Iron (Fe) is essential for plant growth, playing a key role in photosynthesis, respiration, and nitrogen fixation. Despite its abundance, Fe is not easily available for uptake by roots, making Fe deficiency a common issue that reduces crop yields. The ASR (Abscisic acid/Stress/Ripening) proteins are known to be responsive to different abiotic stresses. Two ASR proteins from rice (Oryza sativa L.), OsASR1 and OsASR5, were described as transcription factors involved in aluminum (Al) toxicity response, and were shown to be partially redundant in their function. Here we explored a possible role of ASR proteins in Fe deficiency in rice plants. We showed that rice plants silenced for ASR genes (named OsASR5-RNAi) had increased sensitivity to Fe deficiency, with early and more severe chlorosis, as well as reduced photosynthesis, stunted growth, reduced seed set and altered ionome in roots, leaves and seeds. Transcriptomic analysis indicated that roots of OsASR5-RNAi plants had similar expression of Fe uptake genes, such as OsIRT1 and OsYSL15. However, long distance phloem transporter OsYSL2 was up-regulated in OsASR5-RNAi roots to a larger extent compared to WT, suggesting ASR proteins negatively regulate OsYSL2 expression. We also identified other interesting candidate genes, such as OsZIFL2 and Thionins, that are dependent on ASR proteins for regulation under Fe deficiency. Our work demonstrated that OsASR proteins are important for proper Fe deficiency response in rice.