The Role of Gp130 Signaling in Initiation of Naive Reprogramming of hPSCs
Soutenue publiquement le 27/02/2025 à 14h00, salle de conférence du SBRI
Devant le jury composé de :
KRETZ Carole, Professeure, Université Claude Bernard Lyon 1, Lyon: Présidente
JOUNEAU Alice, Directrice de recherche, INRAE, Paris : Rapportrice
WEBER Michael, Directeur de recherche, CNRS, Strasbourg : Rapporteur
BILLON Nathalie, Chercheuse, CNRS, Nice : Examinatrice
BOURILLOT Pierre-Yves, Chercheur, CNRS, SBRI, Lyon : Directeur de thèse
SAVATIER Pierre, Directeur de recherche, INSERM, SBRI, Lyon : Co-directeur de thèse
Abstract
The conversion of human pluripotent stem cells (hPSCs) from the primed to the naïve state poses significant challenges, with only a small fraction of cells successfully completing this transition. Additionally, the molecular mechanisms driving naïve reprogramming remain poorly understood. To address this, we developed a system using human embryonic stem cells (hESCs) engineered with a genetic switch to induce the transition from the primed to the naïve state. This switch incorporates a hormone-dependent STAT3-ERT2 and a chimeric GCSF:GP130 receptor, which merges the extracellular domain of GCSF-R with the cytoplasmic domain of GP130.
We generated hESC lines expressing STAT3-ERT2 and either the wild-type (WT) GCSF:GP130 receptor or mutants lacking specific downstream signaling capabilities (JAK, STAT3, SHP2, HCK, and YES). While cells with the WT receptor or most mutants retained self-renewal when treated with 4’OHT + GCSF, those with a JAK-deficient receptor differentiated, highlighting the essential role of JAK in naïve conversion. Subsequent RNA interference experiments targeting JAK family kinases identified JAK1 as a critical regulator.
To uncover early response genes involved in reprogramming, RNA-sequencing analysis was conducted on cells treated with 4’OHT + GCSF for 24 hours, revealing differentially expressed genes (DEGs). Among these, 25 genes were expressed in the inner cell mass and epiblast of human blastocysts. Further analysis, including RNA interference screening and gene expression profiling in pre-implantation embryos and established naïve culture conditions, identified IFI16, IFITM1, IFITM2, IFITM3, SPP1, CD44, BHLHE40, FBP1, PROM1, SERPING1, and VWA1 as key regulators of the primed-to-naïve transition.
