Fig. 1

SMYD5 regulates ES cell self-renewal. a Bright-field microscopy of ES cells infected with shLuc or shSmyd5 lentiviral particles and wild-type (WT) SMYD5 or an enzymatically mutant (mut) version of SMYD5 (H315L and C317A) lentiviral particles and stably selected with puromycin and G418. b ES cell colonies were scored by morphology. The percentage of colonies with an ES-like morphology (compact and round vs. flattened) are represented as mean ± SEM. P values were calculated using a t test. c Alkaline phosphatase (AP) staining of ES cells. d ES cells were scored by AP staining. The percentage of AP positive colonies is represented as mean ± SEM. p values were calculated using a t test. e Quantitative RT-PCR (Q-RT-PCR) expression of SMYD5 using primers for three different regions of the SMYD5 coding region. f Scatter plot of RNA sequencing (RNA-Seq) gene expression analysis between shLuc and shSmyd5 ES cells. Log2 adjusted differentially expressed genes are plotted. Genes whose expression is greater than twofold (shLuc vs shSmyd5) and with an RPKM > 1 (reads per kilo bases of exon model per million reads) and FDR < 0.001 are shown in black. g UCSC genome browser view of differential expression of self-renewal genes in shSmyd5 and shLuc control ES cells. h Q-RT-PCR analysis of expression of Smyd5 and self-renewal genes in shLuc and shSmyd5 ES cells. i Gene set enrichment analysis (GSEA) [22] of differentially expressed genes in Smyd5 knockdown ES cells relative to undifferentiated and differentiated embryoid bodies (EBs). j Gene ontology (GO) functional annotation of differentially expressed genes analyzed using DAVID [23]. k Mouse gene atlas expression analysis evaluated using Network2Canvas [66] demonstrates that lineage and ES cell genes are misexpressed in shSmyd5 ES cells. Each node (square) represents a gene list (shLuc vs shSmyd5 DE genes) associated with a gene-set library (mouse gene atlas). The brightness (white) of each node is determined by its p value