The Role And Mechanism Of MiR-487b Targeting Negative Regulation Of KAT6B In The Progression Of Cervical Cancer

Background and ObjectivesCervical cancer is the most common gynecological cancer.In recent years,its incidence tends to be younger.At present,it is known that the loss or gain of function caused by gene change is related to cancer progression,such as gene mutation,deletion,copy number distortion and chromosome weight row.MicroRNA is a single chain non encoding RNA,which plays a role in regulating some key genes after transcription.By activating or inhibiting mRNA,multiple target genes can be regulated,and cell proliferation,invasion,apoptosis,migration and differentiation can be changed.Lysine acetyltransferases(KATS)can acetylate histones and other non histone matrix,which is very important for the structure and function of chromatin.Chromosome translocation of kat6b(also known as MORF and QKF)oncogene has also been found in many cancers.Previous studies have shown that the maladjustment of mir-487 b in colorectal cancer,lung cancer,prostate cancer,gallbladder cancer,glioma and other cancers can promote the growth and proliferation of tumors.The inhibition of kat6 b can induce cell cycle withdrawal,cell aging and cell proliferation in epigenetics.In this project,we intend to explore the role of mir-487 b in cervical cancer cells and the relationship between mir-487 b and kat6 b by cell biology and molecular biology.Research methods and materialsIn order to study the effect of miR-487 b on the proliferation,invasion,migration and invasion of cervical cancer cell line HeLa and c33 a,the antisense sequence of miR-487b(lv-antigomir-487b)was first transfected into cervical cancer cell line,and miR-487 b was detected by fluorescence and PCR.Then,the target gene of miR-487 b,KAT6B,was predicted and regulated by Internet free database(including targetscan,miRBase,etc.).Finally,using CRISPR / cas9-ko method to knock out KAT6 B,on thisbasis,the antisense sequence of miR-487 b precursor or precursor sequence was transferred to study the effect of up-regulation of miR-487 b or down-regulation of miR-487 b on the proliferation,migration,invasion and invasion of cervical cancer cells.In the experiment,cells were continuously cultured and counted under cell microscope to detect cell growth;cell clone formation experiment was used to detect cell proliferation ability;EdU experiment was used to detect cell DNA replication and proliferation ability;scratch experiment was used to detect cell migration movement ability and repair ability;Transwell experiment and Matrigel Transwell experiment were used detect cell migration and invasion ability;flow cytometry was used to detect cell cycle change;Western blot was used to detect protein expression and explore possible signal mechanism.Statistical analysisEach experiment of each cell line was repeated for more than 3 times,and each experiment was set with 3 subsidiary pores.SPSS 22.0 software was used for statistics,and graphpad prism 7.0 software was used for image data sorting.When p <0.05,there was statistical significance.Results1.MiR-487 b has a negative effect on the growth of cervical cancer cells.In cervical cancer cell lines,knockdown of miR-487 b expression increased cell growth rate,colony forming ability,DNA replication and proliferation ability,migration and repair ability,migration and invasion ability.2.KAT6 B is the target gene of miR-487 b.Combined with the free database(including TARGETSCAN,miRbase,etc.),it is predicted that the target of miR-487 b is KAT6 B,which has the potential binding site of miR-487 b.3.After the KAT6 B gene was knocked out,the antisense sequence of miR-487 b precursor or precursor sequence was transferred.WB experiment showed that the expression of KAT6 B had no significant change.In the experiments of cell proliferation,migration and invasion,it was found that the trend of the results was the same as that of KAT6 B knockout,and there was no significant decrease of miR-487 b expression to promote cell growth and proliferation.4.MiR-487 b may regulate the proliferation,migration and invasion of cervicalcancer through AMPK-AKT-mTOR signaling pathway.Western blot showed that the phosphorylation level of AMPK? decreased with the knockdown of miR-487 b,while the phosphorylation level of mTOR,AKT and S6 increased,and the expression of growth factor receptor EGFR,PDGFR? and PDGFR? increased.In contrast to the above results,the phosphorylated AMPK? level increased,the phosphorylated levels of mTOR,AKT and S6 decreased,and the expression of growth factor receptor EGFR,PDGFR? and PDGFR? decreased in the knockout of KAT6 B cells and the up-regulated or down-regulated of miR-487 b cells.Conclusions1.The low expression of miR-487 b promotes the growth,proliferation,migration and invasion of cervical cancer cells.2.The target of miR-487 b is KAT6 B,which can regulate cervical cancer by targeting and combining KAT6 B.3.MiR-487 b regulates the proliferation,migration and invasion of cervical cancer cells by activating AMPK-AKT-MTOR signaling pathway...