Contributions
Abstract: EP337
Type: E-Poster Presentation
Session title: Acute lymphoblastic leukemia - Biology & Translational Research
Background
Abnormal proliferation of leukemia cells is one of the characteristics of acute lymphoblastic leukemia (ALL). But at present, we don’t fully understand the molecular mechanism of ALL cell proliferation, and there is no research about ALL proliferation-related genes from the whole genome level.
Aims
Here, CRISPR/Cas9-mediated genome-wide knockout negative screening method was used to screen ALL proliferation-related genes on genome-wide level.
Methods
After Genome-scale CRISPR-Cas9 knockout (GeCKO) v2 library plasmid amplification, lentivirus packaging, virus infection, Jurkat and Nalm6 cells at day 0 and day 14 were collected for high-throughput sequencing, and two biological replicates were performed. Then gene ontology (GO) and pathway enrichment analysis were conducted by DAVID database to analyze the proliferation-related genes obtained from the screening. And one of the genes was selected as a candidate gene. After knockout of candidate gene using CRISPR/Cas9 technology, MTS method was performed to detect cell proliferation, and flow cytometry was used to detect cell cycle. Using the database to explore the expression of candidate gene and the relationship with the prognosis of ALL.
Results
After successful establishment of the screening model, Jurkat and Nalm6 cells of two screening time point were collected for high-throughput sequencing, and the data met the quality control standards. A total of 155 proliferation-related genes were screened. The GO analysis showed that proliferation-related genes were mainly related to translation and post-transcriptional modification, while pathway enrichment analysis was closely related to metabolic pathways. PTPTM1, which was closely related to metabolism, was selected for further verification. The cell proliferation ability of Jurkat and Nalm6 cells were significantly reduced, and the cell cycle G1 phase was blocked after PTPMT1 being knocked out. Through database analysis, it is found that PTPMT1 was generally highly expressed in tumor cell lines. Comparing with normal people, PTPMT1 had higher expression levels in ALL. In children with B-ALL, those with high PTPMT1 expression have a worse prognosis.
Conclusion
In this study, CRISPR/Cas9-mediated genome-wide screening system was used to screen out 155 proliferation-related genes of Jurkat and Nalm6 cells. PTPMT1, one of the candidate genes, can inhibit the cell proliferation and block G1 phase of Jurkat and Nalm6 cells, and its expression level associated with the prognosis in children ALL.
Keyword(s): Acute lymphoblastic leukemia, Genomics, Proliferation
Abstract: EP337
Type: E-Poster Presentation
Session title: Acute lymphoblastic leukemia - Biology & Translational Research
Background
Abnormal proliferation of leukemia cells is one of the characteristics of acute lymphoblastic leukemia (ALL). But at present, we don’t fully understand the molecular mechanism of ALL cell proliferation, and there is no research about ALL proliferation-related genes from the whole genome level.
Aims
Here, CRISPR/Cas9-mediated genome-wide knockout negative screening method was used to screen ALL proliferation-related genes on genome-wide level.
Methods
After Genome-scale CRISPR-Cas9 knockout (GeCKO) v2 library plasmid amplification, lentivirus packaging, virus infection, Jurkat and Nalm6 cells at day 0 and day 14 were collected for high-throughput sequencing, and two biological replicates were performed. Then gene ontology (GO) and pathway enrichment analysis were conducted by DAVID database to analyze the proliferation-related genes obtained from the screening. And one of the genes was selected as a candidate gene. After knockout of candidate gene using CRISPR/Cas9 technology, MTS method was performed to detect cell proliferation, and flow cytometry was used to detect cell cycle. Using the database to explore the expression of candidate gene and the relationship with the prognosis of ALL.
Results
After successful establishment of the screening model, Jurkat and Nalm6 cells of two screening time point were collected for high-throughput sequencing, and the data met the quality control standards. A total of 155 proliferation-related genes were screened. The GO analysis showed that proliferation-related genes were mainly related to translation and post-transcriptional modification, while pathway enrichment analysis was closely related to metabolic pathways. PTPTM1, which was closely related to metabolism, was selected for further verification. The cell proliferation ability of Jurkat and Nalm6 cells were significantly reduced, and the cell cycle G1 phase was blocked after PTPMT1 being knocked out. Through database analysis, it is found that PTPMT1 was generally highly expressed in tumor cell lines. Comparing with normal people, PTPMT1 had higher expression levels in ALL. In children with B-ALL, those with high PTPMT1 expression have a worse prognosis.
Conclusion
In this study, CRISPR/Cas9-mediated genome-wide screening system was used to screen out 155 proliferation-related genes of Jurkat and Nalm6 cells. PTPMT1, one of the candidate genes, can inhibit the cell proliferation and block G1 phase of Jurkat and Nalm6 cells, and its expression level associated with the prognosis in children ALL.
Keyword(s): Acute lymphoblastic leukemia, Genomics, Proliferation