Dataset Information

Circ_0000045 promotes proliferation, migration, and invasion of head and neck squamous cell carcinomas via regulating HSP70 and MAPK pathway.

ABSTRACT:

Objective

Head and neck squamous cell carcinoma (HNSCC) is one severe malignancy driven by complex cellular and signaling mechanisms. However, the roles of circular RNAs (circRNAs) in HNSCC's development remains poorly understood. Therefore, this study investigated the functions of differentially expressed circRNAs in regulating HNSCC cell functions.

Methods

Differentially expressed circRNAs were characterized through RNA sequencing in HNSCC tissues. CircRNA's identity was then confirmed using RT-PCR and Sanger's sequencing. Next, expression levels of circRNA and mRNA were detected by qRT-PCR, after which protein abundances were measured by Western blotting. Subsequently, the proliferation, migration, and invasion of HNSCC cells was assessed by MTS, wound healing, and Transwell system, respectively, followed by identification of circRNA-binding proteins in HNSCC cells by circRNA pull-down, coupled with mass spectrometry.

Results

Great alterations in circRNA profiles were detected in HNSCC tissues, including the elevated expression of circ_0000045. As observed, silencing of circ_0000045 effectively repressed the proliferation, migration, and invasion of HNSCC cell lines (FaDu and SCC-9). Contrarily, circ_0000045's overexpression promoted the proliferation, migration, and invasion in FaDu and SCC-9 cells. Results also showed that circ_0000045 was associated with multiple RNA-binding proteins in HNSCC cells, such as HSP70. Moreover, circ_0000045 knockdown enhanced HSP70 expression and inhibited JNK2 and P38's expression in HNSCC cells, which were oppositely regulated by circ_0000045's overexpression.

Conclusion

The high expression of circ_0000045; therefore, promoted cell proliferation, migration, and invasion during HNSCC's development through regulating HSP70 protein and mitogen-activated protein kinase signaling.

SUBMITTER: Sun R

PROVIDER: S-EPMC9297571 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:While treatment failure in cases of head and neck squamous cell carcinoma (HNSCC) frequently takes the form of locoregional recurrences and distant metastasis, our understanding of the mechanisms of metastasis in HNSCC is limited. We initially performed the upstream and key nodes analysis together with whole gene microarray analysis characterized by distant metastatic potential in vivo with HNSCC cell lines and identified JunB, a member of the activator protein-1 (AP-1) family, as a key molecule in the regulation of the pathways related to distant metastasis in HNSCC. We have therefore tested the hypothesis that JunB plays a crucial role in distant metastasis in HNSCC.To study the role of JunB on metastatic potential of HNSCC, small interfering RNA (siRNA)-mediated knockdown and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (cas9) system (CRISPR/Cas9)-mediated knockout of JunB in HNSCC cells were established and the abilities of cell invasion and migration in vitro were examined. The efficacy of knockout of JunB was also examined using an experimental lung metastatic mouse model of HNSCC. In addition, to study if the role of JunB in HNSCC cell migration and invasiveness is related to epithelial-to-mesenchymal transition (EMT), cell morphology and expression of mesenchymal or epithelial marker on siRNA mediated JunB knockdown in HNSCC cells were examined with or without TGF-? stimulation.siRNA knockdown and sgRNA knockout of JunB in metastatic HNSCC cells significantly suppressed both cell invasion and migration in vitro. In addition, the knockout of JunB in metastatic HNSCC cells significantly repressed the incidence of lung metastases and prolonged the survival in vivo. However, we did not observe any change in cell morphology with the down-regulation of mesenchymal markers and up-regulation of epithelial markers in response to siRNA-mediated JunB knockdown in HNSCC cells.These results suggested that JunB could play an important role in promoting cell invasion, migration and distant metastasis in HNSCC via pathways other than EMT and that the down-regulation of JunB may become an effective strategy for patients with invasive HNSCC.