Pin1 is a peptidyl‐prolyl cis/trans isomerase which catalyzes the isomerisation of phosphorylated Ser/Thr‐Pro peptide bonds. Pin1 knock‐out mice have a phenotype showing marked abnormalities in their reproductive development and function. However, the molecular mechanisms underlying their reproductive defects are poorly understood. We demonstrate here that Pin1 is required for the basal and GnRH‐induced gonadotropin β‐subunit gene transcription. Knocking down Pin1 expression by small interfering RNA (siRNA) reduces both gonadotropin β‐subunit gene promoter activity and endogenous mRNA levels. In addition, co‐immunoprecipitations and mammalian two hybrid assays show that Pin1 interacts with the transcription factors, SF‐1, Pitx1 and Egr‐1. Pin1 activates transcription of the gonadotropin β‐subunit genes synergistically with these transcription factors, either by modulating their stability or through increasing their protein‐protein interactions. Our results suggest that Pin1 functions as a novel player in GnRH‐induced signal pathway, involved in gonadotropin β‐subunit gene transcription by modulating the activity of various specific transcription factors.