| Objective: Transcriptome sequencing technology is used to detect the transcriptome changes of colorectal cancer cells caused by knockdown PDSS1 expression,and then the functions and pathways reflected behind the differentially expressed genes are analyzed in combination with bioinformatics methods.To provide a theoretical basis for studying the mechanism of PDSS1 promoting the proliferation and metastasis of colorectal cancer.Methods: 1.Constructed colorectal cancer cell lines and control cells with stable knockout and overexpression of PDSS1,and verified the transfection efficiency by q RT-PCR and Western blot technology.2.Using the Illumina platform to perform transcriptome sequencing and quality control of the results in 3 groups of colorectal cancer cell samples.Detecting of gene differential expression,and performed GO function analysis and KEGG pathway function enrichment analysis.3.To further screen the main differentially expressed genes,we verified the correlation between PDSS1 and differentially expressed genes by consulting the literature,q RT-PCR and analysis of TCGA databases.4.Combined with analysis of TCGGA databases,the potential signaling pathway of PDSS1 to the development of colorectal cancer was explored and verified by q RT-PCR.Results: 1.Successfully constructed colorectal cancer cell lines and control strains with stable knockdown and overexpression of PDSS1 by lentiviral infection(SW480-Vector vs.SW480-PDSS1,SW620-Scramble vs.SW620-sh PDSS1).2.A total of 60.78 Gb of data were obtained through transcriptome analysis,of which the Q30 base percentage was more than 94.50%,and the comparison efficiency with the reference genome was more than 94.57%.3.According to the Fold Change ≥ 2 and the error detection rate < 0.05,a total of 5935 differentially expressed genes were screened,accounting for 33.9% of the total number of screenings.Among them,there are 2430 genes expressing upregulation and 3505 expressing downregulation.4.GO function analysis showed that the functional changes of colorectal cancer gene mapping caused by differences in PDSS1 gene expression were mainly concentrated in DNA-templated,mitotic cell cycle,transcription of RNA polymerase II promoters,Nucleic acid binding,RNA binding,RNA polymerase II regulatory region sequence-specific and other cell proliferation and growth-related processes.5.Based on Q value < 0.05,the results of KEGG Pathway functional pathway enrichment showed a total of 26 pathways were correlated with PDSS1 expression levels,including some tumorrelated functional pathways,such as Hippo signaling pathway,MAPK signaling pathway and Wnt signaling pathway.6.Through q RT-PCR and correlation analysis using the data form TCGA databases,we found that PDSS1 expression status was positively associated with the expression levels of MYC,meanwhile PDSS1 may promoted promoted the development of colorectal cancer by increasing the MYC expression.7.Our results of KEGG analysis and q RT-PCR indicated that PDSS1 promotes MYC expression by activating the WNT pathway.Conclusion: 1.PDSS1 expression can affect the expression of multiple genes and related signaling pathways in colorectal cancer.2.PDSS1 may promote the proliferation and metastasis of colorectal cancer by influencing the expression of MYC.3.PDSS1 could promote MYC expression by activating the WNT pathway. |
