The genetic landscape of mutations in Burkitt lymphoma

Abstract

Burkitt lymphoma is characterized by deregulation of MYC, but the contribution of other genetic mutations to the disease is largely unknown. Here, we describe the first completely sequenced genome from a Burkitt lymphoma tumor and germline DNA from the same affected individual. We further sequenced the exomes of 59 Burkitt lymphoma tumors and compared them to sequenced exomes from 94 diffuse large B-cell lymphoma (DLBCL) tumors. We identified 70 genes that were recurrently mutated in Burkitt lymphomas, including ID3, GNA13, RET, PIK3R1 and the SWI/SNF genes ARID1A and SMARCA4. Our data implicate a number of genes in cancer for the first time, including CCT6B, SALL3, FTCD and PC. ID3 mutations occurred in 34% of Burkitt lymphomas and not in DLBCLs. We show experimentally that ID3 mutations promote cell cycle progression and proliferation. Our work thus elucidates commonly occurring gene-coding mutations in Burkitt lymphoma and implicates ID3 as a new tumor suppressor gene.

Figure 1

Results from whole-genome sequencing of a Burkitt lymphoma tumor and germline DNA. The Circos diagram summarizes the somatically acquired genetic variants in a Burkitt lymphoma genome. The outermost ring depicts the chromosome ideogram oriented clockwise, p terminus to q terminus. Centromeres are shown in red. The next three rings indicate somatically acquired mutations falling in intergenic regions, potential regulatory regions and the exome, respectively. The black arc connecting chromosomes 8 and 14 signifies a t(8;14) translocation detected in sequencing data.

Figure 2

Exome sequencing in Burkitt lymphoma. (a) The ratio of somatically acquired transitions and transversions for samples with paired normal tissue are shown for all 14 discovery set samples. (b) The heatmap indicates the mutation patterns of the 19 most frequently implicated genes out of the 70 genes mutated in Burkitt lymphoma. Each column represents an affected individual, and each row represents a gene. (c) The bar graph shows the frequency of variants found per gene across all samples, divided into nonsynonymous and synonymous counts. (d) The bar graph shows the frequency of mutations in all 70 genes mutated in Burkitt lymphoma in each case: orange bars represent samples in the discovery set, and blue bars represent samples in the validation set.

Figure 3

Patterns of exonic mutations in Burkitt lymphoma compared to DLBCL. (a) The bar graph shows the proportion of Burkitt lymphoma and DLBCL samples containing a mutation in each gene. (b) The bar graph shows the number of cases that contain a mutation in the given gene in Burkitt lymphoma and DLBCL. (c) The heatmap shows the association between 55 genes found to be recurrently mutated in Burkitt lymphoma and/or DLBCL. Blue denotes negative association between genes, and orange denotes positive association.

Figure 4

Recurrent ID3 mutations in Burkitt lymphomas. (a) Deep sequencing reads identify recurrent mutations affecting the HLH domain of ID3 in Burkitt lymphomas. Each colored line represents an individual somatic mutation or a rare genetic variant. The conservation of the HLH domain across species is also shown. Red asterisks identify the alterations that were functionally validated. (b) The bar graph shows significantly higher expression of genes involved in the G1 to S-phase transition in ID3-mutant Burkitt lymphoma samples compared to those with wild-type ID3 (FDR = 0.026), as determined in gene set enrichment analysis. Error bars, s.d. (c) Heatmap of genes corresponding to the gene set enrichment msigdb-derived list of those involved in the G1 to S-phase transition. Red denotes high relative expression, and green represents low relative expression across samples. (d) Cell cycle analysis of Jijoye cells expressing mutant ID3 proteins compared to control cells overexpressing GFP, where the red histogram denotes cells in G1 phase, orange denotes S phase, and green denotes G2 phase. The x axis shows the relative fluorescence in the PerCP-Cy5.5 channel. (e) The bar graph summarizes cell cycle analysis from an average of all cells expressing ID3 mutants compared to cells overexpressing GFP (P = 0.03, χ² test). Error bars, s.d. (f) MTT cell proliferation assay performed on Jijoye cells expressing mutant ID3 or control GFP. Absorbance was read 24 h after plating, with higher absorbance indicating more cells and signifying faster proliferation. Error bars, s.d. (g) Cell cycle analysis of BL41 cells expressing wild-type ID3 compared to those expressing GFP control. (h) The bar graph summarizes cell cycle analysis for BL41 cells expressing wild-type ID3 relative to cells expressing GFP control. (i) MTT cell proliferation assay performed on BL41 cells expressing wild-type ID3 or GFP. Error bars, s.d.