Quantitative Proteomics Reveals CBX4 Promotes Cervical Cancer Cell Proliferation by Regulating PFKFB4

Abstract CBX4 is an E3 SUMO ligase involved in the initiation and progression of multiple malignancies. However, the molecular mechanism underlying the functional significance of CBX4 in cervical cancer (CC) cells remains unclear. Here, we used CRISPR/Cas9 genome editing to generate CBX4-knockout cell lines and found that CBX4 loss markedly suppressed HeLa cell proliferation. A label-free quantitative proteomics approach based on data-independent acquisition (DIA) was employed to systematically profile CBX4-regulated proteins in HeLa cells. In total, 263 proteins showed significant changes upon CBX4 knockout, among which 171 proteins were downregulated, and 92 were upregulated. Bioinformatics analysis revealed that these differentially expressed proteins (DEPs) are associated with diverse cellular pathways, especially metabolic pathways. Furthermore, our results demonstrated that CBX4 promotes CC cell proliferation by regulating the expression of 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 4 (PFKFB4). Mechanistically, PFKFB4 is a direct target of the transcriptional activator SP1, and CBX4 interacts with SP1 and increases its SUMOylation. Therefore, CBX4 may exert its effects in CC cells through enhancing SP1-mediated PFKFB4 transcription. In conclusion, the current study establishes a proteome-wide CBX4 regulatory network and provides novel insights into the molecular pathogenesis of CC.