Functional analysis and interpretation of large-scale proteomics and gene expression data require effective use of bioinformatics tools and public knowledge resources coupled with expert-guided examination. An integrated bioinformatics approach was used to analyze cellular pathways in response to ionizing radiation. ATM, or mutated in ataxia-telangiectasia, a serine-threonine protein kinase, plays critical roles in radiation responses, including cell cycle arrest and DNA repair. We analyzed radiation responsive pathways based on 2D-gel/MS proteomics and microarray gene expression data from fibroblasts expressing wild type or mutant ATM gene. The analysis showed that metabolism was significantly affected by radiation in an ATM dependent manner. In particular, purine metabolic pathways were differentially changed in the two cell lines. The expression of ribonucleoside- diphosphate reductase subunit M2 (RRM2) was increased in ATM-wild type cells at both mRNA and protein levels, but no changes were detected in ATM-mutated cells. Increased expression of p53 was observed 30min after irradiation of the ATM-wild type cells. These results suggest that RRM2 is a downstream target of the ATM-p53 pathway that mediates radiation-induced DNA repair. We demonstrated that the integrated bioinformatics approach facilitated pathway analysis, hypothesis generation and target gene/protein identification.