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Cell Reports, Volume 28
Supplemental Information
Glutamine Synthetase Promotes Radiation
Resistance via Facilitating Nucleotide
Metabolism and Subsequent DNA Damage Repair
Shujun Fu, Zhi Li, Lanbo Xiao, Wenfeng Hu, Lu Zhang, Bowen Xie, Qin Zhou, JunjuHe, Yanfang Qiu, Ming Wen, Yanni Peng, Jie Gao, Rong Tan, Yuezhen Deng, LiangWeng, and Lun-Quan Sun
Figure S1. Radiation resistant cancer cells reprogram cellular metabolic flux. Related to Figure 1.(A) Heat map of metabonomics of CNE2, CNE2-IRR, U251 and U251-IRR cell. The content for eachbiochemical factor in each sample is represented as increase in red or decrease in green (n≥4).(B) Pie charts indicating the percentage of biochemical factors with mean levels that were unchangedor significantly (p<0.05) increased or decreased.(C, E and F) Relative cellular levels of metabolites in U251 and U251-IRR cells (n=6).(D) Representative traces showing change in OCR and ECAR during mitochondrial and glycolysisstress test of U251, U251-IRR cells (n=4).Data are represented as mean ± SD. n represents experimental replicates. n.s p>0.05, *p<0.05,**p<0.01, ***p<0.001.
Figure S2. GS promoted cell survival after radiation. Related to Figure 2.(A) Heat map of transcriptomics of CNE2 and CNE2-IRR. The content for each gene expression ineach sample is represented as increase in red or decrease in blue (n=3).(B) Volcano Plot of transcriptomics of CNE2 and CNE2-IRR.(C) GO enrichment analysis of metabolic pathways, such as glucose, glutamine, lipid and nucleotidemetabolism.(D) GO enrichment analysis of pathways GS is involved in.(E) Apoptosis of U251-IRR cells with GS knockdown after 8Gy radiation.(F) Survival fraction of U251-IRR cells to 0, 2, 4, 6, 8, 10Gy radiation after GS was knocked down.Data are represented as mean ± SD. n represents experimental replicates. *p<0.05, **p<0.01.
Figure S3. GS facilitated nucleotide synthesis for DNA repair. Related to Figure 3.(A) Relative cellular nucleotide metabolites of CNE2, CNE2-IRR, U251 and U251-IRR cells (n≥5).(B) Relative cellular nucleotide metabolites of CNE2-IRR shcon, CNE2-IRR shGS1, U251-IRR shconand U251-IRR shGS1 cells (n≥5).(C) γH2A.X foci of CNE2-IRR shcon and shGS cells treated with normal (2mM) or high concentrationglutamine (4mM) at 6h after 8Gy. The scale bars represent 2.5μm.Data are represented as mean ± SD. n represents experimental replicates. n.s p>0.05, *p<0.05,**p<0.01, ***p<0.001.
Table S1 p-value, q-value and lFDR of genes from RNA sequencing. Related to Figure 2.Gene ID p-value q-value lFDRGS 0.0000974 0.0000804 0.000361399GLS 0.120863 0.03261319 0.3885743GLS2 0.02960996 0.009031987 0.09833308GLUD 0.001035973 0.000427536 0.003221275GOT1 0.4604877 0.1095139 1GOT2 0.002371866 0.000853353 0.00731315GPT 0.8636558 0.1954575 1GPT2 0.000149611 0.000110488 0.00052899ME1 0.000754532 0.00033085 0.002370909ME2 0.000176655 0.000118035 0.000614109