PMON145 The Role of Pou3f4 in Pituitary Development and Disease
Authors: Masser Bailey , Camper Sally , Cheung Leonard 
Published: November 2022

Mutations in the pituitary specific transcription factor Prophet of Pit-1 (PROP1) are the most common genetic etiology of combined pituitary hormone deficiency (CPHD), which affects 1 in 8,000 newborns worldwide. CPHD is associated with short stature, attributable to growth hormone (GH) deficiency and/or thyroid stimulating hormone (TSH) deficiency. Pathogenic lesions impair pituitary development and differentiation of endocrine cells. Prop1-mutant mice show increased expression of genes including SRY-Box Transcription Factor 21 (Sox21) and Pou Domain, Class 3, Transcription Factor 4 (Pou3f4). POU3F4 is expressed in the developing inner ear and restricts the proliferation and lineage potential of neural stem cells. Genetic defects cause non-syndromic X-linked deafness (DFNX2) in humans and mice. The function of POU3F4 in pituitary development is largely unknown and the implications of its overexpression in Prop1-mutant pituitaries remain unexplored. Therefore, we aim to address the role of Pou3f4 in pituitary development and determine the contributions of Pou3f4 upregulation to pituitary disease by utilizing Pou3f4-mutant mice as well as double mutant mice lacking both Prop1 and Pou3f4.

Utilizing immunofluorescent analysis at critical timepoints during pituitary development, we have characterized the timeline for endogenous POU3F4 expression. Initial expression is detected at e11.5 in differentiating cells on the ventral side of Rathke's pouch and is restricted to the proliferating anterior lobe at e13.5, continuing postnatally, similar to previously published expression data. POU3F4 expression then decreases and becomes gradually restricted to the periluminal stem cell niche at e15.5 and p0. We further validated the upregulation of POU3F4 in histological sections of Prop1-mutant mice at e15 and p0, finding increased expression in the developing anterior lobe of mutants compared to wildtypes. Single-cell regulatory network inference and clustering (SCENIC) analyses, which utilize single-cell transcription factor co-expression with potential direct downstream targets that contain the corresponding transcription factors’ binding sequence to infer active transcription networks, predict increased activity of the Pou3f4 gene regulatory network in a subpopulation of Sox2-expressing stem cells in Prop1-mutant mice. Preliminary analyses indicate that Pou3f4 mutant have normal morphology at P1, together with grossly normal expression of Sox2 and Pou1f1. We are performing quantitative PCR to determine possible compensatory mechanisms in response to loss of Pou3f4 expression. To investigate whether any Prop1-mutant phenotypes, including dysmorphology and ectopic expression of Sox21 and Otog, are attributable to Pou3f4 upregulation, we are studying Pou3f4;Prop1 mice. We will determine the impacts of POU3F4 upregulation on cellular morphology, stem cell maintenance and endocrine cell differentiation. The mechanisms of action in the regulation of stem cell differentiation constitutes a gap in our current understanding of the pathogenesis of CPHD. These studies will provide novel insights into the mechanistic function of Pou3f4 during embryonic pituitary development and potential novel etiologies of CPHD in human patients.

Presentation: Monday, June 13, 2022 12:30 p.m. - 2:30 p.m.