To understand why this finding matters, it helps to begin with the biology of protein turnover. Inside every cell, proteins that are no longer needed, damaged, or dangerous are tagged with small marker proteins called ubiquitins. Enzymes assemble these tags into chains, and the way the chains are linked determines the tagged protein’s fate. K48-linked polyubiquitin chains, in which ubiquitin molecules are connected through a specific lysine residue, act as a molecular kiss of death: they flag the protein for delivery to the proteasome, the cell’s garbage disposal, where it is chopped into fragments. Deubiquitinating enzymes, or DUBs, do the opposite. They snip ubiquitin tags off proteins, rescuing them from degradation. Among the largest DUB family is the ubiquitin-specific proteases, or USPs, and when one of these enzymes removes death marks from a protein that promotes cancer, the effect can be dramatic: the oncoprotein accumulates, and the cell drifts further toward uncontrolled growth.

CIAP2, also known as baculoviral IAP repeat containing protein 2 and a member of the inhibitor of apoptosis protein family, was already known to be overexpressed in hepatocellular carcinoma and to function there as an oncogene. What remained incompletely understood was how tumor cells manage to keep CIAP2 levels so high. The research team approached this question systematically. Rather than guessing which of the many ubiquitin-specific proteases might be responsible, they screened USPs that could plausibly regulate CIAP2 expression and identified USP45 as a key regulator that enhances CIAP2 levels in hepatocellular carcinoma cells. That screening result set the stage for a deeper interrogation of what USP45 is doing in these tumors and whether its activity has real consequences for cancer behavior.

Bioinformatic analysis of clinical data provided the first hint that USP45 is more than a laboratory curiosity. The researchers found that USP45 is overexpressed in hepatocellular carcinoma and, critically, that elevated USP45 is associated with worse patient survival. In other words, the enzyme is not merely present in these tumors; its abundance tracks with the outcome that matters most to patients. This kind of correlation does not by itself prove causation, but it supplied the rationale for the functional experiments that followed. If USP45 truly helps drive the disease, then removing it from cancer cells should blunt their malignant behavior in measurable ways.

That is precisely what the team observed. When the researchers knocked down USP45 in two hepatocellular carcinoma cell lines, SNU449 and SK-Hep-1, the consequences were broad and consistent. The cells lost viability, and their ability to form colonies in culture was impaired. The cell cycle was disrupted, with cells accumulating in an arrested state rather than progressing smoothly through division. Apoptosis, the programmed self-destruction that cancer cells typically resist, increased. Beyond growth and survival, the cells’ metastatic potential also weakened: migration and invasion, the two behaviors that allow a tumor to spread beyond its original site, were attenuated. Finally, sphere formation, an in vitro assay that reflects the self-renewal capacity associated with cancer stem cells, was compromised, suggesting that USP45 supports not just bulk tumor growth but also the stem-like subpopulation that often fuels relapse and treatment resistance.

The downstream signaling changes reinforced this picture. USP45 deficiency suppressed the NF-κB signaling pathway, a major pro-survival and pro-inflammatory cascade that many cancers exploit to keep growing under hostile conditions. At the same time, loss of USP45 reduced the expression of key transcription factors involved in cancer cell stemness, the molecular program that endows a subset of tumor cells with the self-renewal and differentiation plasticity of stem cells. Taken together, these results indicate that USP45 sits upstream of at least two malignant programs in hepatocellular carcinoma cells: the inflammatory survival signaling that keeps cells alive and the stemness program that maintains a reservoir of aggressive, therapy-resistant cells.

The mechanistic core of the study lies in the physical and functional relationship between USP45 and CIAP2. Using co-immunoprecipitation, the researchers showed that USP45 interacts with CIAP2 in hepatocellular carcinoma cells, and endogenous co-immunoprecipitation experiments, which detect interactions between the proteins as they naturally occur in cells rather than in artificial overexpression systems, confirmed that this partnership is physiological. Functionally, USP45 removes ubiquitination from CIAP2 and enhances its stability, meaning the enzyme directly extends the lifespan of the oncoprotein inside the cell. The team then narrowed the mechanism further with ubiquitin mutant assays. By testing cells co-transfected with Flag-tagged CIAP2, HA-tagged USP45, and either wild-type ubiquitin or mutant ubiquitin lacking the K48 or K63 linkage sites, they demonstrated that USP45 specifically targets K48-linked polyubiquitin chains on CIAP2. In plain terms, USP45 strips off precisely the ubiquitin marks that would otherwise condemn CIAP2 to the proteasome, sparing the K63-linked chains that serve other signaling roles.

A crucial rescue experiment established that CIAP2 is not merely one target among many but the functionally relevant mediator of USP45’s oncogenic activity. When the researchers overexpressed CIAP2 in USP45-deficient hepatocellular carcinoma cells, the malignant traits that had been lost came back: the cells regained the aggressive behaviors that USP45 depletion had taken away. This epistasis-style logic, in which restoring a downstream factor rescues a phenotype caused by losing an upstream factor, is a strong indication that the USP45–CIAP2 axis is the operative pathway rather than a side effect of some other change. It also suggests that the amount of CIAP2 available to the tumor cell, which USP45 controls by editing its ubiquitin tags, is a limiting factor for the malignant state.

The final piece of evidence came from an animal model. In a hepatocellular carcinoma xenograft mouse model, in which human cancer cells are grown as tumors in immunodeficient mice, depletion of USP45 inhibited tumor growth and disrupted various malignant characteristics of the tumors. This in vivo result matters because cell culture experiments, however thorough, cannot fully capture the complexity of a growing tumor with its nutrient gradients, stromal interactions and selective pressures. The xenograft data indicate that the USP45–CIAP2 mechanism is not an artifact of the culture dish but operates in the context of an actual expanding tumor, strengthening the case that the pathway is relevant to real disease progression.

Collectively, the findings position USP45 as an oncogene in hepatocellular carcinoma and define its mechanism of action with unusual precision: USP45 binds CIAP2, removes K48-linked polyubiquitin chains from it, stabilizes the protein, and thereby sustains the survival, proliferation, motility and stem-like behavior of liver cancer cells, with the axis confirmed both in cultured cells and in xenograft tumors. The study was approved by the Ethics Committee of The Affiliated Hospital of Putian University and was supported by the Fujian Provincial Natural Science Foundation and the Joint Funds for the Innovation of Science and Technology of Fujian Province. For the field, the work adds USP45 to the growing list of deubiquitinating enzymes implicated in cancer and nominates the USP45–CIAP2 pair as a potential therapeutic axis. Because the enzyme–substrate relationship is defined at the level of a specific ubiquitin linkage, it offers a concrete biochemical handle for future drug discovery efforts aimed at destabilizing CIAP2 in liver tumors, although translating such mechanistic insight into clinical interventions will require the usual long road of validation and development.

Subject of Research: The role of the deubiquitinating enzyme USP45 in stabilizing CIAP2 and promoting hepatocellular carcinoma progression

Article Title: USP45 Promotes hepatocellular carcinoma progression by regulating the ubiquitination and stability of CIAP2

Article References: Jiang, X., Chen, J., Lin, W., Huang, K., Wang, L., & Lin, J. (2026). USP45 Promotes hepatocellular carcinoma progression by regulating the ubiquitination and stability of CIAP2. Cellular and Molecular Life Sciences. https://doi.org/10.1007/s00018-026-06409-5

Image Credits: AI Generated

DOI: 10.1007/s00018-026-06409-5

Keywords: hepatocellular carcinoma, USP45, CIAP2, deubiquitinating enzymes, ubiquitination, K48-linked polyubiquitin, proteasomal degradation, NF-kB signaling, cancer stem cells, xenograft mouse model, liver cancer, oncoprotein stability