Extracellular pyruvate kinase M2 induces cell migration through p-Tyr42 RhoA-mediated superoxide generation and epithelial-mesenchymal transition

• Published in: Free radical biology & medicine 2023-11, Vol.208, p.614-629
• Main Authors: Hamza, Amir, Cho, Jung Yoon, Cap, Kim Cuong, Hossain, Abu Jubayer, Kim, Jae-Gyu, Park, Jae-Bong
• Format: Article
• Language: English
• Subjects: Animals; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition - genetics; Humans; Mice; Neoplasms; p-p47phox; p-Tyr42 RhoA; PKM2; Pyruvate Kinase - genetics; Pyruvate Kinase - metabolism; Signal Transduction; Superoxides; TAM; Thyroid Hormone-Binding Proteins; TME; Tumor Microenvironment - genetics
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/j.freeradbiomed.2023.09.016

In the tumor microenvironment (TME), communication between cancer cells and tumor-associated macrophages (TAMs) through secreted extracellular proteins promotes cancer progression. Here, we observed that co-culturing cancer cells (4T1) and macrophage cells (Raw264.7) significantly enhanced superoxide production in both cell types. Using MALDI-TOF, we identified PKM2 as a highly secreted protein by Raw264.7 cells and bone marrow-derived monocytes. The extracellular recombinant PKM2 protein not only enhanced cancer cell migration and invasion but also increased superoxide production. Additionally, PKM2 was found to associate with the cell surface, and its binding to integrin α5/β1 receptor was inhibited by antibodies specifically targeting it. Furthermore, we investigated downstream signaling pathways involved in PKM2-induced superoxide production. We found that knock-down of RhoA and p47phox using siRNAs effectively abolished superoxide generation in response to extracellular PKM2. Notably, extracellular PKM2 triggered the phosphorylation of p47phox at Ser345 residue and RhoA at Tyr42 residue (p-Tyr42 RhoA). Moreover, extracellular PKM2 exerted regulatory control over the expression of key epithelial-mesenchymal transition (EMT) markers, including ZEB1, Snail1, vimentin, and E-cadherin. Interestingly, p-Tyr42 RhoA translocated to the nucleus, where it bound to the ZEB1 promoter region. In light of these findings, we propose that extracellular PKM2 within the TME plays a critical role in tumorigenesis by promoting cancer cell migration and invasion through RhoA/p47phox signaling pathway. [Display omitted] •Extracellular PKM2 secreted from macrophages stimulates superoxide production in cancer cells.•Superoxide production is regulated by p-Tyrosine42 RhoA and p-p47phox.•p-Tyrosine42 RhoA translocates to the nucleus, where it binds to the promoter of ZEB1 and induces its expression.•Extracellular PKM2 binds to integrin α5β1 and phosphoenol pyruvate (PEP) inhibits PKM2 binding to the integrin α5β1.