RNA-seq analysis dataset: FACS-sorted Ptf1a-lineage cells from E14.5 hypothalamus of Ptf1a hetero and homozygous mice

Because Ptf1a is a transcription factor, this protein should regulate transcription of various genes in Ptf1a-expressing cells of the hypothalamus. To detect those downstream genes, we purified cells in the Ptf1a-lineage from the hypothalamus at E14.5 by FACS sorting, subsequently performed RNA-seq analysis and obtained 706 differentially expressed genes. 386 genes had lower and 320 had higher expression in Ptf1a-lineage cells from homozygous Ptf1a deficient mice. The down-regulated genes included Prdm13, Corl2 and Lhx5 that were known downstream targets of Ptf1a, in addition, Sox3 and Nkx2-1, which are known to be required for hypothalamic development. Interestingly, among those affected genes, we detected several genes encoding transmembrane and/or secreted proteins, such as Jag2 (ligand for Notch receptors), Sema6b, Sema4g (ligands for Plexin family proteins). These genes may have a non-cell autonomous role in the sexual differentiation of hypothalamic cells adjacent to Ptf1a-lineage cells. Kiss1 expression was not altered, as was expected by the finding that most Kiss1-expressing cells were not in the Ptf1a lineage. Hypothalamic Ptf1a-lineage cell mRNA profiles of E14.5 Ptf1a-cre heterozygous and homozygous mice with Ai9 reporter were generated by FACS-sorting and RNA-sequencing, using BD Bioscience FACSAria II SORP flow cytometer and Illumina NextSeq500.

鉴于Ptf1a属于转录因子（transcription factor），该蛋白应可在下丘脑表达Ptf1a的细胞中调控多种基因的转录过程。为鉴定此类下游靶基因，我们于胚胎发育第14.5天（E14.5）通过荧光激活细胞分选（fluorescence-activated cell sorting, FACS）从下丘脑中分离纯化得到Ptf1a谱系细胞，随后开展RNA测序（RNA sequencing, RNA-seq）分析，最终获得706个差异表达基因（differentially expressed genes）。在纯合子Ptf1a缺陷小鼠的Ptf1a谱系细胞中，386个基因的表达水平下调，320个基因的表达水平上调。下调基因中包含已被报道的Ptf1a下游靶标Prdm13、Corl2与Lhx5，此外还有Sox3与Nkx2-1——这两个基因已被证实对下丘脑发育不可或缺。值得注意的是，在受调控的基因中，我们鉴定到多个编码跨膜蛋白和/或分泌蛋白的基因，例如Notch受体（Notch receptors）的配体Jag2，以及Plexin家族蛋白（Plexin family proteins）的配体Sema6b与Sema4g。此类基因可能在Ptf1a谱系细胞邻近的下丘脑细胞的性别分化过程中发挥非细胞自主调控功能。Kiss1基因的表达水平未发生改变，这与预期相符——因为绝大多数表达Kiss1的细胞并不属于Ptf1a谱系。本数据集通过荧光激活细胞分选与RNA测序技术，结合BD生物科学FACSAria II SORP流式细胞仪与Illumina NextSeq500测序平台，获取了携带Ai9报告基因（Ai9 reporter）的E14.5天Ptf1a-cre杂合子与纯合子小鼠的下丘脑Ptf1a谱系细胞的mRNA表达谱。