DOI: 10.5061/dryad.gxd2547w7
title: Data from: α-Ketoglutarate inhibits the pluripotent-to-totipotent state transition

description: In early mouse embryogenesis, the distinct enrichment of α-ketoglutarate
 (αKG) in blastocysts and L-2-hydroxyglutarate (L-2HG) in 2-cell (2C)
 embryos serves as a key metabolic signature. While elevated L-2HG levels
 inhibit the resolution of totipotency during the transition from the 2C
 stage to the blastocyst, the role of αKG remains elusive. Mouse embryonic
 stem cells (mESCs) cultured in vitro naturally harbor a subpopulation that
 transitions dynamically into a 2C-like totipotent state, providing a
 convenient model to investigate the role of αKG in totipotency
 reprogramming. This study demonstrates that αKG significantly inhibits the
 pluripotency to totipotency transition through upregulating ten-eleven
 translocation (TET) DNA hydroxylases. We further show that reducing
 endogenous αKG levels via glutamine withdrawal or inhibiting αKG-dependent
 dioxygenases by blocking succinate dehydrogenase (SDH) markedly enhances
 the induction of 2C-like cells (2CLCs). Finally, leveraging the potent SDH
 inhibitor dimethyl malonate (DMM), we have developed a highly efficient
 protocol for 2CLC induction, producing cells that transcriptionally
 resemble mid-to-late 2C embryos. Our findings deepen the understanding of
 the metabolic regulation of totipotency and provide a previously
 undescribed approach for capturing totipotent-like stem cells in vitro.
description: # README: Supplementary Data Description ## General Information This
 README file provides details about the supplementary tables included in
 the dataset. These tables contain shRNA sequence information, primer
 details,  RNA sequencing, and scRNA sequencing data related to MERVL-GFP
 cell line under different experimental conditions. ## File Description The
 provided Excel file consists of the following supplementary tables: ###
 **Supplementary Table 1: shRNA Sequences** * This table contains the
 sequences of short hairpin RNAs (shRNAs) used in the study. * Column
 descriptions: * **Gene**: Gene targeted by the shRNA * **Target
 Sequence**: Nucleotide sequence of the shRNA ### **Supplementary Table 2:
 Primer Sequences for RT-qPCR** * This table lists the primer sequences
 used for reverse transcription quantitative PCR (RT-qPCR). * Column
 descriptions: * **Gene:** Target gene for qPCR * **Forward Primer
 (5'→3')**: Forward primer sequence * **Reverse Primer
 (5'→3')**: Reverse primer sequence ### **Supplementary Table 3:
 MERVL-GFP Bulk RNA Sequencing Data** * This table reports bulk RNA
 sequencing data for MERVL-GFP-expressing cells. * Includes differential
 expression analysis for the following comparison: * **DMM 30 mM treated
 cells vs. Control cells** * Column descriptions: * **Gene
 symbol**: Official gene symbol corresponding to the differentially
 expressed gene. * **Ensembl ID**: Unique Ensembl identifier for the gene.
 * **D30 Log2 FC**: Log2 fold change in gene expression for DMM 30 mM vs.
 SL (Control). * **D30 padj**: Adjusted p-value for the differential
 expression analysis of DMM 30 mM vs. SL (Control). * **VTD Log2 FC**: Log2
 fold change in gene expression for VTD-treated vs. SL (Control). *
 **VTD padj:** Adjusted p-value for the differential expression analysis of
 VTD-treated vs. SL (Control). ### **Supplementary Table 4: Single-Cell RNA
 Sequencing Data of MERVL-GFP cell line (VTD Treatment)** * This table
 reports single-cell RNA sequencing data for MERVL-GFP cell line. *
 Includes differential expression analysis for the following comparison: *
 **VTD treated cells vs. Control cells** * Column descriptions: * **Gene
 symbol**: Official gene symbol corresponding to the differentially
 expressed gene. * **Ensembl ID**: Unique Ensembl identifier for the gene.
 * **VTD Log2 FC**: Log2 fold change in gene expression for VTD-treated vs.
 SL (Control). * **VTD padj:** Adjusted p-value for the differential
 expression analysis of VTD-treated vs. SL (Control). ### **Supplementary
 Table 5: Single-Cell RNA Sequencing Data of Selected MERVL-GFP Cells (VTD
 Treatment)** * This table reports single-cell RNA sequencing data for
 MERVL-GFP cell line. * Includes differential expression analysis for the
 following comparison: * VTD-2CLC cells (top 200 VTD-treated cells selected
 based on totipotency gene expression) vs. Control cells (randomly selected
 200 Control cells) * Column descriptions: * **Gene symbol**: Official gene
 symbol corresponding to the differentially expressed gene. * **Ensembl
 ID**: Unique Ensembl identifier for the gene. * **VTD-2CLC Log2 FC**: Log2
 fold change in gene expression for VTD-2CLC vs. SL cells (Control). *
 **VTD-2CLC padj:** Adjusted p-value for the differential expression
 analysis of for VTD-2CLC vs. SL cells (Control). ## Notes * Data are
 provided in standard tabular format (.xlsx).