Oct4, Nanog and HP1 dynamics in embryonic stem cells

OSES CAMILA; COSENTINO SOLEDAD; ALVAREZ YANINA; VAZQUEZ ECHEGARAY CAMILA; GUBERMAN ALEJANDRA; LEVI VALERIA

La Plata

Congreso; XLVII SAB anual meeting; 2018

Transcription factors (TFs) dynamic interactions with DNA targets depend, among multiple parameters, on chromatin landscape. Despite TFs-chromatin interactions play a relevant role in the modulation of gene expression we still do not know how the complex temporal and spatial network of interactions defines transcriptional response in stem cells (SCs).Embryonic SCs (ESC) studies are relevant for understanding embryo development and for their potential clinical applications. These cells have two important properties: an unlimited possibility of self-renewal and pluripotency, which depends on specific TFs such as Oct4, Sox2 and Nanog that repress genes involved in differentiation and induce genes that preserve an undifferentiated state.In this work, we used fluorescence correlation spectroscopy (FCS) analysis to quantitatively explore the dynamical organization of TFs in the nucleus of ESC wild type (wt) and an ESC line knock-out for a chromatin remodeler, the histone acetyltransferase Kat6B (Kat6B-/-). Since histone acetylation usually makes a more permissive chromatin, we expected a different dynamical distribution of TFs and heterochromatin associated proteins between Kat6B -/- and wt ESCs. With the aim of comparing these dynamics in ESC wild type with Kat6B-/-, we transfected the cells with vectors encoding Oct4, Nanog or HP1 fused to enhanced green fluorescent protein (GFP), and measured fluorescence fluctuations as a function of time using confocal microscopy. We analyzed the FCS data with a model that considers fast and slow interactions with DNA targets. Results showed statistical significant differences between both cell lines for both TFs and HP1, which are consistent with a less permissive chromatin in Kat6B -/- ESC line. These results may provide important clues for understanding the transcriptional response. Furthermore, they show that FCS is a useful tool to evaluate how biological processes modulate TFs partition in the nucleus and could help us to understand early embryo development in future studies.