[en] The pituitary is a complex gland comprising different cell types each secreting specific hormones. The extensive network of signaling molecules and transcription factors required for determination and terminal differentiation of specific cell types is still not fully understood. The SRY-like HMG-box (SOX) transcription factor Sox4 plays important roles in many developmental processes and has two homologs in zebrafish, Sox4a and Sox4b. We show that the sox4b gene is expressed in the pituitary anlagen starting at 24 h after fertilization (hpf) and later in the entire head region including the pituitary. At 48 hpf, sox4b mRNA colocalizes with that for TSH (tshbeta), glycoprotein subunit alpha (gsualpha), and the Zn finger transcription factor Gata2a. Loss of Sox4b function, using morpholino knockdown or expression of a dominant-negative Sox4 mutant, leads to a drastic decrease in tshbeta and gsualpha expression and reduced levels of gh, whereas other anterior pituitary gland markers including prl, slbeta, pomc, and lim3 are not affected. Sox4b is also required for expression of gata2a in the pituitary. Knockdown of gata2a leads to decreased tshbeta and gsualpha expression at 48 hpf, similar to sox4b morphants. Injection of gata2a mRNA into sox4b morphants rescued tshbeta and gsualpha expression in thyrotrope cells. Finally, sox4b or gata2a knockdown causes a significant decrease of gonadotropin expression (lhbeta and fshbeta) at 4 d after fertilization. In summary, our results indicate that Sox4b is expressed in zebrafish during pituitary development and plays a crucial role in the differentiation of thyrotrope and gonadotrope cells through induction of gata2a expression in the developing pituitary.