Function And Associated Molecular Mechanisms Of Mitochondrial Ribosomal Gene MRPL35 In Esophageal Carcinoma

Background: Esophageal cancer is a common cancer type worldwide,ranking seventh in incidence among all malignant tumors and sixth in overall mortality.The incidence of esophageal cancer in China has been increasing and is currently the highest in the world.Despite advancements in diagnosis and treatment,the prognosis of esophageal cancer remains poor,with the overall five-year survival rate below 20% globally and in China.Clinical evidence suggests that if the tumor is clinically staged early at the time of diagnosis,the five-year survival rate can increase to 80%-90%.However,due to the lack of specific clinical manifestations in early-stage esophageal cancer,patients often seek medical attention during the advanced stages.The use of biomarker-based non-endoscopic disease detection has improved the accuracy and availability of early esophageal cancer screening.Therefore,establishing molecular targets for population-wide early screening and high-risk population warning can enhance the early diagnosis rate of esophageal cancer and explore new molecular treatment strategies,which holds significant clinical significance.Apoptosis inhibition and reprogramming of energy metabolism are important characteristic features of cancer.Mitochondria play a crucial role in cellular apoptosis and energy metabolism and have a relatively independent gene expression mechanism.Proteins associated with mitochondrial oxidative phosphorylation are located within the inner membrane of mitochondria and are translated and synthesized by mitochondrial ribosomes.Mitochondrial ribosomes consist of three RNA components encoded by mitochondrial DNA and 82 specific protein components encoded by nuclear DNA.They differ in complexity,structure,and mechanisms of action compared to cytoplasmic ribosomes.In the past few decades,the structure and basic functions of mitochondrial ribosomes have been elucidated.In recent years,with the rapid development of high-throughput sequencing and bioinformatics,the connection between mitochondrial ribosomes and tumors has gradually been unveiled.Relevant studies have shown that some mitochondrial ribosomal genes are upregulated in malignant tumors and are closely associated with tumor occurrence and development.The mitochondrial ribosomal large subunit protein coding gene MRPL35(Mitochondrial ribosomal protein L35)is essential for mitochondrial oxidative phosphorylation.Recent research suggests that MRPL35 is upregulated in gastric and colon cancer,and reducing its expression in in vitro experiments leads to DNA damage,cell cycle arrest,and autophagy in tumor cells.This indicates that MRPL35 has the potential to become a new molecular target for the diagnosis and treatment of malignant tumors.However,in the field of esophageal cancer,there is still a lack of transcriptomic analysis,gene function,and molecular mechanism research on MRPL35.Therefore,it is worth studying whether the gene MRPL35 has potential as a biomarker for earlystage esophageal cancer.Objective:In this study,we focused on the mitochondrial ribosomal large subunit protein coding gene MRPL35 as our research subject.Firstly,we analyzed the expression of MRPL35 and its clinical relevance in esophageal cancer by combining tumor-related databases and clinical samples.This aimed to determine the potential of MRPL35 as a biomarker for early-stage esophageal cancer and its subsequent research value.Next,we used RNA interference to suppress the expression of the target gene MRPL35 in esophageal squamous cell carcinoma cell lines(KYSE410 and TE-1)and investigated its gene function in both in vivo and in vitro settings.Finally,we integrated cell functional assays,molecular biology techniques,in vivo experiments using nude mice,high-throughput sequencing,and bioinformatics analysis to identify the signaling pathways and key molecules involved.We aim to explore the role of MRPL35 in the development of esophageal cancer and provide new molecular strategies and theoretical foundations for the selection of biomarkers and treatment targets in early-stage esophageal cancer.Methods:In this study,we utilized cancer-related databases to analyze the gene expression of MRPL35 and its clinical relevance in esophageal squamous cell carcinoma.We further validated these findings using clinical samples.To target MRPL35 gene expression,we designed RNA interference targeting sequences based on the MRPL35 gene and prepared viral vectors capable of downregulating the target gene.We infected cells with the viral vectors to reduce the expression of the target gene and screened and cultured esophageal cancer cell lines with stable low MRPL35 expression.Using techniques such as flow cytometry and transmission electron microscopy,we demonstrated that MRPL35 knockdown induces changes in cellular activities such as apoptosis and abnormalities in mitochondrial structure and function.We performed high-throughput sequencing and transcriptomic analysis to detect the changes in gene expression throughout the whole genome after MRPL35 gene knockdown.After screening for differentially expressed downstream genes,we conducted gene functional annotation and enrichment analysis.To further validate our findings,we performed in vivo and in vitro experiments and conducted in-depth research into the signaling pathways and molecular mechanisms involved.By using these approaches,we aim to gain a comprehensive understanding of the role of MRPL35 in esophageal cancer development and explore its potential as a molecular target for early diagnosis and treatment strategies.Results:1.MRPL35 is upregulated in esophageal squamous cell carcinoma and correlates with clinical features such as tumor staging.2.In vitro studies demonstrate that reducing MRPL35 expression leads to changes in mitochondrial structure and function,including alterations in mitochondrial membrane potential and increased reactive oxygen species,ultimately resulting in cellular apoptosis.3.In vivo studies show that reducing MRPL35 expression can attenuate the formation of esophageal cancer xenografts.4.Consistent findings from both in vivo and in vitro studies suggest that reducing MRPL35 expression can activate the MAPK signaling pathway,regulating apoptosis through the JNK/p53 signaling axis.Conclusions:This study suggests that the MRPL35 gene is upregulated in esophageal squamous cell carcinoma.Lowering the expression of MRPL35 leads to changes in mitochondrial structure and function and induces apoptosis through the activation of the MAPK signaling pathway.Additionally,it can attenuate the formation of esophageal cancer xenografts.MRPL35 shows potential as a biomarker for esophageal squamous cell carcinoma and a novel therapeutic target.