CSN5/HSF/SPI1/PU.1轴调节低细胞性骨髓增生异常综合征患者的细胞增殖。

The CSN5/HSF/SPI1/PU.1 Axis Regulates Cell Proliferation in Hypocellular Myelodysplastic Syndrome Patients.

作者信息

Yu Zheng-Ping, Jian Zi-Ying, Sun Ai-Ning, Chen Bao-An, Ge Zheng

机构信息

Department of Hematology (Key Department of Jiangsu Medicine), Zhong Da Hospital, Southeast University, Nanjing.

Hematology Division, Suzhou Medical University, Suzhou, China.

出版信息

J Pediatr Hematol Oncol. 2023 Oct 1;45(7):e873-e878. doi: 10.1097/MPH.0000000000002712. Epub 2023 Aug 1.

DOI:10.1097/MPH.0000000000002712

PMID:37526438

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10521780/

Abstract

OBJECTIVE

This study explored the relationship between the activation of the jak/stat3 signaling pathway and the CSN5 gene transcript and protein expression levels in the hematopoietic stem cells of patients with myelodysplastic syndromes (MDSs). This study also aimed to investigate the correlation between the expression level of CSN5 and the deubiquitination of HSF1, as well as the transcript level of the spi1/pu.1 genes to explore the pathogenesis of MDS.

MATERIALS AND METHODS

We isolated cells from normal individuals and MDS patients, and the mRNA and protein expression levels of spi1/pu.1 in cd34+ cells (hematopoietic stem cells) were measured by PCR and western blotting, respectively. A ChIP assay was used to detect the binding of HSF1 to the spi1/pu.1 promoter in cd34+ cells. The ubiquitination of HSF1 in cd34+ cells was detected by CO-IP. The binding of HSF1 and Fbxw7α was detected in in cd34+ cells by CO-IP. The binding of HSF1 and CSN5 was evaluated. A luciferase reporter assay was used to detect the effect of STAT3 on CSN5 promoter activation in cd34+ cells. Western blotting was used to detect the phosphorylation of STAT3 in cd34+ cells of MDS patients. The binding of STAT3 and C/EBP beta in cd34+ cells was detected by CO-IP.

RESULTS

Inhibition of SPI1/PU.1 expression was observed in MDS samples with low proliferation ability. Further experiments proved that phosphorylation of STAT3 affected CSN5 function and mediated the ubiquitination of HSF, the upstream regulator of SPI1/PU.1 transcription, which led to the inhibition of SPI1/PU.1 expression. Restoration of CSN5 rescued the inhibition of HSF1 ubiquitination, causing SPI1/PU.1 transcription to resume and increasing SPI1/PU.1 expression, promoting the recovery of cell proliferation in hypocellular MDS.

CONCLUSIONS

Our research revealed the regulatory role of the CSN5/HSF/SPI1/PU.1 axis in hypocellular MDS, providing a probable target for clinical intervention.

摘要

目的

本研究探讨骨髓增生异常综合征（MDS）患者造血干细胞中jak/stat3信号通路激活与CSN5基因转录及蛋白表达水平之间的关系。本研究还旨在研究CSN5表达水平与HSF1去泛素化以及spi1/pu.1基因转录水平之间的相关性，以探讨MDS的发病机制。

材料与方法

我们从正常个体和MDS患者中分离细胞，分别通过PCR和蛋白质印迹法检测cd34+细胞（造血干细胞）中spi1/pu.1的mRNA和蛋白表达水平。采用染色质免疫沉淀（ChIP）试验检测cd34+细胞中HSF1与spi1/pu.1启动子的结合情况。通过免疫共沉淀（CO-IP）检测cd34+细胞中HSF1的泛素化。通过CO-IP检测cd34+细胞中HSF1与Fbxw7α的结合情况。评估HSF1与CSN5的结合情况。采用荧光素酶报告基因试验检测STAT3对cd34+细胞中CSN5启动子激活的影响。用蛋白质印迹法检测MDS患者cd34+细胞中STAT3的磷酸化情况。通过CO-IP检测cd34+细胞中STAT3与C/EBPβ的结合情况。

结果

在增殖能力低的MDS样本中观察到SPI1/PU.1表达受到抑制。进一步实验证明，STAT3的磷酸化影响CSN5功能并介导SPI1/PU.1转录上游调节因子HSF的泛素化，从而导致SPI1/PU.1表达受到抑制。CSN5的恢复挽救了HSF1泛素化的抑制，使SPI1/PU.1转录恢复并增加SPI1/PU.1表达，促进低细胞性MDS中细胞增殖的恢复。