天津医药

天津医药 ›› 2021, Vol. 49 ›› Issue (7): 688-693.doi: 10.11958/20210070

• 细胞与分子生物学 • 上一篇    下一篇

RPL15在K562细胞向红系分化过程中的功能和机制研究

彭涛，周唯君，周栋珍，李化，兰金芝，李丹，周艳华，舒莉萍   

  1. 1贵州医科大学细胞工程生物医药技术国家地方联合工程实验室，贵州省再生医学重点实验室，贵州医科大学基础医学院免疫 教研室（邮编550004）；2中国医学科学院成体干细胞转化研究重点实验室
  • 收稿日期:2021-01-13 修回日期:2021-04-14 出版日期:2021-07-15 发布日期:2021-07-12
  • 作者简介:彭涛（1996），男，硕士在读，主要从事干细胞及血液免疫相关研究。E-mail：496875493@qq.com
  • 基金资助:
    国家自然科学基金资助项目（31860325）；贵州省科技计划项目（黔科合基础［2019］1436）；贵州省科技计划项目（黔科中引 ［2019］4008）

Study on the function and mechanism of RPL15 in erythroid differentiation of K562 cells

PENG Tao, ZHOU Wei-jun, ZHOU Dong-zhen, LI Hua, LAN Jin-zhi, LI Dan, ZHOU Yan-hua, SHU Li-ping   

  1. 1 National & Guizhou Joint Engineering Laboratory for Cell Engineering and Biomedicine Technique, Guizhou Province Key Laboratory of Regenerative Medicine, Department of Immunology, School of Basic Medicine, Guizhou Medical University, Guiyang 550004,China; 2 Key Laboratory of Adult Stem Cell Translational Research, Chinese Academy of Medical Sciences
  • Received:2021-01-13 Revised:2021-04-14 Published:2021-07-15 Online:2021-07-12

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摘要： 目的　研究RPL15基因在K562细胞向红系诱导分化过程中的表达变化，并确定其对珠蛋白表达及红系分化的影响。方法　用氯化血红素诱导K562细胞向红系分化，采用实时荧光定量PCR（qPCR）及Western blot法检测RPL15在K562红系分化过程中的表达情况。利用RNA干扰（RNAi）技术抑制K562细胞中RPL15的表达，进而通过四甲基联苯胺染色、qPCR及流式细胞技术检测K562细胞中血红蛋白、红系分化相关基因以及CD235a和CD71表达的变化。结果　在K562细胞向红系分化过程中，RPL15的mRNA及蛋白水平均呈先升高后下降趋势。与对照组相比，干扰RPL15表达后，向红系分化24、48和72 h的K562细胞血红蛋白染色阳性率均明显下降；向红系分化的K562细胞中α-、β-、ε-、γ-珠蛋白及红系分化相关基因GATA1和HSP70相对表达量均明显下调（均P＜0.05）；而抑癌基因P53的mRNA表达水平显著上调（P＜0.01）。同时CD235a+CD71+ K562细胞比例在RPL15干扰后显著降低（P＜0.01）。结论　抑制RPL15表达可抑制K562细胞向红系分化，提示RPL15正调控红系分化过程。同时P53相关通路、GATA1及HSP70可能参与了RPL15对红系分化的调控过程。

关键词: K562细胞, 核糖体蛋白质类, GATA1转录因子, 基因, p53, HSP70热休克蛋白质类, RPL15基因, 红系分化

Abstract: Objective　To study the expression changes of RPL15 during erythroid differentiation of K562 cells and to determine its effect on globin expression and erythroid differentiation. Methods　Erythroid differentiation of K562 cells was induced by hemin. The expression of RPL15 in K562 erythroid differentiation was detected by quantitative Real-Time PCR(qPCR) and Western blot assay. RNA interference (RNAi) technology was used for the inhibition of RPL15 expression in K562 cells. The expression of hemoglobin, erythroid differentiation-related genes and the expression of CD235a and CD71 in K562 cells were detected by tetramethyl benzidine staining, qPCR and flow cytometry. Results　In the process of erythroid differentiation of K562 cells, the mRNA and protein levels of RPL15 were up-regulated at first and then decreased. Compared with the control group, the interference with the RPL15 expression significantly decreased the positive rate of hemoglobin staining in K562 cells differentiated into erythroid at 24, 48 and 72 hours. The relative expressions of α-, β-, ε-, γ-globin and erythroid differentiation-related genes GATA1 and HSP70 were significantly down-regulated in erythroid differentiated K562 cells (P＜0.05). The relative expression of tumor suppressor gene P53 was significantly up-regulated (P＜0.01). Meanwhile, the proportion of CD235a+CD71+ K562 cells decreased significantly after RPL15 interference (P＜0.01). Conclusion　Inhibition of RPL15 expression can inhibit erythroid differentiation of K562 cells, suggesting that RPL15 is regulating the process of erythroid differentiation. At the same time, P53-related pathway, GATA1 and HSP70 may be involved in the regulation of erythroid differentiation by RPL15.

Key words: K562 cells, ribosomal proteins, GATA1 transcription factor, genes, p53, HSP70 heat-shock proteins, RPL15 gene, erythroid differentiation