Journal: EMBO Reports

Article Title: NANOG governs cell metabolism and redox homeostasis in human naïve embryonic stem cells

doi: 10.1038/s44319-025-00629-9

Figure Lengend Snippet: ( A ) A Venn diagram illustrating the overlap between NANOG- occupied genes and DEGs induced by NANOG deletion for three days in naïve hESCs. ( B ) The list of multiple metabolic pathways enriched by the GO analysis of 362 NANOG- occupied and NANOG depletion- downregulated DEGs in naïve hESCs. ( C ) Genome browser snapshots showing levels of NANOG occupancy near DLAT, CPT1A, ACADS, ALDH2, MPC2 , and ACSS2 in primed and naïve SHhES8 hESCs, as well as histone modifications and chromatin openness in naïve NANOG iKO SHhES8 hESCs, either untreated or treated with DOX for two or three days. ( D ) The results for ChIP-qPCR validation of NANOG occupancy on the promoter region of DLAT and MPC2 . Two pairs of primers were used for each gene. Data are presented as mean ± SEM ( n = 3). ( E ) RT-qPCR analysis results for relative mRNA levels of six NANOG target genes in naïve NANOG iKO hESCs, either untreated or treated with DOX for two or three days. Data are presented as mean ± SEM ( n = 3). ( F ) The representative western blot analysis result for steady-state levels of proteins examined in naïve NT or NANOG iKO hESCs, either untreated or treated with DOX for two or three days. Alpha-TUBULIN served as a loading control (left). The quantitative analysis results of relative protein levels from four independent experiments are shown on the right side. Data are presented as mean ± SEM ( n = 4). ( G ) The representative western blot analysis result for levels of proteins examined in naïve i NANOG-mut _ NANOG iKO SHhES8 hESCs, either untreated or treated with DOX for three days. Alpha-TUBULIN served as a loading control. ( H ) RT-qPCR analysis results for relative mRNA levels of indicated genes in naïve i NANOG-mut _ NANOG iKO SHhES8 hESCs, either untreated or treated with DOX for three days. Data are presented as mean ± SEM ( n = 3). The unpaired two-tailed Student’s t -test was used for the statistical analysis in ( D – F , H ). .

Article Snippet: MPC2 , Proteintech , Cat# 20049-1-AP.

Techniques: ChIP-qPCR, Biomarker Discovery, Quantitative RT-PCR, Western Blot, Control, Two Tailed Test

Journal: EMBO Reports

Article Title: NANOG governs cell metabolism and redox homeostasis in human naïve embryonic stem cells

doi: 10.1038/s44319-025-00629-9

Figure Lengend Snippet: ( A ) The GO analysis of 473 NANOG- occupied and NANOG depletion- upregulated (top) as well as 362 NANOG-occupied and NANOG depletion- downregulated (bottom) DEGs in naïve hESCs. The top 20 enriched terms are presented. ( B ) The heatmap depicting normalized mRNA levels of metabolism- related genes in enriched pathways shown in Fig. , measured by our RNA-seq assays. The color represents Z-scores. Three biological replicates were used for each condition ( n = 3). ( C ) A schematic diagram of metabolic pathways of acetyl-CoA synthesis and putative NANOG targets associated with the pathway. ( D ) The representative western blot analysis result for protein levels of NANOG, histone acetylation modifications, and proteins related to acetyl-CoA synthesis in naïve H9 NANOG iKO hESCs cultured under the 5iLA condition, either untreated or treated with DOX for two or three days. Alpha-TUBULIN and histone 3 served as loading controls (left). The quantitative analysis results of relative protein levels are shown on the right side. Data are presented as mean ± SEM ( n = 3). DLAT, Row 1, p = 2.93 × 10 −7 ; NANOG, Row 1, p = 2.60 × 10 −5 . ( E ) The representative western blot analysis result for levels of proteins indicated in naïve NT and NANOG iKO SHhES8 hESCs cultured under the PXGL condition, either untreated or treated with DOX for two or three days. Alpha-TUBULIN and histone 3 served as loading controls (left). The quantitative analysis results of relative protein levels are shown on the right side. Data are presented as mean ± SEM ( n = 3). CPT1A, p = 1.08 × 10 −5 ; NANOG, Row 1, p = 1.06 × 10 −6 . ( F ) RT-qPCR analysis results for relative mRNA levels of the indicated genes in naïve NANOG iKD SHhES8 hESCs, either untreated or treated with DOX for 3 days. Data are presented as mean ± SEM ( n = 3). NANOG , p = 9.39 × 10 −5 ; ALDH2 , p = 9.35 × 10 −6 ; MPC2 , p = 3.62 × 10 −5 . ( G ) The representative western blot analysis result for levels of indicated proteins in naïve NANOG iKD SHhES8 hESCs, either untreated or treated with DOX for 3 days. dCas9 was tagged by HA. Alpha-TUBULIN and histone 3 served as loading controls. The unpaired two-tailed student’s t -test was used for the statistical analysis in ( D – F ).

Article Snippet: MPC2 , Proteintech , Cat# 20049-1-AP.

Techniques: RNA Sequencing, Western Blot, Cell Culture, Quantitative RT-PCR, Two Tailed Test