SALL4 controls cell fate in response to DNA base composition. Pantier et al.

Mammalian genomes contain long domains with distinct average compositions of A/T versus G/C base pairs. In a screen for proteins that might interpret base composition by binding to AT-rich motifs, we identified the stem cell factor SALL4 which contains multiple zinc-fingers. Mutation of the domain responsible for AT binding drastically reduced SALL4 genome occupancy and prematurely up-regulated genes in proportion to their AT content. Inactivation of this single AT-binding zinc-finger cluster mimicked defects seen in Sall4-null cells, including precocious differentiation of embryonic stem cells and embryonic lethality in mice. In contrast, deletion of two other zinc-finger clusters was phenotypically neutral. Our data indicate that loss of pluripotency is triggered by down-regulation of SALL4, leading to de-repression of a set of AT-rich genes that promotes neuronal differentiation. We conclude that base composition is not merely a passive by-product of genome evolution, but constitutes a signal that aids control of cell fate. Python scripts and source code used for bioinformatic analyses, raw Western blot and microscopy images, as well as other types of unprocessed and processed data used to generate the figures are available here. "Figure 1.zip", "Figure 4.zip", "Figure 5.zip", "Figure 6.zip", "Figure S1.zip", "Figure S2.zip", "Figure S3.zip", "Figure S4.zip", "Figure S5.zip" and "Figure S6.zip" contain the raw western blot, microscopy images, processed and unprocessed data associated with figure panels in respective figures. "bioinformatics.zip" contains the respective ipynb (jupyter notebook) files for Figure 1, 2, 3, 6 and 7. Each notebook contains the source code to generate main+supplemental figure panels and the sub-folder "data" contain the processed data associated with the respective figure. "selex_scripts.zip" contains the Python scripts and all the necessary information to reproduce the analysis of HT-SELEX data. Raw and processed high-throughput sequencing data is deposited on Array Express, as described below. E-MTAB-7343 RNA-seq of WT, S4KO, ZFC4mut and ZFC4Δ ESCs E-MTAB-7655 RNA-seq of WT, S4KO, ZFC4mut and ZFC1-2Δ ESCs E-MTAB-9197 SALL4 ChIP-seq in WT, S4KO, ZFC4mut and ZFC1-2Δ ESCs E-MTAB-9198 Timecourse RNA-seq during differentiation (day 0, 2 and 5) of WT, S4KO, ZFC4mut and ZFC1-2Δ ESCs E-MTAB-9202 RNA-seq of S4KO cells carrying Sall4 cDNA or EGFP cDNA under a doxycycline inducible promoter E-MTAB-9236 HT-SELEX of recombinant C2H2 zinc-finger domains of SALL4 E-MTAB-9245 ATAC-seq in WT ESCs

哺乳动物基因组包含具有不同A/T与G/C碱基平均组成的长结构域。在通过结合AT富集基序来解读碱基组成的蛋白质筛选中，我们鉴定出了干细胞因子SALL4，其包含多个锌指结构域。负责AT结合的结构域发生突变后，SALL4在基因组中的占据率大幅降低，且会按照基因的AT含量比例提前上调基因表达。灭活该单个AT结合锌指簇的表型与Sall4敲除细胞中的缺陷一致，包括胚胎干细胞的过早分化以及小鼠的胚胎致死。相比之下，缺失另外两个锌指簇则表型中性。我们的数据表明，多能性的丧失由SALL4的下调触发，进而导致一组促进神经元分化的AT富集基因去抑制。我们得出结论：碱基组成不仅仅是基因组进化的被动副产物，而是构成了有助于调控细胞命运的信号。本研究用于生物信息学分析的Python脚本与源代码、原始蛋白质印迹（Western blot）和显微镜图像，以及用于生成图表的各类未处理与已处理数据均可在此获取。"Figure 1.zip"、"Figure 4.zip"、"Figure 5.zip"、"Figure 6.zip"、"Figure S1.zip"、"Figure S2.zip"、"Figure S3.zip"、"Figure S4.zip"、"Figure S5.zip"及"Figure S6.zip"包含对应图表面板相关的原始蛋白质印迹图像、显微镜图像、已处理与未处理数据。"bioinformatics.zip"包含用于生成图1、2、3、6及7的相关Jupyter Notebook（ipynb）文件，每个Notebook均包含生成主图与补充图面板的源代码，其子文件夹"data"中存放对应图表的已处理数据。"selex_scripts.zip"包含用于复现高通量指数富集配体系统进化（HT-SELEX）数据分析的Python脚本及所有必要信息。原始与已处理的高通量测序数据已提交至Array Express，详情如下：E-MTAB-7343：野生型（WT）、S4KO、ZFC4mut及ZFC4Δ胚胎干细胞（ESC）的RNA测序（RNA-seq）；E-MTAB-7655：野生型（WT）、S4KO、ZFC4mut及ZFC1-2Δ胚胎干细胞（ESC）的RNA测序（RNA-seq）；E-MTAB-9197：野生型（WT）、S4KO、ZFC4mut及ZFC1-2Δ胚胎干细胞（ESC）中的SALL4染色质免疫沉淀测序（ChIP-seq）；E-MTAB-9198：野生型（WT）、S4KO、ZFC4mut及ZFC1-2Δ胚胎干细胞（ESC）在分化过程（第0、2、5天）的时间序列RNA测序（RNA-seq）；E-MTAB-9202：携带由强力霉素（doxycycline）诱导型启动子调控的Sall4互补DNA（cDNA）或增强绿色荧光蛋白（EGFP）cDNA的S4KO细胞的RNA测序（RNA-seq）；E-MTAB-9236：重组SALL4的C2H2锌指结构域的HT-SELEX测序；E-MTAB-9245：野生型（WT）胚胎干细胞（ESC）的转座酶可及性测序（ATAC-seq）