Do H3K4 methyltransferases MLL4 and MLL3 instruct the mammalian body plan?

主要的表观遗传调节因子MLL4在许多癌症中发生显著突变，是六种哺乳动物组蛋白3赖氨酸4 （H3K4）甲基转移酶之一。MLL4的杂合突变引发歌舞伎综合征，包括智力迟钝、面部和其他形态改变以及代谢紊乱。尽管具有重要的医学意义，但MLL4在小鼠发育中的作用仅得到部分研究。我们已经系统地记录了所有六种H3K4甲基转移酶在胚胎中的作用，并报道了由于在早期发育过程中建立前内脏内胚层（AVE）以及前后轴（a -p）的缺陷，Mll4缺失胚胎会发生有缺陷的原肠胚形成。为了进一步研究开发，我们生成了一个新的loxP条件Mll4等位基因。使用Sox2Cre，我们确定了MLL4在外胚层中的作用，并在不同时间使用R26CreERT2与他莫昔芬诱导，发现了MLL4在E13.5完成a - p轴延伸之前的作用。我们认为，MLL4在神经中胚层祖细胞（NMP）的躯干伸长、分割和尾部规范中是持续需要的，可能是通过调控Hox簇共线来发挥作用的。条件诱变还显示，直到出生时才需要的MLL3可以在A-P轴伸长期间替代MLL4。通过在子宫和胚胎干细胞系（ESCs）中进行条件诱变，我们的目标是剖析MLL4（和MLL3）对哺乳动物身体计划的建立的贡献。我们将研究在缺乏MLL4 （+/-MLL3）的情况下，是否可以通过NMPs和原肠样蛋白在体外重现胚胎发育，重点研究Hox簇表达的共线性诱导。这些分析包括使用将MLL4 BAC转座子序列删除到条件MLL4 ESCs中，对MLL4进行结构/功能解剖。利用原位杂交、成像技术和部分由我们实验室开发的条件诱变方法，以及表达谱、深度测序和体外胚胎发生，我们的目标是确定哺乳动物身体计划规范中MLL4（和MLL3）之间的关系。

The major epigenetic regulator, MLL4, is prominently mutated in many cancers and is one of six mammalian histone 3 lysine 4 (H3K4) methyltransferases. Heterozygous mutations of MLL4 provoke Kabuki syndrome, which involves mental retardation, facial and other morphological changes, and metabolic disorders. Despite its medical importance, the role of MLL4 in mouse development has only been partially examined. We have been systematically documenting the embryonic roles of all six H3K4 methyltransferases and report that Mll4 null embryos undergo defective gastrulation owing to a defect in establishing the anterior visceral endoderm (AVE) and consequently, the anterior-posterior (A-P) axis during early development. To probe later into development, we generated a new loxP conditional Mll4 allele. Using Sox2Cre, we identified a role for MLL4 in the epiblast and using R26CreERT2 with tamoxifen-induction at different times, found evidence for MLL4 contribution until the completion of A-P axis elongation at E13.5. We propose that MLL4 is continuously required for neuromesodermal progenitor (NMP) specification of trunk elongation, segmentation and the tail, potentially exerted through regulation of Hox cluster colinearity. Conditional mutagenesis also revealed that MLL3, which is not required until birth, can substitute for MLL4 during the elongation of the A-P axis. Using conditional mutagenesis in utero and in embryonic stem cell lines (ESCs) established from our conditional knockout mouse lines, we aim to dissect the contributions of MLL4 (and MLL3) to the establishment of the mammalian body plan. We will investigate whether embryonic development can be recapitulated in vitro through NMPs and gastruloids in the absence of MLL4 (+/-MLL3) with a focus on colinear induction of Hox cluster expression. These analyses include a structure/function dissection of MLL4 using a deletion series of Mll4 BAC transposons introduced into the conditional Mll4 ESCs. Using in situ hybridization, imaging techniques, and conditional mutagenesis methods developed in part by our lab, together with expression profiling, deep sequencing and in vitro embryogenesis, we aim to identify the relationship between MLL4 (and MLL3) in specification of the mammalian body plan.