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. 2015 Jul 2;162(1):45-58.
doi: 10.1016/j.cell.2015.06.001. Epub 2015 Jun 18.

Critical Role for the DNA Sensor AIM2 in Stem Cell Proliferation and Cancer

Si Ming Man  1 , Qifan Zhu  2 , Liqin Zhu  3 , Zhiping Liu  4 , Rajendra Karki  1 , Ankit Malik  1 , Deepika Sharma  1 , Liyuan Li  3 , R K Subbarao Malireddi  1 , Prajwal Gurung  1 , Geoffrey Neale  5 , Scott R Olsen  5 , Robert A Carter  6 , Daniel J McGoldrick  6 , Gang Wu  6 , David Finkelstein  6 , Peter Vogel  7 , Richard J Gilbertson  8 , Thirumala-Devi Kanneganti  9
Affiliations

Critical Role for the DNA Sensor AIM2 in Stem Cell Proliferation and Cancer

Si Ming Man et al. Cell. .

Abstract

Colorectal cancer is a leading cause of cancer-related deaths. Mutations in the innate immune sensor AIM2 are frequently identified in patients with colorectal cancer, but how AIM2 modulates colonic tumorigenesis is unknown. Here, we found that Aim2-deficient mice were hypersusceptible to colonic tumor development. Production of inflammasome-associated cytokines and other inflammatory mediators was largely intact in Aim2-deficient mice; however, intestinal stem cells were prone to uncontrolled proliferation. Aberrant Wnt signaling expanded a population of tumor-initiating stem cells in the absence of AIM2. Susceptibility of Aim2-deficient mice to colorectal tumorigenesis was enhanced by a dysbiotic gut microbiota, which was reduced by reciprocal exchange of gut microbiota with healthy wild-type mice. These findings uncover a synergy between a specific host genetic factor and gut microbiota in determining the susceptibility to colorectal cancer. Therapeutic modulation of AIM2 expression and microbiota has the potential to prevent colorectal cancer.

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Figures

Figure 1
Figure 1. AIM2 prevents colitis-associated colorectal tumorigenesis. See also Figure S1
(A) Relative expression of the gene encoding AIM2 in tissues of WT C57BL/6 mice. (B) Relative expression of the gene encoding AIM2 in colon tissues of untreated WT mice or WT mice 14 days after injection of AOM. (C) Relative expression of the gene encoding AIM2 in colon tissues of untreated WT mice or WT mice 80 days after injection of AOM. (D) Colon tumors in WT and Aim2−/− mice 80 days after injection of AOM. (E and F) Number and size of colon tumors in WT (n=34) and Aim2−/− (n=27) mice. (G) Body weight change of mice pooled from six independent experiments. (H) Histological scores 80 days after injection of AOM. (I) Percentages of mice with dysplasia 80 days after injection of AOM. (J) H&E staining of colon tumors. Original magnification, (J) 10×. Each symbol represents an individual mouse (E,H). * P<0.05; ** P<0.01; *** P<0.001; **** P<0.0001; NS, not statistically significant. (two tailed t-test (B,C,E,F,H) or One-way ANOVA (G)). Data represent two (A–C) or six independent experiments (D–J; mean and s.e.m. in B,C,E,F,G,H).
Figure 2
Figure 2. AIM2 governs colorectal tumorigenic susceptibility independently of inflammasomes and a number of inflammatory mediators. See also Figure S2
(A) Caspase-1 processing in colon lysates from WT and Aim2−/− mice 14 days after injection of AOM. (B) Levels of IL-18 in the colon tissue or serum (C) Levels of IL-1β in colon tissues (D) The lengths of colon from WT and Aim2−/− mice 14 days after injection of AOM. (E) Immunoblot analysis (left) and densitometric quantification of band intensity of phosphorylated and total ERK and IκBα (right) in colon tissues of WT and Aim2−/− mice 14 days after injection of AOM. (F) Levels of IL-6, TNF, G-CSF and KC in colon tissues. (G) Percentages of CD4+ cells, CD4+ cells expressing IFN-γ or IL-17, CD8+ cells, CD8+ cells expressing IFN-γ, and CD11b+ cells, CD11b+ cells expressing IL-6 or TNF. Each symbol represents an individual mouse (B–D,F,G). * P<0.05; ** P<0.01; NS, not statistically significant. (two tailed t-test (B–D,F,G)). Data represent one experiment representative of two independent experiment (A–C,G) or pooled from three independent experiments (D–F; mean and s.e.m. in B–G).
Figure 3
Figure 3. AIM2 suppresses overt proliferation. See also Figure S3
(A) Quantification of the number of BrdU+ cells in each crypt of WT (day 8 n=3, day 14 n=5) and Aim2−/− (day 8, n=4; day 14, n=5) mice. Quantification of the number of Ki67+ in each crypt of WT (day 0, n=4; day 14, n=4) and Aim2−/− (day 0, n=3, day 14, n=4) mice. (B) Microarray analysis of genes encoding molecules involved in proliferation with significantly higher or lower expression in Aim2−/− mice 14 days after injection of AOM compared with their WT counterparts. (C) Real time qRT-PCR analysis of genes encoding S100A9, SNRPD1 and DBF4 in WT and Aim2−/− mice 14 days after injection of AOM. (D) Immunoblot analysis of phosphorylated and total AKT and phosphorylated and total PTEN from colon tissues. (E) Immunoblot analysis of c-Myc, HIF1-α and GAPDH (loading control) from colon tissues. (F) Real-time qRT-PCR analysis of the gene encoding c-Myc in colon tissues of mice 14 days after injection of AOM. (G and H) Immunoblot analysis of caspase-3 (G) and caspase-7 activation (H) in colon tissues mice 8 days after injection of AOM. At least 20 crypts were counted in each animal (A). Each symbol represents one crypt (A) or one mouse (C and F). * P <0.05; ** P <0.01; ***P <0.001; ****P <0.0001; NS, no statistical significance (two tailed t-test (A,C,F)). Data represent one experiment with two biological replicates per genotype (B) or from one experiment representative of two independent experiments (A,C,D–H) (mean and s.e.m. in A,C,F).
Figure 4
Figure 4. AIM2 controls proliferation of intestinal progenitor stem cells
Formation of organoids derived from colonic stem cells harvested from untreated WT and Aim2−/− mice. Quantification of the number (top right) and size of organoids (bottom right). Scale bars, 50 μm (left) or 200 μm (right). Data represent one of three independent experiments.
Figure 5
Figure 5. AIM2 controls expansion of intestinal stem cells. See also Figure S4
(A) Lineage tracing (left) of Prom1+ cells (green) in the large intestine three weeks post tamoxifen (TMX) induction. Blue staining indicates DAPI. Scale bar, 100 μm. Staining for phosphorylated AKT (p-AKT, Ser473), total AKT and c-Myc (right) in the large intestine six weeks post TMX induction. Scale bar, 200 μm. (B) H&E staining and staining for p-AKT (Ser473), total AKT, c-Myc and Ki67 staining in the small intestine six weeks post TMX induction. Arrowheads indicate early lesions with intense staining for the respective markers. Scale bars, 250 μm (H&E), 100 μm (p-AKT, total AKT, c-Myc and Ki67 staining).
Figure 6
Figure 6. The role of hematopoietic and non-hematopoietic compartments in AIM2-mediated protection of colorectal tumorigenesis
(A) Four groups of mice were generated by bone marrow transplantation: (1) WT>WT n=9; (2) Aim2−/−>WT n=9; (3) WT>Aim2−/− n=7; (4) Aim2−/−>Aim2−/− n=8. Six weeks following bone marrow transplantation, mice were treated with AOM followed by DSS and colon tumors were counted after 80 days. Numbers of tumors in the colon and percentages of tumors of less than 2 mm or 2–4 mm in diameter. (B) Representative images of colon tumors. (C) Representative H&E images of colon tumors. Magnification: 20×. Each symbol in the graph represents one mouse (A). Data represent one experiment (mean and s.e.m.). *P<0.05; **P<0.01; ***P<0.001. One-way ANOVA (A).
Figure 7
Figure 7. Reciprocal exchange of gut microbiota reduces susceptibility of Aim2−/− mice to colon cancer. See also Figure S5
(A) Bacterial species from feces which had the greatest difference in abundance between single housed WT and Aim2−/− mice and their relative abundance in co-housed WT and Aim2−/− mice, based on analysis of the 16S rRNA gene, are shown in a heatmap in alphabetical order. (B) Principal Component Analysis of A. (C) The total number of tumors observed in the colon of separately housed and co-housed mice 80 days after injection of AOM. (D) Representative images of colons in C. Each column represents one mouse (A). Each symbol represents one mouse (B and C). Data represent one experiment (A and B) or pooled from two independent experiments (C and D) (mean and s.e.m. in C). *P<0.05; ****P<0.0001. One-way ANOVA (C).
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Comment in

  • AIMing 2 Curtail Cancer.
    Rommereim LM, Subramanian N. Rommereim LM, et al. Cell. 2015 Jul 2;162(1):18-20. doi: 10.1016/j.cell.2015.06.041. Cell. 2015. PMID: 26140588

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