mTOR/HK2信号通路的激活减轻了结直肠细胞中PYCR2缺失的影响。

Activation of mTOR/HK2 signaling mitigates effects of PYCR2 depletion in colorectal cells.

作者信息

Chen Li, Yuan Yuan, Zhang Nian, Huang Qianqian, Zhou Yu

机构信息

Zhuhai Hospital of Integrated Traditional Chinese and Western Medicine, China.

出版信息

Tissue Cell. 2025 Apr;93:102729. doi: 10.1016/j.tice.2025.102729. Epub 2025 Jan 10.

DOI:10.1016/j.tice.2025.102729

PMID:39808866

Abstract

BACKGROUND

Colorectal cancer (CRC) is one of the aggressive malignant tumors. Studies have shown that glycolysis promotes the proliferation of colorectal cancer cells and that PYCR2 is involved in cancer progression by affecting cellular glycolysis. In addition, PYCR2 is upregulated in colorectal cancer cell lines and can affect cellular autophagy.

METHODS

Si-PYCR2 was used to interfere with PYCR2 in colorectal cancer cells, and the cells were treated with the addition of autophagy inhibitor 3-MA or mTOR agonist MHY1485. The expression of LC3B was detected by immunofluorescence, and the expression of autophagy and glycolytic proteins was detected by Western blot. XF96 extracellular flux analyzer was used to detect the ECAR and OCR of the cells, and biochemical kits were used to detect the levels of glucose consumption, lactate secretion, and ATP production in the cells.

RESULTS

PYCR2 expression was up-regulated in colorectal cancer cell lines. si-PYCR2 interference enhanced the fluorescence intensity of LC3B in the cells, inhibited the expression of p62 proteins but enhanced the expression of ATG5, ATG7, and LC3-II/I proteins, which indicated an enhanced level of autophagy in colorectal cancer cells. In addition, PYCR2 depletion also inhibited cellular glycolysis as well as mTOR/HK2 signaling. However, the addition of 3-MA resulted in an increase in cellular ECAR while a decrease in OCR, and an increase in the levels of glucose consumption, lactate and ATP production, as well as the expressions of glycolytic proteins (GLUT1, PGK1, ENO1, PKM2), which suggested the glycolysis of cells was enhanced. In addition, MHY1485 treatment not only inhibited autophagy but also enhanced glycolysis in colorectal cancer cells.

CONCLUSION

Interference with PYCR2 corrected autophagy-dependent glycolysis levels in colorectal cancer cells via mTOR/HK2 signaling. Activation of mTOR/HK2 signaling mitigated the effects of PYCR2 depletion in colorectal cells.

摘要

背景

结直肠癌（CRC）是侵袭性恶性肿瘤之一。研究表明，糖酵解促进结直肠癌细胞增殖，且PYCR2通过影响细胞糖酵解参与癌症进展。此外，PYCR2在结直肠癌细胞系中上调，并且能够影响细胞自噬。

方法

使用Si-PYCR2干扰结直肠癌细胞中的PYCR2，并添加自噬抑制剂3-MA或mTOR激动剂MHY1485处理细胞。通过免疫荧光检测LC3B的表达，通过蛋白质免疫印迹法检测自噬和糖酵解相关蛋白的表达。使用XF96细胞外流量分析仪检测细胞的细胞外酸化率（ECAR）和氧消耗率（OCR），并使用生化试剂盒检测细胞中葡萄糖消耗、乳酸分泌和ATP产生水平。

结果

PYCR2在结直肠癌细胞系中表达上调。si-PYCR2干扰增强了细胞中LC3B的荧光强度，抑制了p62蛋白的表达，但增强了ATG5、ATG7和LC3-II/I蛋白的表达，这表明结直肠癌细胞中的自噬水平增强。此外，PYCR2缺失还抑制了细胞糖酵解以及mTOR/HK2信号传导。然而，添加3-MA导致细胞ECAR增加而OCR降低，葡萄糖消耗、乳酸和ATP产生水平增加，以及糖酵解蛋白（GLUT1、PGK1、ENO1、PKM2）的表达增加，这表明细胞的糖酵解增强。此外，MHY1485处理不仅抑制了结直肠癌细胞的自噬，还增强了糖酵解。