| Background Glioblastoma(GBM)is the most common primary brain tumor and among the most lethal forms of human cancer,Chromosomal translocations generate fusion genes and produce oncogenic proteins.Many fusion genes have been identified and studied in hematological malignancies and solid tumors.Transcriptome sequencing is a powerful tool for detecting gene fusions in tumors.Increasing numbers of recurrent fusion genes have been identified using next-generation sequencing technologies.FGFR3-TACC3 fusion was the first reported in glioblastoma multiforme(GBM)cases using high-throughput transcriptome sequencing.The FGFR3-TACC3 fusion protein induces constitutive kinase activation and mitotic and chromosomal segregation defects.EGFR-SEPT14 was identified as the most frequent recurrent fusion transcript to activate STAT3 signaling.Several fusion genes have recently been detected by transcriptome sequencing of glioblastoma multiforme(GBM)cases in the Chinese Glioma Genome Atlas(CGGA).PTPRZ1-MET fusion transcripts wereidentified and revealed in secondary GBMs(s GBMs)cases from CGGA.A new PTEN-COL17A1 gene fusion was also detected in two GBM cases,and a new EGFR-RP11-745C15.2 gene fusion was also detected in four GBM cases from CGGA,their properties and functions remain to be characterized.Methods In the present study,we used RT-PCR together with Sanger sequencing to verify the presence of the above-mentioned PTEN-COL17A1 and EGFR-RP11-745C15.2 fusion transcript in Positive samples of CGGA and Another40 samples,and performed the Western blotting to measure the protein expression of PTEN-COL17A1 gene fusion.In vitro experiments,COL17A1 knockdown was achieved following transient transfection using three different human fascin si RNA duplexes in U251 cells,The expression vector(pc DNA3.1)that express COL17A1(pc DNA3.1-COL17A1)and a negative control(pc DNA3.1-NC)were transfected into H4 cells or U87 cells.CCK8,plate colony and migration assays were used to examine proliferation,Invasion and immigration.EGFR-RP11-745C15.2 fusion sequences were generated by over lap PCR,and then constructed EGFR-RP11-745C15.2 fusion plasmids.EGFR-RP11-745C15.2 fusion was transfected into the U87 cell line and then functional studies were performed.Results PTEN-COL17A1 chimera,comprising intron 1 of PTEN fused to intron14 of COL17A1,was identified in one positive samples of transcriptome sequencing from CGGA.EGFR-RP11-745C15.2 chimera,comprising exon 23 of EGFR fused to RP11-745C15.2 was identified in two positive samples of transcriptome sequencing from CGGA and four samples in Another40 samples.The 4 positive samples were all high-grade gliomas.We observe the COL17A1 protein over expression in the positive sample of PTEN-COL17A1 gene fusion,that revealed the PTEN-COL17A1 gene fusion might have resulted in COL17A1 protein over expression.Knocked down Collagen XVII expression in U251 glioma cell lines resulted in decreased tumor invasiveness,along with significant reduction of MMP9 expression,while increased collagen XVII expression promotes invasive activities of H4 and U87 glioma cells along with significant elevated the protein expression of MMP9.EGFR-RP11-745C15.2 fusion plasmids were constructed successfully.In the U87 cell line,overexpression of the EGFR-RP11-745C15.2 fusion gene was observed to inhibit tumor migration and proliferation,and to promote apoptosis.Conclusions The study validated the existence of this PTEN-COL17A1 and EGFR-RP11-745C15.2fusion gene.EGFR-RP11-745C15.2 fusion gene might highly correlated with high-grade gliomas especially WHO Ⅳgrade gliomas We observe the COL17A1 protein over expression in positive sample of PTEN-COL17A1 gene fusion,that revealed the PTEN-COL17A1 gene fusion might resulted in COL17A1 protein over expression.COL17A1 may promote cell invasion through the MMP9-hydrolyzed cell matrix.EGFR-RP11-745C15.2 fusion gene may inhibit tumor migration and proliferation,and promote apoptosis of cells... |
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