Transcriptional regulation of interleukin-6 receptor by PML::RARα fusion protein

doi:10.16138/j.1673-6087.2025.02.08

Abstract

Abstract:

Objective To investigate the regulatory mechanism of PML::RARα fusion protein on interleukin-6 receptor (IL-6R) and the effects of IL-6R on the proliferation and differentiation of acute promyelocytic leukemia (APL) cells.Methods The expression levels of IL-6R in APL cells were analyzed using the GSE12662 and GSE10358 datasets. Reverse transcription quantitative real-time quantitative PCR（RT-qPCR） was performed to detect IL-6R mRNA expression in NB4 cells before and after all-trans retinoic acid (ATRA) treatment, as well as in PR9 cells before and after Zn²⁺ induction. Chromatin immunoprecipitation (ChIP)-seq data analysis, ChIP-qPCR experiments, and luciferase reporter gene activity assays were performed to explore the regulatory mechanism of PML::RARα on IL-6R. An IL-6R expression plasmid was constructed for NB4 cells via retrovirus. Cell proliferation was assessed using the cell counting kit-8 (CCK-8) assay, and CD11b expression was detected by flow cytometry.Results Analysis of the GSE12662 dataset revealed that the expression level of IL-6R in APL cells (12.20 ± 0.41) was significantly lower than that in normal promyelocytes (13.14 ± 0.47, t = 4.289, P < 0.001) and polymorphonuclear cells (14.82 ± 0.40, t = 12.35, P < 0.001). Moreover, analysis of the GSE10358 dataset showed that IL-6R expression in APL patients (5.93 ± 0.84) was significantly lower than that in non-APL AML patients (6.50 ± 0.87, t = 3.91, P < 0.001). PML::RARα directly bound to the promoter region of IL-6R to inhibit its transcriptional activity resulting in the low expression. Overexpression of IL-6R in the APL-derived NB4 cells significantly inhibited cell proliferation. Four days after transfection, the optical density values measured by the CCK-8 assay were 0.86 ± 0.01 and 0.40 ± 0.01, respectively (t = 32.66, P < 0.001). Simultaneously, cell differentiation was significantly enhanced. The ratio of the CD11b positive cells increased from 3.10 % ± 1.22 % to 14.4 % ± 1.11 % (t = 11.84, P < 0.001).Conclusions IL-6R is a target gene of PML::RARα, demonstrating that PML::RARα can suppress IL-6R transcription by binding to its promoter region. It is illustrated that IL-6R inhibited the cell proliferation and induced partial differentiation in APL cells.

Key words: Interleukin-6 receptor, Acute promyelocytic leukemia, PML::RARα, Cell proliferation, Cell differentiation

CLC Number:

R733.7

ZHAO Lingling, CUI Canqi, MI Jianqing. Transcriptional regulation of interleukin-6 receptor by PML::RARα fusion protein[J]. Journal of Internal Medicine Concepts & Practice, 2025, 20(02): 146-151.

Figures/Tables 3

References 27

[1]	Chen Z, Chen SJ. Poisoning the devil[J]. Cell, 2017, 168(4): 556-560. doi: S0092-8674(17)30114-9 pmid: 28187275
[2]	《中国急性早幼粒细胞白血病诊疗指南(2018年版)》发布[J]. 中华医学信息导报, 2018, 33(7): 9.
[3]	de thé H, Pandolfi PP, Chen Z. Acute promyelocytic leukemia: a paradigm for oncoprotein-targeted cure[J]. Cancer Cell, 2017, 32(5): 552-560. doi: S1535-6108(17)30455-5 pmid: 29136503
[4]	Tan Y, Wang X, Song H, et al. A PML/RARalpha direct target atlas redefines transcriptional deregulation in acute promyelocytic leukemia[J]. Blood, 2021, 137(11):1503-1516.
[5]	Villiers W, Kelly A, He X, et al. Multi-omics and machine learning reveal context-specific gene regulatory activities of PML::RARA in acute promyelocytic leukemia[J]. Nat Commun, 2023, 14(1): 724. doi: 10.1038/s41467-023-36262-0 pmid: 36759620
[6]	Rose-John S. Interleukin-6 signalling in health and disease[EB/J]. F1000Res, 2020. https://pmc.ncbi.nlm.nih.gov/articles/PMC7443778/.
[7]	Garbers C, Heink S, Korn T, et al. Interleukin-6: designing specific therapeutics for a complex cytokine[J]. Nat Rev Drug Discov, 2018, 17(6): 395-412. doi: 10.1038/nrd.2018.45 pmid: 29725131
[8]	Mei Y, Ren K, Liu Y, et al. Bone marrow-confined IL-6 signaling mediates the progression of myelodysplastic syndromes to acute myeloid leukemia[J]. J Clin Invest, 2022, 132(17): e152673.
[9]	张丽霞, 杜思博, 李琳, 等. 白介素-6及其受体与多种健康结局:孟德尔随机化的证据图[J]. 兰州大学学报(医学版), 2024, 50(10): 62-72.
[10]	Wang MJ, Zhang HL, Chen F, et al. The double-edged effects of IL-6 in liver regeneration, aging, inflammation, and diseases[J]. Exp Hematol Oncol, 2024, 13(1): 62.
[11]	Stahl M, Tallman MS. Differentiation syndrome in acute promyelocytic leukaemia[J]. Br J Haematol, 2019, 187(2): 157-162.
[12]	Hui H, Yang H, Dai Q, et al. Oroxylin A inhibits ATRA-induced IL-6 expression involved in retinoic acid syndrome by down-regulating CHOP[J]. Gene, 2014, 551(2):230-235. doi: 10.1016/j.gene.2014.08.061 pmid: 25192658
[13]	赵世香, 葛圆圆, 李增政, 等. 细胞因子对初诊急性早幼粒细胞白血病患者早期死亡的影响[J]. 中国实验血液学杂志, 2023, 31(5): 1315-1321.
[14]	陈芳, 马凤霞, 李洋, 等. 人脐带间充质干细胞分泌白介素-6对白血病细胞分化的影响[J]. 中国医学科学院学报, 2016, 38(2): 164-168. doi: 10.3881/j.issn.1000-503X.2016.02.007
[15]	Wang X, Tan Y, Li Y, et al. Repression of CDKN2C caused by PML/RARalpha binding promotes the proliferation and differentiation block in acute promyelocytic leukemia[J]. Front Med, 2016, 10(4): 420-429.
[16]	Payton JE, Grieselhuber NR, Chang LW, et al. High throughput digital quantification of mRNA abundance in primary human acute myeloid leukemia samples[J]. J Clin Invest, 2009, 119(6): 1714-1726. doi: 10.1172/JCI38248 pmid: 19451695
[17]	Tomasson MH, Xiang Z, Walgren R, et al. Somatic mutations and germline sequence variants in the expressed tyrosine kinase genes of patients with de novo acute myeloid leukemia[J]. Blood, 2008, 111(9): 4797-808. doi: 10.1182/blood-2007-09-113027 pmid: 18270328
[18]	Qian M, Jin W, Zhu X, et al. Structurally differentiated cis-elements that interact with PU.1 are functionally distinguishable in acute promyelocytic leukemia[J]. J Hematol Oncol, 2013, 6:25.
[19]	de thé H, Chen Z. Acute promyelocytic leukaemia: novel insights into the mechanisms of cure[J]. Nat Rev Cancer, 2010, 10(11):775-783. doi: 10.1038/nrc2943 pmid: 20966922
[20]	Kishimoto T, Akira S, Taga T. Interleukin-6 and its receptor: a paradigm for cytokines[J]. Science, 1992, 258(5082):593-597. doi: 10.1126/science.1411569 pmid: 1411569
[21]	Inoue K, Sugiyama H, Ogawa H, et al. Expression of the interleukin-6 (IL-6), IL-6 receptor, and gp130 genes in acute leukemia[J]. Blood, 1994, 84(8): 2672-2680. pmid: 7919380
[22]	刘爽, 奚永志, 郭斯启, 等. IL-6受体α亚单位mRNA和蛋白在人白血病细胞中的表达[J]. 中国实验血液学杂志, 2002, 10(1): 22-26.
[23]	Scheller J, Chalaris A, Schmidt-Arras D, et al. The pro- and anti-inflammatory properties of the cytokine interleukin-6[J]. Biochim Biophys Acta, 2011, 1813(5):878-888. doi: 10.1016/j.bbamcr.2011.01.034 pmid: 21296109
[24]	Reeh H, Rudolph N, Billing U, et al. Response to IL-6 trans- and IL-6 classic signalling is determined by the ratio of the IL-6 receptor alpha to gp130 expression: fusing experimental insights and dynamic modelling[J]. Cell Commun Signal, 2019, 17(1): 46.
[25]	Schreiber S, Aden K, Bernardes JP, et al. Therapeutic interleukin-6 trans-signaling inhibition by olamkicept (sgp130fc) in patients with active inflammatory bowel disease[J]. Gastroenterology, 2021, 160(7): 2354-2366. doi: 10.1053/j.gastro.2021.02.062 pmid: 33667488
[26]	Zhang S, Chen B, Wang B, et al. Effect of induction therapy with olamkicept vs placebo on clinical response in patients with active ulcerative colitis[J]. JAMA, 2023, 329(9): 725-734.
[27]	Kantarjian HM, DiNardo CD, Kadia TM, et al. Acute myeloid leukemia management and research in 2025[J]. CA Cancer J Clin, 2025, 75(1): 46-67.

转染天数（d）	空载	IL-6R	t	P
0	0.24 ± 0.00	0.24 ± 0.00	0.13	0.90
1	0.34 ± 0.02	0.31 ± 0.01	2.22	0.09
2	0.39 ± 0.01	0.34 ± 0.02	4.01	0.02
3	0.54 ± 0.01	0.35 ± 0.01	35.74	<0.0001
4	0.86 ± 0.01	0.40 ± 0.01	32.66	<0.0001