Molecular Mechanisms Of Spt6 Involved In Colon Cancer Growth Through Regulating HTERT Expression

Objective:hTERT(human telomerase reverse transcriptase)is a ribonucleoprotein polymerase.To prevent the ends of chromosomes become shorter,maintaining chromosomal stability,chromatin repair to ensure the potential of chromosomal replication in normal human cells plays an important role.It plays an important role in cell senescence.It was lowly expressed in normal somatic cells.The abnormal expression of hTERT in normal somatic cells is often closely related to tumor formation.Numerous studies have shown that,hTERT had a higher expression in lung cancer,esophageal cancer,stomach cancer,breast cancer,colon cancer,and hepatocellular carcinoma.It has shown that SP1,Akt and c-Myc,STAT3 were involved in the regulation of transcriptional and posttranscriptional gene expression of hTERT and regulating the development of prostate cancer.hTERT mRNA levels in patients with nasopharyngeal carcinoma can be a early diagnosis and therapeutic target,by means of radiotherapy and chemotherapy after reducing hTERT mRNA,survival status of patients with nasopharyngeal carcinoma have also improved,indicating that hTERT is an important factor in the development of nasopharyngeal carcinoma.Studies have found that on hTERT expression in tumor tissues or cells,hEST2 are also high.Therefore,we think that in cell immortalization and tumorigenesis,hEST2 may be the activator of hTERT.Research has also found that long-term high level expression of cell cycle regulatory factors MCRS2,will lead to the cell received inhibition of hTERT activity and telomere length becoming shorter,indicating that hTERT processes may be involved in cell cycle regulation.Studies also show that PI3K/Akt pathways can be the upstream of hTERT.There is a positive feedback pathway inside the cell.In turn,hTERT can adjust the PI3K/Akt signaling pathway.The PI3K/Akt signaling pathway is related to cell proliferation.It has shown to be activated in tumor cells.There are reports,compared with hTERT expression in normal cells,hTERT gene promoter activity showed high levels in cancer cells with hTERT expression activated.Because of the complexity of gene regulation and cancer cause diversity and uncertainty,there are other regulators of hTERT.This study was to discover new hTERT transcription factors,and identify its role in the transcriptional regulation of hTERT mechanism as well as the significance of cancer research.Methods:(1)In colon cancer cell lines,we used the new Biotin-Streptavidin promoter fishing system to separate and identify hTERT promoter binding proteins.(2)The effect of siRNA-specific inhibition of the hTERT promoter binding protein on hTERT gene expression,cell growth,proliferation,migration and apoptosis were tested.(3)We established animal model of mice to verify the effect of hTERT promoter binding protein on tumor growth and hTERT gene expression.(4)The expression of hTERT promoter binding protein was detected by Western blot,RT-PCR,experimental techniques in cell lines tumor tissues.We further analyzed the clinical data to determine the relationship between hTERT promoter binding protein and diagnosis,prognosis,transfer and recurrence.Results:(1)We used Streptavidin-agarose pulldown and proteomics assay and identified Spt6 as a hTERT promoter-binding protein.(2)Knockdown of Spt6 inhibited telomerase activity,hTERT promoter activity,cancer cell proliferation and migration,and tumor cell apoptosis.(3)In vivo experiments showed that knockdown Spt6 inhibited tumor growth and hTERT gene expression.(4)Immunohistochemistry and immunoblot experiments verified the correlation of Spt6 with hTERT in tumor tissues and cells.The expression of Spt6 is implicated in cancer diagnosis and prognostic significance.Conclusions:We identified Spt6 as an hTERT promoter binding protein and verified the role of Spt6 in the regulation of hTERT expression and tumor growth.