dsup plants and potential mechanisms of Dsup function in crops. Our findings provide novel insights for the breeding of crop radiation resistance.