This study reveals a critical role of PKCη in triple-negative breast cancer (TNBC) metastasis, elucidating that PKCη activates the Hippo-YAP pathway by phosphorylating YAP at Ser128 and suppresses AKT signaling, thereby promoting EMT and tumor stemness. Additionally, it identifies that a micropeptide encoded by an evolutionarily conserved upstream open reading frame (uORF) can target PKCη for degradation, offering potential therapeutic value.
Literature Overview
The article titled 'PKC-eta promotes breast cancer metastasis by regulating the Hippo–YAP signaling pathway,' published in Signal Transduction and Targeted Therapy, reviews and summarizes the molecular mechanisms of protein kinase C-eta (PKCη) in the metastasis of triple-negative breast cancer (TNBC). The study reveals that PKCη promotes epithelial-mesenchymal transition (EMT) and tumor stemness by regulating the Hippo-YAP signaling pathway, thereby driving tumor progression and distant metastasis. It also identifies that a micropeptide encoded by an evolutionarily conserved upstream open reading frame (uORF) can act as a PKCη degrader, activating the Hippo pathway and suppressing YAP function. The entire passage is coherent and logical, ending with a Chinese period.Background Knowledge
Triple-negative breast cancer (TNBC) is an aggressive and highly metastatic subtype of breast cancer, accounting for 10%–20% of all breast cancer cases, with limited treatment options. The absence of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression renders targeted therapies ineffective, leaving chemotherapy as the primary treatment, which is often hindered by high rates of drug resistance and recurrence. Therefore, identifying key signaling pathways driving TNBC metastasis is crucial for developing novel therapeutic strategies.
Epithelial–mesenchymal transition (EMT) is a vital biological process enabling tumor cells to acquire migratory and invasive capabilities, closely associated with the tumor stem cell (CSC) phenotype, promoting tumor initiation, metastatic colonization, and therapy resistance. The Hippo-YAP signaling pathway, an evolutionarily conserved pathway, regulates organ size, tissue homeostasis, and cancer development. When inactive, YAP/TAZ translocate into the nucleus and bind to TEAD transcription factors, activating genes that promote proliferation and metastasis. In various cancers, hyperactivation of YAP is linked to poor prognosis.
The protein kinase C (PKC) family serves as critical downstream effectors of G protein–coupled receptors (GPCRs), regulating diverse cellular functions. Among them, PKCη belongs to the novel PKC subfamily and exhibits anti-apoptotic functions, with aberrant expression observed in multiple cancers. Although previous studies have linked PKCη to poor prognosis in breast cancer, its specific role in TNBC metastasis remains unclear. Furthermore, upstream open reading frames (uORFs), typically located in the 5'UTR of mRNAs, were traditionally considered to inhibit translation of the main ORF. However, recent studies have shown that certain uORFs can encode functional micropeptides involved in regulating protein stability and signaling pathways.
This study focuses on the role of PKCη in TNBC metastasis, systematically dissecting its regulatory mechanisms on the Hippo-YAP pathway and exploring the therapeutic potential of uORF-encoded micropeptides, providing novel molecular targets and intervention strategies for TNBC. This work fills a gap in understanding PKCη's regulation of the Hippo pathway and expands our knowledge of the kinase–transcriptional co-activator interaction network, holding significant theoretical and translational implications.
Research Methods and Experiments
The researchers first analyzed the expression profile of PRKCH (encoding PKCη) across different breast cancer subtypes using public databases such as TCGA and METABRIC, and assessed its correlation with EMT features via GSEA enrichment analysis. Immunohistochemistry (IHC) was performed on tissue microarrays to validate the relationship between PKCη expression and tumor grade, stage, and subcellular localization. Functionally, CRISPR/Cas9 technology was used to generate PKCη-knockout 4T1 and MDA-MB-231 cell lines, followed by in vitro migration, invasion, sphere formation, and limiting dilution assays to evaluate the impact on EMT and stemness. In vivo, orthotopic transplantation and tail vein injection were used to establish xenograft mouse models to monitor tumor growth and lung metastasis, along with survival analysis.
To investigate the molecular mechanism of PKCη, the authors validated the physical interaction between PKCη and YAP using Co-IP, ELISA, and molecular docking analyses, and confirmed direct phosphorylation of YAP at Ser128 by PKCη using in vitro kinase assays. Subcellular fractionation and immunofluorescence were used to analyze changes in YAP localization, and cycloheximide chase experiments assessed YAP protein stability. Additionally, the impact of PKCη loss on the AKT-MST1-LATS1 pathway was examined, with signal crosstalk verified using siRNA knockdown and pharmacological inhibitors. Finally, the effect of the uORF-encoded micropeptide uPEP2 on PKCη and YAP expression was studied, and its anti-tumor and anti-metastatic effects were evaluated in both cellular and animal models.Key Conclusions and Perspectives
Research Significance and Prospects
This study is the first to systematically reveal PKCη as an upstream negative regulator of the Hippo-YAP pathway, playing a central role in TNBC metastasis. By phosphorylating YAP at Ser128, PKCη not only enhances YAP stability but may also impede LATS1-mediated phosphorylation at Ser127, synergistically promoting YAP nuclear localization and transcriptional activity. This finding expands our understanding of the regulatory network of the Hippo pathway, particularly emphasizing functional specificity differences among PKC family members.
Moreover, the study identifies that the micropeptide uPEP2 can activate the Hippo pathway by degrading PKCη, significantly suppressing tumor progression and metastasis, offering a novel targeted strategy for TNBC treatment. Compared to traditional small-molecule kinase inhibitors, targeted protein degradation (e.g., PROTAC or micropeptide-mediated degradation) may offer greater selectivity and sustained effects. Future studies could further explore delivery methods, pharmacokinetics of uPEP2, and its potential for combination with other therapies such as immunotherapy.
Although the mechanistic analysis in this study is thorough, further validation of PKCη's universality in other TNBC models is needed, along with exploration of its role in the tumor microenvironment. Additionally, the stability and in vivo bioavailability of uPEP2 require optimization to advance its clinical translation. Overall, this work provides a robust new target and proof-of-concept for precision therapy in TNBC.
Conclusion
This study systematically elucidates the critical oncogenic role of PKCη in triple-negative breast cancer metastasis and its underlying molecular mechanisms. By integrating multi-omics analyses, gene-edited cell models, and animal experiments, the study demonstrates that PKCη is highly expressed in the ‘claudin-low’ subtype and significantly correlates with EMT features and poor prognosis. Mechanistically, PKCη activates YAP through a dual pathway: directly phosphorylating YAP at Ser128 to enhance its stability and nuclear translocation, and inhibiting the AKT-MST1-LATS1 kinase cascade to relieve Hippo pathway suppression of YAP. Functional experiments show that loss of PKCη significantly impairs tumor cell migration, invasion, and stemness, and suppresses primary tumor growth and lung metastasis in mouse models. More importantly, the study reveals that an evolutionarily conserved uORF-encoded micropeptide, uPEP2, can selectively degrade PKCη, thereby activating the Hippo pathway, promoting YAP degradation, and inhibiting metastasis. These findings not only expand our understanding of crosstalk between the PKC family and the Hippo-YAP signaling pathway but also provide new intervention targets and potential therapeutic molecules for triple-negative breast cancer, offering significant translational medical value.
