Gene transcriptomics reveals that PAK5 regulates the pathogenesis of Esophageal Squamous Cell Carcinoma
Abstract Background Esophageal Squamous Cell Carcinoma (ESCC) is one of the leading causes of cancer death worldwide, especially in Asia. Because the early symptoms are not obvious, most patients are already in an advanced stage when diagnosed, which greatly limits the effectiveness of treatment and affects the patient's prognosis. Therefore, it is of great significance to explore the occurrence and development mechanism of ESCC and to search for effective biomarkers and therapeutic targets for improving patient prognosis. P21 activated protein kinase 5 (PAK5) belongs to the mitogen-activated protein kinase family, which has been extensively studied in recent years and is thought to play an important role in a variety of malignant tumors. PAK5 promotes tumor development by regulating processes such as cell cycle, cell migration and invasion. However, the specific role of PAK5 in ESCC and its potential mechanisms have not been fully elucidated. Methods We initially employed immunohistochemical techniques to assess PAK5 expression levels in ESCC tissue samples and compared these levels with those in normal esophageal tissues. Kaplan-Meier survival analysis was subsequently conducted to evaluate the correlation between PAK5 expression levels and patient survival in ESCC. To further investigate the functional role of PAK5 in ESCC cells, we constructed PAK5 overexpression (PAK5-OE) and PAK5 silencing (PAK5-si) ESCC cell models.  The effects of PAK5 on cell proliferation, clonogenicity, migration, and invasion were assessed using CCK-8 assays, Cell Plate Cloning assays, and Transwell assays. Additionally, transcriptomic analyses were performed using mRNA sequencing techniques to identify downstream effector molecules and signaling pathways influenced by PAK5. Results The results showed that the expression of PAK5 in ESCC tissue was significantly higher than that in normal esophageal tissue, and high PAK5 level was closely associated with poor prognosis in ESCC patients.  Functional experiments showed that PAK5 could significantly enhance the proliferation, clonal formation, migration and invasion of ESCC cells.  Further transcriptomic analysis revealed multiple downstream genes and signaling pathways regulated by PAK5, particularly those related to cell cycle progression and anti-apoptosis. Most notably, this study found that PAK5 forms a new signaling axis by interacting with GAREM1, which is critical for maintaining the transfer behavior of ESCC cells.These findings underscore PAK5's tumorigenic potential in ESCC, positioning it as a promising prognostic marker and therapeutic target. Keywords: PAK5, Esophageal squamous cell carcinoma, GAREM1 Overall design: The KYSE-150 cell line, modified with either PAK5 overexpression or PAK5 silencing, was used for mRNA sequencing to identify differentially expressed genes (DEGs). Sequencing was performed by Beijing Novozymes Biotechnology Co., Ltd. using standard protocols. DEGs between PAK5-si-KYSE-150 and WT-KYSE-150 cells were analyzed using the DESeq2 R package, with significance thresholds set at FDR < 0.05 and |log2(Fold Change)| = 1.