PROP1 regulates expression of proteins involved in EMT to lead/direct pituitary differentiation

MARTI MA; CAMPER SA; MERCOGLIANO FLORENCIA; SEILICOVICH A; DE PAUL ANA; PEREZ MILLAN MI

Mar del Plata

Congreso; Reunion SAIC 2018; 2018

Sociedad Argentina de Investigación Clínica (SAIC)

Mutations in PROP1, a key transcription factor of pituitary stem cell differentiation, are the most common known cause of hypopituitarism. We determined that PROP1 is essential for stimulating stem cells to undergo an epithelial to mesenchymal transition-like (EMT) process necessary for cell migration and differentiation. The mechanism whereby PROP1 regulates this process is not completely understood. Our goal is to further our understanding of the factors regulating embryonic pituitary progenitor cells. We genetically engineered the GHFT1 mouse pituitary cell line to express biotin-tagged PROP1 and use them to identify Prop1-mediated changes in gene expression with RNA-Seq and PROP1-interacting proteins with mass spectrometry. Gene expression profiling revealed that Prop1 upregulates genes that are involved in migration (adhesion/dispersion) and in degrading extracellular matrix proteins, like matrix metalloproteinases, MMP2, 15 and 19, and downregulates tissue inhibitor of metalloproteinases, like TIMP-3. MMPs play key roles in embryonic development and are involved in EMT in the morphogenesis of many tissues. We examined expression of MMP2 during pituitary development in control and Prop1df/df mice. At e12.5, e14.5 and e16.5, MMP2 is normally expressed in the pituitary gland in normal mice and it is almost absent in Prop1 mutant pituitaries. Furthermore, we observed an increased in cell-cell junctions by TEM in P1 pituitaries from Prop1df/df mice. Using immunoprecipitation coupled to mass spectrometry, we identified 93 proteins that specifically interact with PROP1. These proteins were tested for GO biological process enrichment, and two predominant themes were identified: Cell-Cell Adherens Junction and Cell-Cell adhesion. Several genes from these groups of proteins are candidates for involvement in migration and EMT that fails in Prop1 mutants. Together these approaches are uncovering the specific players that Prop1 regulates or interacts with to direct EMT and differentiation. This basic knowledge could implicate candidate genes to explain cases of hypopituitarism with unknown etiology and thereby, yield better diagnosis.