PcG家族CBX2基因的发现及其在哺乳动物性发育中的作用

Polycomb group complex (PcG)作为发挥转录抑制作用的重要表观遗传调控复合物参与发育、衰老以及肿瘤发生等重要生理过程。Pc G家族成员,包括PRC1 (Polycomb repressive complex 1)与PRC2 (Polycomb repressive complex 2)两种复合物,各组分间功能既协同,又不失特性,色素框同源蛋白(Chromobox homolog CBX)是PRC1的主要核心蛋白,其家族的成员之一CBX2近年来成为研究的热点。本研究综述了CBX2基因的发现、基因和蛋白质的结构组成及其在人和鼠不同组织的表达情况,阐述了CBX2在哺乳动物性发育中的作用及研究进展,发现CBX2除参与PcG复合体发挥转录抑制作用外,还具有基因转录激活作用,它作为哺乳动物性别决定级联中SRY (Sex region of Y chromosome)的上游基因可直接激活靶基因SF1/NR5A1的表达,如果CBX2基因突变或缺失将使雄性小鼠发生性反转,在人类中造成46,XY性发育障碍(disorders of sex development, DSD),为进一步研究CBX2基因在动物性发育调控中的作用提供依据。     

多梳蛋白CBX家族在哺乳动物中结构与功能多样性的研究进展

多梳蛋白家族(PcG)是一类重要的表观遗传调控因子,主要参与维持特定基因的转录沉默,与多种干细胞的干性维系、细胞分化、细胞周期的调控、细胞衰老、X染色体失活、基因印记、癌症的发生发展等一系列细胞生理及病理活动密切相关.PcG蛋白主要形成PRC1和PRC2两类多梳蛋白抑制复合体,它们彼此相互协同调控基因的表达.其中Chromobox同源蛋白即CBX蛋白作为PRC1复合体的核心组分协助招募并稳定PRC1到染色质的特定区域.在哺乳动物中,CBX蛋白家族包含5个成员,它们在结构和功能方面既有共性又有特性,本文主要就多梳蛋白CBX家族在哺乳动物中结构与功能多样性的研究进展作一综述.     

PcG蛋白转录抑制复合物组份的功能及构成的时空特征

PcG蛋白主要以PRC1和PRC2两组复合物的形式存在,通过参与核小体组蛋白翻译后修饰等机制,发挥调控靶基因转录的功能. PRC1复合体中的RING1A/B具有使组蛋白H2AK119泛素化的活性;PRC2中的EZH2具有使组蛋白H3K27三甲基化的活性,形成PRC1锚定到核小体上的位点. PcG蛋白的表达特征具有发育阶段和细胞类型时空特异性. 长链非编码RNA等反式作用因子能募集PcG蛋白结合于靶基因,发挥靶向作用. 本文就PcG蛋白功能、构成的时空特异性、募集机制及其与疾病发生的关系研究进展做一综述.     

PcG蛋白转录抑制复合物组分的功能及构成的时空特征

PcG蛋白主要以PRC1和PRC2两组复合物的形式存在,通过参与核小体组蛋白翻译后修饰等机制,发挥调控靶基因转录的功能.PRC1复合体中的RING1A/B具有使组蛋白H2AK119泛素化的活性;PRC2中的EZH2具有使组蛋白H3K27三甲基化的活性,形成PRC1锚定到核小体上的位点.PcG蛋白的表达特征具有发育阶段和细胞类型时空特异性.长链非编码RNA等反式作用因子能募集PcG蛋白结合于靶基因,发挥靶向作用.本文就PcG蛋白功能、构成的时空特异性、募集机制及其与疾病发生的关系研究进展做一综述.     

PcG蛋白在干细胞调控中的作用

PcG (polycomb group)蛋白作为一种表观遗传修饰系统,在动物和植物中具有 保守性.从功能上讲,PcG蛋白可以分为PRC1（polycomb repressive complex 1）和 PRC2（polycomb repressive complex 2）两个核心蛋白复合体. PRC2含有组蛋白甲 基化酶的活性,而PRC1在泛素连接酶E3介导的组蛋白泛素化中发挥作用,二者通过对 组蛋白的修饰控制靶基因转录. 近来研究表明,PcG蛋白对干细胞数量维持和命运转变 有重要的调控作用,其成员表达失调或缺失导致许多恶性肿瘤的发生或导致植物细胞 丧失分化能力、形成愈伤组织. 本文简要综述了PcG蛋白的组成及其在干细胞调控中 的作用.     

去泛素化酶USP29调控CBX6蛋白质稳定性

多梳家族(polycomb group,PcG)是一类控制细胞命运和胚胎发育的转录抑制因子,主要以转录抑制复合物(polycomb repressive complex,PRC)的形式发挥功能.染色体盒蛋白质同源物6(chromobox protein homolog 6,CBX6)是PRC1的核心蛋白质亚基之一,在基...     

Polycomb group蛋白复合体

Polycomb group (PcG) 蛋白是一组通过染色质修饰调控靶基因的转录抑制子, 从生化和功能上它可以分成两个主要的核心蛋白复合体PRC1(Polycomb repressive complex 1)和PRC2(Polycomb repressive complex 2)。研究发现PcG蛋白不仅控制个体正确的发育模式, 而且与细胞的增殖、分化和肿瘤发生有关。文章就PcG蛋白的组成、作用机制及功能进行综述, 并对PcG未来的研究方向进行展望。     

拟南芥印迹基因MEA

MEDEA(MEA)是第1个在拟南芥胚乳中发现的印迹基因.MEA基因的印迹受甲基转移酶MET1和DNA糖基酶DME之间的拮抗作用调控.MEA基因编码polycomb group(PcG)蛋白,并与FIE、MSI1等其他PcG蛋白形成复合物,维持与细胞增殖相关基因的表达抑制状态.mea突变会启动未受精状态下中央细胞核的复制,而使种子发生败育.目前已发现Ⅰ型MADSbox基因PHERES1是受MEA直接作用的.研究MEA基因对种子发育的调控,不仅有助于阐明原胚及胚乳启动的机制,在无融合生殖研究中也有意义.PcG蛋白在动植物中的强保守性说明,PcG基因印迹在个体发育过程中有调控作用.     

Nestin基因在髓系白血病的表达及意义

Nestin蛋白是一种神经干细胞标志物,参与组织修复,并且在一些肿瘤细胞中表达。最近研究报道称nestin可能参与肿瘤细胞恶性增殖及细胞侵袭。在急性淋巴细胞白血病(acute lymphoid leukemia,ALL)小鼠模型中,白血病增殖细胞(leukemia-propagating cells,LPCs)可形成一个抗药性niche,其中nestin阳性细胞参与该niche形成。为探索nestin基因在白血病患者的临床表达情况,通过RT-PCR和Western-blot实验方法 ,检测了50位髓系白血病患者的nestin基因的临床表达情况。结果发现,在50位髓系白血病患者(38位AML,12位CML)中,42位患者表达nestin基因mRNA,6位患者表达nestin蛋白,并且nestin基因表达水平和患者血红蛋白、血小板、细胞因子和T淋巴细胞数并不存在相关性,但是高白细胞白血病患者表现出nestin基因mRNA水平高表达。虽然nestin基因和患者完全缓解率(complete remission,CR)并不存在联系,但是nestin基因在髓系白血病异常表达,说明其可能作为一种诊断AML或CML的生物标记物。     

HOXA10直接调控人子宫内膜间质细胞中p/CAF启动子的活性

目的：筛选与鉴定转录因子同源框蛋白（HOX）A10下游靶基因。方法：用Ad-HOXA10重组腺病毒感染人子宫内膜间质细胞,通过染色体免疫共沉淀（ChIP）方法,筛选HOXA10的下游靶基因;采用萤光素酶报告基因实验结合腺病毒介导的HOXA10过表达和小干扰RNA介导的基因沉默实验,分析HOXA10对下游靶基因的转录调控作用。结果：ChIP-PCR筛选并鉴定p/CAF（p300/CBP相关因子）为HOXA10直接作用的靶基因;过表达HOXA10抑制p/CAF启动子活性达60%;基因沉默内源性HOXA10的表达可以激活p/CAF启动子活性超过2倍以上。结论：p/CAF是HOXA10新的靶基因,HOXA10可能通过调节p/CAF的表达来调控子宫内膜的发育。     

Negative Regulation of Erythroid Differentiation via the CBX8-TRIM28 Axis

Although the mechanism of chronic myeloid leukemia (CML) initiation through BCR/ABL oncogene has been well characterized, CML cell differentiation into erythroid lineage cells remains poorly understood. Using CRISPR-Cas9 screening, we identify Chromobox 8 (CBX8) as a negative regulator of K562 cell differentiation into erythrocytes. CBX8 is degraded via proteasomal pathway during K562 cell differentiation, which activates the expression of erythroid differentiation-related genes that are repressed by CBX8 in the complex of PRC1. During the differentiation process, the serine/threonine-protein kinase PIM1 phosphorylates serine 196 on CBX8, which contributes to CBX8 reduction. When CD235A expression levels are analyzed, the result reveals that the knockdown of PIM1 inhibits K562 cell differentiation. We also identify TRIM28 as another interaction partner of CBX8 by proteomic analysis. Intriguingly, TRIM28 maintains protein stability of CBX8 and TRIM28 loss significantly induces proteasomal degradation of CBX8, resulting in an acceleration of erythroid differentiation. Here, we demonstrate the involvement of the CBX8-TRIM28 axis during CML cell differentiation, suggesting that CBX8 and TRIM28 are promising novel targets for CML research.     

CBX8 interacts with chromatin PTEN and is involved in regulating mitotic progression

ObjectivesBesides its role in regulating phosphatidylinositol‐3 kinase (PI3K) signalling in the cytosol, PTEN also has a nuclear function. In this study, we attempted to understand the mechanism of chromatin PTEN in suppressing chromosomal instability during cell division.Materials and methodsImmunocoprecipitation, ectopic expression, and deletional analyses were used to identify the physical interaction between Chromobox Homolog protein 8 (CBX8) and PTEN, as well as the functional domain(s) of PTEN mediating the interaction. Cell synchronization followed by immunoblotting was employed to study cell cycle regulation of CBX8 and the functional interaction between chromatin PTEN and CBX8. Small interfering RNAs (siRNAs) were used to study the role of PTEN and CBX8 in modulating histone epigenetic markers during the cell cycle.ResultsPolycomb group (PcG) proteins including CBXs function to repress gene expression in a wide range of organisms including mammals. We recently showed that PTEN interacted with CBX8, a component of Polycomb Repressing Complex 1 (PRC1), and that CBX8 co‐localized with PTEN in the nucleus. CBX8 levels were high, coinciding with its phosphorylation in mitosis. Phosphorylation of CBX8 was associated with monoubiquitinated PTEN and phosphorylated‐BubR1 on chromatin. Moreover, CBX8 played an important role in cell proliferation and mitotic progression. Significantly, downregulation of either PTEN or CBX8 induced H3K27Me3 epigenetic marker in mitotic cells.ConclusionCBX8 is a new component that physically interacts with chromatin PTEN, playing an important role in regulating mitotic progression.     

CBX7 and miR‐9 are part of an autoregulatory loop controlling p16INK4a

Polycomb repressive complexes (PRC1 and PRC2) are epigenetic regulators that act in coordination to influence multiple cellular processes including pluripotency, differentiation, cancer and senescence. The role of PRCs in senescence can be mostly explained by their ability to repress the INK4/ARF locus. CBX7 is one of five mammalian orthologues of Drosophila Polycomb that forms part of PRC1. Despite the relevance of CBX7 for regulating senescence and pluripotency, we have a limited understanding of how the expression of CBX7 is regulated. Here we report that the miR‐9 family of microRNAs (miRNAS) downregulates the expression of CBX7. In turn, CBX7 represses miR‐9‐1 and miR‐9‐2 as part of a regulatory negative feedback loop. The miR‐9/CBX7 feedback loop is a regulatory module contributing to induction of the cyclin‐dependent kinase inhibitor (CDKI) p16^INK4a during senescence. The ability of the miR‐9 family to regulate senescence could have implications for understanding the role of miR‐9 in cancer and aging.     

Spred2 Modulates the Erythroid Differentiation Induced by Imatinib in Chronic Myeloid Leukemia Cells

Differentiation induction is currently considered as an alternative strategy for treating chronic myelogenous leukemia (CML). Our previous work has demonstrated that Sprouty-related EVH1 domainprotein2 (Spred2) was involved in imatinib mediated cytotoxicity in CML cells. However, its roles in growth and lineage differentiation of CML cells remain unknown. In this study, we found that CML CD34⁺ cells expressed lower level of Spred2 compared with normal hematopoietic progenitor cells, and adenovirus mediated restoration of Spred2 promoted the erythroid differentiation of CML cells. Imatinib could induce Spred2 expression and enhance erythroid differentiation in K562 cells. However, the imatinib induced erythroid differentiation could be blocked by Spred2 silence using lentiviral vector PLKO.1-shSpred2. Spred2 interference activated phosphorylated-ERK (p-ERK) and inhibited erythroid differentiation, while ERK inhibitor, PD98059, could restore the erythroid differentiation, suggesting Spred2 regulated the erythroid differentiation partly through ERK signaling. Furthermore, Spred2 interference partly restored p-ERK level leading to inhibition of erythroid differentiation in imatinib treated K562 cells. In conclusion, Spred2 was involved in erythroid differentiation of CML cells and participated in imatinib induced erythroid differentiation partly through ERK signaling.     

Inhibition of PCGF2 enhances granulocytic differentiation of acute promyelocytic leukemia cell line HL-60 via induction of HOXA7

This study tested the hypothesis that Polycomb Repressive Complex 1 (PRC1) may play a negative role in the granulocytic differentiation of acute promyelocytic leukemia (APL) cells. We first examined the expression of PRC1 genes during all-trans retinoic acid (ATRA)-mediated differentiation of human HL-60 cells, and identified PCGF2 as a gene down-regulated by ATRA in a time-dependent manner. Upon gene silencing of PCGF2 with lentiviral short hairpin RNA, granulocytic differentiation was induced as assessed by differentiation marker gene expression, nitroblue tetrazolium staining, Wright-Giemsa staining, and cell cycle analysis. We next identified HOXA7 as a homeobox gene up-regulated by ATRA and successfully induced granulocytic differentiation by overexpression of HOXA7. We next tested the relationship between PCGF2 and HOXA7 by quantifying the changes in HOXA7 and PCGF2 expression upon PCGF2 gene silencing and HOXA7 overexpression, respectively. HOXA7 expression was up-regulated by PCGF2 gene silencing, while PCGF2 expression remained unchanged by ectopic HOXA7 expression, suggesting PCGF2 as acting upstream of HOXA7. Finally, chromatin immunoprecipitation assay was performed with HOXA7 chromatin. We observed gene-specific reduction in direct binding of Pcgf2 protein to HOXA7 chromatin upon PCGF2 gene silencing. Taken together, these results support the notion that down-regulation of PCGF2 is sufficient to induce granulocytic differentiation of HL-60 cells via de-repression of HOXA7 gene expression. In conclusion, we report that PCGF2, a PRC1 gene, played a negative role in the granulocytic differentiation of human APL cells.