In search of mechanisms of function of NAG7 gene, a powerful new tool for the unambiguous characterization of gel-separated proteins is accomplished by the combination of mass spectrometry and sequence database searching. NAG7, a novel putative tumor suppressor gene, located on 3p25.3, was introduced into HNE1 cells by liposome transfection. We used two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) to identify proteins that were overexpressed in NAG7 transfected cells. After staining and image analysis, spots of interest were isolated and subjected to mass spectrometry (MS). We found nine proteins, which up-regulated in NAG7 transfected cells and be identified by MS. These proteins included growth arrest specific protein, DNA binding protein, c-myc promoter-binding protein and caspase 6 etc., which involved in cell cycling, transcription regulation, and apoptosis. NAG7 may exert the function by up-regulating the expression of these proteins.