Study On The Role Of TRIM25 In The Progression Of Papillary Thyroid Carcinoma

Background Papillary Thyroid Carcinoma(PTC)is the most common type of thyroid carcinoma(TC),its incidence is increasing in the past years.Studies have shown that the reduced expression of sodium/iodide symporter(NIS)is one of the factors that can lead to dedifferentiation in PTC.TRIM25,an E3 ubiquitin ligase,implicated in the regulation of metabolic reprogramming,cell proliferation,migration and apoptosis in multiple tumors.It has been found to be abnormally low expressed in thyroid cancer,and bioinformatics analysis has shown that it is positively correlated with NIS.However,the exact role of TRIM25 in PTC is not yet clear.Pituitary TumorTransforming Gene 1 Protein-Interacting Protein(PBF)is an oncogenic protein that has been found to inhibit the functional expression of NIS in thyroid cancer.The Protein Interaction Information Database(Biogrid)suggests that TRIM25 interacts with PBF.Accordingly,we hypothesized that TRIM25 may affect the expression of NIS through regulating PBF,which in turn affects the dedifferentiation process of PTC.Objective To study whether TRIM25 affects the expression of NIS through regulating PBF in papillary thyroid carcinoma,and thereby affects the progression of PTC.It may provide a new target for clinical treatment.Materials and methods1.Bioinformatics analysis of the expression of TRIM25 in thyroid carcinoma and its relationship with clinicopathological features.The data of gene expression and clinicopathological features of thyroid carcinoma were obtained from the TCGA and GTEx database.The difference of TRIM25 expression between normal thyroid and thyroid carcinoma was analyzed by the MannWhitney U test.After screening the data without clinical information,the expression of TRIM25 in different clinicopathological features were statistically analyzed by the Mann-Whitney U test,such as T-stage,N-stage,extrathyroidal extension,and pathological subtypes.The Kaplan-Meier Plotter tool was used to analyze the relationship between the TRIM25 and the prognosis of thyroid carcinoma2.Verifying the expression of TRIM25 in normal thyroid and PTC through experimentsSix patients who were pathologically diagnosed with PTC were randomly selected,and specimens of PTC tissues and normal thyroid tissues were collected.The expression of TRIM25 was verified by Western Blot and immunohistochemistry.The expression of TRIM25 at the m RNA and protein levels in the normal thyroid cells Nthy-ori 3-1,papillary thyroid cancer cells K1 and TPC-1 were detected by PCR and Western Blot.3.Effects of overexpression of TRIM25 on the proliferation and migration in PTC cells in vitroK1 and TPC-1 cells were randomly divided into 2 groups,vector group(vector)and overexpression group(TRIM25).The effect of over-expressing TRIM25 on the proliferation of PTC cells was detected by CCK8 and colony formation assay;the effect of over-expressing TRIM25 on the migration of PTC cells was detected by Transwell and wound healing.4.Effect of knockdown of TRIM25 on the proliferation and migration of PTC cells in vitroK1 and TPC-1 cells were randomly divided into three groups,negative control group(si TRIM25-NC),TRIM25-interfering group 1(si TRIM25-2),and TRIM25-interfering group 2(si TRIM25-3).The effect of the knockdown of TRIM25 on the proliferation of PTC cells was detected by CCK8 and colony formation;the effect of knockdown of TRIM25 on the migration of PTC cells was detected through transwell and wound healing.5.The relationship between TRIM25 and NIS,TgOverexpression/knockdown of TRIM25 in K1 and TPC-1 cells,using Western Blot assay to verify the relationship between overexpression/knockdown of TRIM25 and NIS,Tg protein expression in PTC cells.6.Study on the regulatory effect of TRIM25 and PBF(1)Overexpression/knockdown of TRIM25 in K1 and TPC-1 cells,and detect the expression of PBF using Western Blot.(2)The interaction between TRIM25 and PBF protein was detected using CO-IP assay.Results1.Comparing with normal thyroid,the expression of TRIM25 was significantly decreased in PTC tissues and cells.There is a correlation between the expression of TRIM25 and various factors in thyroid carcinoma such as T-stage,N-stage,extrathyroidal extension,pathological subtype,and disease-free survival.2.Overexpression/knockdown of TRIM25 could inhibit/promote PTC proliferation and migration in PTC cells.3.Overexpression/knockdown of TRIM25 could promote/inhibit the expression of NIS and Tg in PTC cells.4.TRIM25 interacted with PBF and overexpression/knockdown of TRIM25 could inhibit/promote the expression of PBF in PTC cells.Conclusion In papillary thyroid carcinoma,TRIM25 may affect the expression of NIS by inhibiting PBF,and affect the proliferation and migration of PTC cells,which may be related to the de-differentiation process of papillary thyroid carcinoma.