〔Abstract〕 To investigate the expression and interaction of activating transcription factor 3 (ATF3) and smad family member 4 (Smad4) in pterygium . Methods In this study , two sets of data GSE51995 and GSE2513 in NCBI database were analyzed by bioinformatics methods to screen out the key difference-expressed gene ATF3 in pterygium when normal conjunctiva was used as the control . The differential genes identified by bioinformatics anal- ysis were further detected by reverse transcription quantitative polymerase chain reaction (RT-qPCR) . Finally , the literatures were searched and The Gene Transcription Regulation Database ( GTRD) was used to predict whether there was a target gene Smad4 that might bind to the target gene . The expression of Smad4 in normal conjunctival and pterygium tissues was verified by RT-qPCR , and the interaction between the target protein ATF3 and Smad4 in the pterygium was investigated through Co-Immunoprecipitation (Co-IP) . Results The results of bioinformatics a- nalysis showed that compared with normal conjunctival tissue , there was significantly different expression of ATF3 gene in pterygium tissue (P < 0. 05) . RT-qPCR confirmed that ATF3 was less expressed in pterygium tissue than normal conjunctival tissue (P < 0. 001) . Genomic data from the GTRD database revealed that Smad4 contains two ATF3 binding motifs , suggesting functional interaction potential . Compared with normal conjunctiva , RT-qPCR re- vealed Smad4 downregulation in pterygium tissues (P < 0. 01) ; Co-IP demonstrated enhanced interaction between Smad4 and ATF3 in pterygium tissues following immunoprecipitation with anti-ATF3 antibodies (P < 0. 05) . Con- clusion ATF3 interacts with Smad4 in pterygium , and the low expression of both in pterygium tissues and their in- teraction may be associated with the pathogenesis of pterygium .