The roles of Zbtb11 in embryonic stem cell proliferation and survival

Zbtb11 is a transcription factor conserved in vertebrates. Mutations in its human gene cause a form of inherited intellectual disablity, while in zebrafish Zbtb11 mutations lead to impaired hematopoesis and neruonal development. To determine the genes regulated by Zbtb11 we generated an inducible KO ES cell line (Zbtb11 lox/lox, Rosa26 ERt2-Cre) that allows the rapid depletion of Zbtb11 upon treatment with 4-hydroxytamoxyfen (4OHT). We subsequently performed RNA-seq in Zbtb11 KO and control cells to determine differentially expressed genes. Upon Zbtb11 deletion 310 genes changed expression. Integration of the RNA-seq data with Zbtb11 ChIP-seq data (GEO series GSE125047) allowed us to distinguish between genes directly regulated by Zbtb11 and indirect changes. 204 differentially expressed genes were bound by Zbtb11, while 106 did not have Zbtb11 binding sites. Directly regulated genes were significantly enriched in genes with mitochondrial functions, while indirectly controlled genes were enriched for p53 targets. In agreement with these findings Zbtb11 KO led to impaired respiration, proliferation arrest and cell death. Overall design: RNA-seq to determine differentially expressed genes upon induction of Zbtb11 KO. Directional RNA-seq was performed to compare transcripts in Zbtb11 lox/lox, Rosa26 ERt2-Cre mouse ES cells treated with 4-hydroxytamoxifen (4OHT) (Zbtb11 KO) or ethanol (EtOH) (control) for 72 hours. To control for non-specific effects of 4OHT on gene expression, we also compared EtOH-treated Zbtb11 lox/lox, Rosa26 ERt2-Cre cells with 4OHT-treated Zbtb11 wt/wt, Rosa26 wt cells.