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HTm4基因在造血细胞周期调控中的作用

李锦, 谢超, 谢小燕, 王冬梅, 裴雪涛

军事医学科学院输血研究所干细胞研究室.北京 100850

摘要

为了研究HTm4基因在造血细胞细胞周期调控中的作用,以佛波酯(phorbol 12--myristate 13--acetate,PMA)诱导K562细胞分化为模型,利用流式细胞术(FACS)及半定量RT--PCR对分化过程中细胞周期的变化及HTm4基因的表达进行了分析,并利用Tet--Off调控表达系统,将HTm4基因以及C端功能域缺失的HTm4--ct转染K562细胞,观察对细胞周期的影响。结果表明,PMA同时引起了K562细胞的增殖和分化,G(0)/G_(1)期细胞的比例以及HTm4基因的表达均呈现出波浪形的变化趋势,说明HTm4基因可能参与了细胞退出细胞周期的过程。HTm4基因转染后引起K562细胞滞留于G(0)/G_(1)期,但C端功能域缺失的HTm4--ct没有此作用,说明C端功能域在HTm4基因调控细胞周期的功能中发挥重要作用。

关键词： 细胞周期; HTm4基因; K562细胞

Regulatory role of HTm4 gene in hematopoietic cell cycle

Li Jin, Xie Chao, Xie Xiaoyan, Wang Dongmei, Pei Xuetao

Department of Stem Cell Biology, Beijing Institute of Transfusion Medicine.Beijing 100850

Abstract

Cell cycle progression is tightly regulated in hematopoietic stem cells. The cycle state decides cells'fates, which include self-renewal, proliferation and differentiation. Proper cell cycle regulation is a pivotal element for the maintenance of hematopoiesis homeostasis. Htm4 is a newly identified specific cell cycle regulator of the hematopoietic cell. Through interacting with KAP-CDK2 complex, it arrests cells in G(0)/G_(1) phase. K562 is a human chronic myelogenous leukemia cell; it could be induced to megakaryoblast by PMA. Such differentiation must associate with cell cycle change. To further clarify Htm4's function in hematopoietic cell cycle regulation, K562 cells were treated with PMA. Cell cycle change was analysed using flow cytometric system. And during the induction process gene expression of HTm4 as well as CycleE and CDK2, which are responsible for G_(1) to S transition, were analysed using semi-quantitative RT-PCR. The C-terminal domain of HTm4 protein has been shown to be important for HTm4's binding with KAP-CDK2 complex. To determine its impact on HTm4's function, HTm4 and C-terminal truncated HTm4 (HTm4-ct) were transfected into K562 cells using Tet-Off regulation expression system. Their influence on cell cycle was observed. Results showed that PMA induced both expansion and differentiation of K562 cells as measured by cell number count and NBT staining respectively. During PMA treatment, G(0)/G_(1) cell proportion and HTm4 expression displayed coordinated change, which suggested that HTm4 might drive K562 cells out of cell cycle but was not involved in the quiescence maintenance. Additionally, transfection of HTm4 caused G(0)/G_(1) arrest in K562 cells, while transfection of HTm4-ct did not. .It is therefore suggested that the C-terminal domain is important for the function of HTm4 in cell cycle regulation.

Key words: Cell cycle;HTm4 gene;K562 cells

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引用本文：

李锦, 谢超, 谢小燕, 王冬梅, 裴雪涛. HTm4基因在造血细胞周期调控中的作用[J]. 生理学报 2005; 57 (2): .

Li Jin, Xie Chao, Xie Xiaoyan, Wang Dongmei, Pei Xuetao. Regulatory role of HTm4 gene in hematopoietic cell cycle. Acta Physiol Sin 2005; 57 (2): (in Chinese with English abstract).