All- trans retinoic acid (atRA), bioactive component of vitamin A, plays a pivotal role in various biological processes. Essential for embryonic development, bone growth and maintenance, and immune function, atRA exerts its regulatory effects primarily through its interaction with the nuclear receptor RARα. Upon binding to RARα, atRA enters the nucleus and forms a heterodimer with RXR. This heterodimer can then interact with various transcription factors to form regulatory complexes that influence gene expression. While the role of atRA in regulating the type 2 immune response has been studied, the role atRA in Th2 cell differentiation is warranted to fully elucidate its mechanisms and potential therapeutic applications. Our study shows that atRA suppressed Th2 phenotype by down-regulating type 2 transcription factors such as Spi1 and cMaf. atRA also down-regulated IL-4 and IL-13 production in Th2 cells, potentially by up-regulating Gfi1, a transcriptional repressor. atRA increased the Gfi1 recruitment to the Il4 and Il13 promoters, along with the common enhancer Ecr. RARα, which is typically an inducer of Il4 and Il13, was observed to decrease recruitment to these loci in atRA-treated Th2 cells. Comparative gene expression analysis revealed a reduction in inflammatory responses in atRA-treated Th2 cells. Furthermore, these cells exhibited a negative correlation with epigenetic modifications, nuclear receptor activity among other biological processes. Collectively, our findings suggest that atRA can effectively suppress the Th2 phenotype in vitro, through the regulation of key type 2 transcription factors and associated biological pathways, indicating its potential therapeutic implications for limiting type 2 immune responses.