Section of Hematology/Oncology, Department of Medicine, The University of Chicago, Chicago, IL 60637, USA.
Center for Research Informatics, The University of Chicago, Chicago, IL 60637, USA.
Cell Rep. 2014 Jan 16;6(1):231-244. doi: 10.1016/j.celrep.2013.11.044. Epub 2013 Dec 27.
Hematopoietic stem cell differentiation involves the silencing of self-renewal genes and induction of a specific transcriptional program. Identification of multiple covalent cytosine modifications raises the question of how these derivatized bases influence stem cell commitment. Using a replicative primary human hematopoietic stem/progenitor cell differentiation system, we demonstrate dynamic changes of 5-hydroxymethylcytosine (5-hmC) during stem cell commitment and differentiation to the erythroid lineage. Genomic loci that maintain or gain 5-hmC density throughout erythroid differentiation contain binding sites for erythroid transcription factors and several factors not previously recognized as erythroid-specific factors. The functional importance of 5-hmC was demonstrated by impaired erythroid differentiation, with augmentation of myeloid potential, and disrupted 5-hmC patterning in leukemia patient-derived CD34+ stem/early progenitor cells with TET methylcytosine dioxygenase 2 (TET2) mutations. Thus, chemical conjugation and affinity purification of 5-hmC-enriched sequences followed by sequencing serve as resources for deciphering functional implications for gene expression during stem cell commitment and differentiation along a particular lineage.
造血干细胞分化涉及自我更新基因的沉默和特定转录程序的诱导。多个共价胞嘧啶修饰的鉴定提出了这样一个问题，即这些衍生碱基如何影响干细胞的定向分化。我们利用可复制的原发性人类造血干/祖细胞分化系统，在造血干细胞向红细胞系定向分化的过程中，证明了 5-羟甲基胞嘧啶（5-hmC）的动态变化。在红细胞分化过程中保持或增加 5-hmC 密度的基因组位点包含红细胞转录因子的结合位点，以及几个以前未被认为是红细胞特异性因子的因子。5-hmC 的功能重要性通过以下实验得到证明：在白血病患者来源的 CD34+干细胞/早期祖细胞中，TET 甲基胞嘧啶双加氧酶 2（TET2）突变导致 5-hmC 模式紊乱，从而导致红细胞分化受损，髓系潜能增强。因此，5-hmC 富集序列的化学偶联和亲和纯化，然后进行测序，可作为解析干细胞定向分化过程中基因表达功能意义的资源。
