COORDINATE EXPRESSION AND DEVELOPMENTAL ROLE OF ID2 PROTEIN AND TAL1/E2A HETERODIMER IN ERYTHROID PROGENITOR DIFFERENTIATION
COORDINATE EXPRESSION AND DEVELOPMENTAL ROLE OF ID2 PROTEIN AND TAL1/E2A HETERODIMER IN ERYTHROID PROGENITOR DIFFERENTIATION
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DOI：
10.1182/blood.v86.1.164.bloodjournal861164
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发表时间：
1995-07-01
期刊：
影响因子：
20.3
通讯作者：
PESCHLE, C
中科院分区：
文献类型：
--
作者：
CONDORELLI, G;VITELLI, L;PESCHLE, C
Id蛋白和碱性螺旋-环-螺旋（bHLH）蛋白在不同生物环境中指定细胞命运决定中起主要作用。对永生化细胞系的研究表明Id和TAL 1/E2 A bHLH基因在造血中具有潜在的作用。然而，尚不确定这些观察结果是否反映了正常的造血。我们研究了在液体悬浮培养的纯化的造血祖细胞（HPCs）的表达模式Id 2和TAL 1/E2 A基因进行红系或粒系分化的第一个培养周和成熟的终末细胞在第二周。在静止的，新鲜纯化的HPC，Id 2 mRNA的检测通过逆转录酶-聚合酶链反应（RT-PCR），而TAL 1和E2 A的mRNA没有。在红系分化开始时，Id 2 mRNA下调，而E2 A和TAL 1 mRNA伴随上调：它们的表达在成红细胞水平进一步增加。相反，除了在第10至12天的晚期下降外，Id 2在粒细胞生成培养物中不被下调，而TAL 1和E2 A仅在粒细胞生成分化的第一周被短暂诱导。通过迁移率变动分析评估的TAL 1/E2 A异二聚体的表达模式与RT-PCR结果一致（除了在红系成熟晚期异二聚体的水平较低）。通过Western印迹分析的TAL 1蛋白水平显示出与凝胶位移结果一致的模式。对纯化的HPC用Id 2或TAL 1 mRNA的硫代磷酸反义寡脱氧核苷酸处理进行功能实验。结果与表达研究严格一致：与未处理或乱序寡聚体处理的对照HPC相比，抗Id 2寡聚体（α-Id 2）导致红系细胞集落形成的显著剂量依赖性增加，而α-TAL 1诱导对红系细胞集落的选择性剂量相关抑制作用。最后，当将鼠和人谷胱甘肽-S-转移酶（GST）-Id 2多肽添加到源自红系培养细胞的核提取物中时，其竞争TAL 1/E2 A特异性DNA结合活性，从而表明Id 2与TAL 1/E2 A复合物的生物化学和功能相互作用。这些新的观察结果表明，在正常红系分化中，抑制性Id蛋白（Id 2）和刺激性bHLH/bHLH异源二聚体（TAL 1/E2 A）的协调表达和功能。(C)1995年，美国血液学会。
The Id proteins and basic helix-loop-helix (bHLH) proteins play major roles in specifying cell fate decisions in diverse biologic settings. A potential role for Id and TAL 1/E2A bHLH genes in hematopoiesis has been suggested by studies on immortalized cell lines. However, it is uncertain whether these observations reflect normal hematopoiesis. We have investigated the expression pattern of Id2 and TAL1/E2A genes in liquid suspension culture of purified hematopoietic progenitor cell (HPCs) undergoing erythroid or granulopoietic differentiation in the first culture week and maturation to terminal cells in the second week. In quiescent, freshly purified HPCs, Id2 mRNA is detected by reverse transcriptase-polymerase chain reaction (RT-PCR), whereas TAL1 and E2A mRNAs are not. At the onset of erythroid differentiation, Id2 mRNA is downregulated, while E2A and TAL1 mRNAs are concomitantly upregulated: their expression is further increased at erythroblast level. Conversely, Id2 is not downmodulated in granulopoietic culture, except for a late decline at day 10 to 12, while TAL1 and E2A are only transiently induced in the first week of granulopoietic differentiation. The expression pattern of the TAL1/E2A heterodimer, as evaluated by mobility shift assay, is consistent with RT-PCR results (except for lower levels of the heterodimer in late erythroid maturation). TAL1 protein level, analyzed by Western blot, shows a pattern consistent with gel-shift results. Functional experiments were performed on purified HPCs treated with phosphorothioate antisense oligodeoxynucleotides to Id2 or TAL1 mRNA. The results are strictly consistent with the expression studies: anti-Id2 oligomer (alpha-Id2) causes a significant dose-dependent increase of erythroid colony formation, whereas alpha-TAL1 induces a selective dose-related inhibitory effect on erythroid colonies, as compared with untreated or scrambled oligomer-treated control HPCs. Finally, murine and human glutathione-S-transferase (GST)-Id2 polypeptides compete the TAL1/E2A-specific DNA binding activity when added to the nuclear extracts derived from erythroid culture cells, thus indicating biochemical and suggesting functional interaction of Id2 with the TAL1/E2A complex. These novel observations indicate a coordinate expression and function of an inhibitory Id protein (Id2) and a stimulatory bHLH/bHLH heterodimer (TAL1/E2A) in normal erythroid differentiation. (C) 1995 by The American Society of Hematology.