OR17-02 Combining Transcriptomics And Proteomics To Unveil Possible PROP1 Interactors During Embryonic Pituitary Development

Abstract

Abstract Disclosure: L. Iglesias Garcia: None. M.F. Mercogliano: None. C. Schuster: None. L. Cheung: None. S.A. Camper: None. M. Marti: None. M.I. Perez-Millan: None. Congenital hypopituitarism (CH) is a deficiency of one or more hormones produced by the anterior pituitary. The most frequently mutated gene in CH is PROP1, which has a very dynamic expression pattern. In Prop1 mutant mice, the pituitary is dysmorphic, with increased apoptosis and abnormal vascularization. These mutant mice are dwarfs and develop hypopituitarism. In previous work we have shown that PROP1 has a key role in stimulating progenitors to undergo an EMT-like transition before differentiation. The general objective of this work was to elucidate the PROP1 protein complexes in order to understand its role during pituitary development and to evaluate them as candidate genes that can cause congenital hypopituitarism. A GHFT-1 murine cell line expressing biotinylated Prop1 was used and co-immunoprecipitation (Co-IP) assays were performed, followed by Rapid Immunoprecipitation Mass spectrometry of Endogenous (RIME). We obtained an initial set of 786 proteins that co precipitate with PROP1. Ontology enrichment analysis shows that PROP1 immunoprecipitated with several nuclear proteins related to transcriptional regulation and splicing and also with many non nuclear proteins related to cell adhesion and migration. We used LISA and TRANSFAC to predict transcriptional regulators using as input the genes that intersected from Prop1-ChIP-Seq and RNA-seq experiments and obtained 536 genes. We compared this result with the potential PROP1 interaction proteins derived from the RIME to obtain a final list of 37 transcription factors. One possible candidate from this list is the Nuclear Factor I/B or NFIB. Mutations in this gene are related to macrocephaly with impaired intellectual development. We performed a scRNA-Seq at P4 and it showed that NFIB expresses in the same cell clusters as Prop1. Also, we have preliminary results indicating that the pituitary in NFIB KO mice at E13.5 is dysmorphic. Further experimental work is needed to validate Prop1 interaction partners and to have a better understanding of Prop1 mechanism during pituitary development. Presentation: Saturday, June 17, 2023