The Function And Mechanism Of Yb1 In Sunitinib Tolerance And Metastasis Of Renal Cell Carcinoma

The incidence of renal cell carcinoma(RCC)is increasing and one third of newly diagnosed cases yet are metastatic.The metastatic spread of solid malignant tumors results in RCC uncurable by surgical operation and therefore more difficult to treat.New molecular-targeted therapies have played a key role in RCC treatment.Unfortunately,tumors frequently evolve resistance into these targeted therapies letting cancer cells to continue to survive.YB1(Y-Box-binding protein 1)is a transcription/translation regulatory factor which is a member of cold-shock domain protein superfamily.It was studied to regulate miscellaneous gene expressions through binding to the Y-box regulatory element of target gene promoters.Furthermore,YB1 was discovered to form complexes with RNA and played many roles in RNA stabilization and messenger RNA(m RNA)splicing.Clinical research have shown the raised nuclear YB1 expression in large-scale of human carcinomas,including prostate,breast,lung,colon,and bladder.Additionally,overexpression of YB1 was closely correlated with the tumor growth and invasive cancer phenotype,resulting in poor prognosis of cancer patients.Mechanistically,YB1 raised cell proliferation through upregulating the expressions of EGFR(epidermal growth factor receptor)in breast cancers.YB1 has also been proved to regulate the inherent MDR1(multidrug resistance gene 1)expression,hinting its contribution to drug resistance.Anyway,the underlying molecular mechanism of the association of increased YB1 expression with renal cell carcinoma remain abstruse.This study is divided into three units:Part ? The expression of YB1 in cc RCC and the effects of YB1 on the biological behavior of cc RCC cellsObjectives: To analyze the expression level of YB1 in cc RCC and investigate the effect of YB1 on the biological behavior of cc RCC cells.Methods: Doing with the bioinformatics analysis of the TCGA database,we detect the expression level of YB1 in cc RCC.The relationship between the expression of YB1 and clinical characteristics of cc RCC patients was further analyzed.Through analyzing immunohistochemical staining and western blotting of cc RCC tissue samples,we confirmed the TCGA database information.Transwell assay was used to investigate the effect of knockdown or overexpression of YB1 on the migration and invasion of renal cell lines.Results: Compared to adjacent tissues,YB1 was highly expressed in cc RCC tissues.The high expression of YB1 correlates positively with the high stage of tumors.After knocking down YB1,the migration and invasion rates of 786-O and ACHN cells decreased significantly.After overexpression of YB1,the migration and invasion rates of 786-O and ACHN cells increased significantly.Conclusion: YB1 was highly expressed in cc RCC tissues compared to adjacent tissues and its expression level is closely related to the progression of renal cell carcinoma.Part ? The mechanisms of YB1 affecting the biological behaviors of cc RCC cellsObjectives: To clarify the mechanisms of YB1 affecting migration and invasion of cc RCCMethods: Using bioinformatic analysis and RNA-sequencing to explore the YB1-involved gene set and signal pathway molecules.Chromatin immunoprecipitation and proteolysis was used to detect the way of YB1 regulate the downstream target molecule.A series of rescue experiments were conducted to research the mechanisms of YB1 affecting the biological behaviors of cc RCC cells.Results: High YB1 expression largely enriched in the metastasis and drug resistance pathways.After knockdown or overexpression of YB1,the expression level of Eph A2 was down-regulated or up-regulated,respectively.Protein degradation experiments and rescue experiments showed that YB1 binds to Eph A2,inhibiting the ubiquitination and maintain the protein stability of Eph A2,thereby promoting migration and invasion of cc RCC cells.Conclusion: YB1 regulates the expression level of Eph A2 at the translation and affecting the biological behaviors of cc RCC cells consequently.Part ? The effects of YB1 on sunitinib tolerance and metastasis of renal cell carcinomaObjectives: To determine the effects of YB1 on sunitinib tolerance and metastasis of renal carcinoma by up-regulate the expression of Eph A2.Methods: In vitro,CCK-8 Proliferation experiments was used to determine whether sunitinib sensitivity of RCC cells was changed when YB1 and/or Eph A2 was knocked down or overexpressed.In vivo,nude mice metastasis model was established to value the effects of YB1 on metastasis of renal carcinoma.Results: YB1 promote sunitinib resistance and metastasis of renal carcinoma by up-regulate the expression of Eph A2.Conclusion: YB1 promoting sunitinib tolerance and metastasis in renal cell carcinoma by up-regulating Eph A2.