In vivo RNA interference demonstrates a role for Nramp1 in modifying susceptibility to type 1 diabetes

Abstract

Type 1 diabetes is an autoimmune disease influenced by multiple genetic loci. Although more than 20 insulin-dependent diabetes (Idd) loci have been implicated in the nonobese diabetic (NOD) mouse model1,2, few causal gene variants have been identified3,4. Here we show that RNA interference5,6 (RNAi) can be used to probe candidate genes in this disease model. Slc11a1 encodes a phagosomal ion transporter, Nramp1, that affects resistance to intracellular pathogens and influences antigen presentation7,8,9. This gene is the strongest candidate among the 42 genes in the Idd5.2 region; a naturally occurring mutation in the protective Idd5.2 haplotype results in loss of function of the Nramp1 protein10. Using lentiviral transgenesis11, we generated NOD mice in which Slc11a1 is silenced by RNAi. Silencing reduced the frequency of type 1 diabetes, mimicking the protective Idd5.2 region. Our results demonstrate a role for Slc11a1 in modifying susceptibility to type 1 diabetes and illustrate that RNAi can be used to study causal genes in a mammalian model organism.

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Figure 1: Dose effect of the protective B10-derived Idd5.2 region.
Figure 2: Design of a lentiviral vector for Nramp1 knockdown.
Figure 3: Generation of Nramp1-knockdown Idd5.1 congenic NOD mice.
Figure 4: Nramp1 silencing affects S. enterica infection and diabetes frequency.

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Acknowledgements

The authors thank J. Blackwell for the gift of Slc11a1 cDNA; J. Bode for the gift of the SAR DNA; J. Todd for advice and support; P. Sandy, C. Dillon, B. Haines and A. Ventura for technical discussions and A. Ayerdi, M. Serrano and T. Chavarria for technical assistance. This work was supported in part by the Cambridge-MIT Institute. L.S.W. and K.H. are supported by grants from the Juvenile Diabetes Research Foundation (JDRF) and the Wellcome Trust. The availability of NOD congenic mice through the Taconic Farms Emerging Models Program has been supported by grants from the Merck Genome Research Institute, the US National Institute of Allergy and Infectious Disease and the JDRF.

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Correspondence to Stephan Kissler.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Dose effect of protective alleles at Idd5.2 in the absence of protective alleles at Idd5.1. (PDF 340 kb)

Supplementary Fig. 2

Variegated expression in pLL3.7 transgenic mice. (PDF 241 kb)

Supplementary Fig. 3

Lentiviral construct expression in pLB-915 founder. (PDF 300 kb)

Supplementary Fig. 4

Reduced expression caused by interference between lentiviral constructs. (PDF 346 kb)

Supplementary Fig. 5

Nramp1 silencing protects from experimental autoimmune encephalomyelitis (EAE). (PDF 356 kb)

Supplementary Table 1

Primer sequences used for cloning and target sequences used for shRNA design. (PDF 10 kb)

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Kissler, S., Stern, P., Takahashi, K. et al. In vivo RNA interference demonstrates a role for Nramp1 in modifying susceptibility to type 1 diabetes. Nat Genet 38, 479–483 (2006). https://doi.org/10.1038/ng1766

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