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

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.