Sequencing of Idd regions in the NOD mouse genome

Background

Idd regions currently annotated by HAVANA. [Genome Research Limited]

Type 1 diabetes is characterised by hyperglycaemia that results from progressive autoimmune T cell-mediated destruction of the insulin-producing β-cells of the islets of Langerhans in the pancreas and is fatal if not treated. Type 1 diabetes is typically associated with specific allelic variants of the MHC class I and class II genes within the Major Histocompatability Complex (MHC), a region that is critical for mounting immune and autoimmune responses. Type 1 diabetes is a complex disease with nearly 50 loci known to be involved. To date however, the MHC is the only locus that has been found to be essential for the manifestation of this disease. The NOD mouse spontaneously develops type 1 diabetes and as such represents a valuable tool for studying the genetics of type 1 diabetes and for evaluating therapeutic interventions, since it shares multiple characteristics with the human disease such as genetic polymorphisms affecting shared pathways, common antigenic targets, and the expression of class II MHC molecules displaying related peptides.

Funding

The NOD bacterial artificial chromosome (BAC) sequencing was funded by Immune Tolerance Network (ITN) Contract AI 15416, which was sponsored by the National Institute of Allergy and Infectious Diseases (NIAID), the National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK), and the Juvenile Diabetes Research Foundation International (JDRF).

Collaborations

We are working closely with the following organisations:

• Diabetes and Inflammation Laboratory
• The Hospital for Sick Children
• Institut Pasteur
• Institut de recherches cliniques de Montréal
• University of Cincinnati College of Medicine
• University of Virginia School of Medicine

Clone libraries

Two NOD mouse BAC libraries - the DIL NOD and the CHORI-29 NOD, were constructed. We generated a clone map from these two libraries by mapping the BAC end-sequences to the latest assembly of the C57BL/6J mouse reference genome sequence and displaying them in Ensembl.

NOD mouse libraries

Targeted sequencing

In conjunction with external collaborators studying the genetics of NOD mice, clones covering defined Idd candidate regions were selected for whole BAC sequencing from either of the two NOD mouse libraries using the BAC end-sequence alignments in Ensembl. Sequencing was then carried out using T7 and SP6 primers on the vector, and big dye terminator chemistry. In parallel, the quality of the corresponding sequence in the C57BL/6J reference mouse has been checked. Clones from the DIL NOD BAC library constructed by RIKEN Genomic Sciences Centre (Japan) in conjunction with the Diabetes and Inflammation Laboratory (DIL) (University of Cambridge) from the NOD/MrkTac mouse strain are designated DIL. Clones from the CHORI-29 NOD BAC library constructed by Pieter de Jong (Children's Hospital, Oakland, California, USA) from the NOD/ShiLtJ mouse strain are designated CHORI-29.

Targeted regions

The approximate coordinates for the syntenic Idd regions in the Black 6 mouse, based upon available NOD sequence are shown below and are not necessarily exact. To access the NOD sequence click on the region.

Software & data resources

All NOD mouse sequences have been submitted to the International Nucleotide Sequence Database Consortium (INSDC), deposited in the NCBI trace archive and can also be downloaded from the NOD clone assembly status webpage or our ftp site. We have generated a Distributed Annotation System (DAS) source to display both the DIL NOD clones and the CHORI-29 NOD clones. These BAC end-sequence alignments can then be visualized in the Ensembl mouse genome browser where the alignments of both NOD BAC libraries can be accessed through the DAS sources menu (configured in the Other DNA alignments section under Configure this page) and viewed against the reference C57BL/6J genome. DIL NOD clones are displayed as red and black lines depending on the orientation of the insert in the vector, while CHORI-29 NOD clones are displayed similarly as green and blue lines.

The Mouse Genomes Project has used the Illumina platform to sequence the entire NOD/ShiLtJ genome and this should help to position unaligned BAC end-sequences to novel non-reference regions of the NOD genome. Further information about the BAC end-sequences, such as their alignment, variation data and Ensembl gene coverage, can be obtained from the NOD mouse ftp site.

Finished clones from the targeted Idd candidate regions are displayed in the NOD clone sequence section of the website, where they can be downloaded either as individual clone sequences or larger contigs that make up the accession golden path (AGP). To access all the sequence for a specific region, select the Idd region from the relevant chromosome dropdown menu and then click on "Show AGP". Clicking to the right of the vertical green bar will download the complete sequence for a contig. The importance and utility of these high quality finished sequences is demonstrated further by the essential role that the NOD/ShiLtJ strain derived CHORI-29 NOD BACs played in calibrating the variation calling software for the Mouse Genomes Project.

Analysis and manual annotation

Analysed and manually annotated sequences have been generated using in-house developed software in accordance with the manual annotation guidelines, and are available through the Vertebrate Genome Annotation browser Vega. Completed C57BL/6J annotation can also be viewed in the Vega genome browser alongside the NOD sequence. This allows comparison of the genomic sequence and genes in the candidate loci between diabetes resistant and diabetes sensitive strains, looking for example for SNPs, and is a useful way of identifying regions of difference between the two mouse strains.

Publications

• The B10 Idd9.3 locus mediates accumulation of functionally superior CD137(+) regulatory T cells in the nonobese diabetic type 1 diabetes model.

  Kachapati K, Adams DE, Wu Y, Steward CA, Rainbow DB, Wicker LS, Mittler RS and Ridgway WM

  Journal of immunology (Baltimore, Md. : 1950) 2012;189;10;5001-15

  PUBMED: 23066155; PMC: 3505683; DOI: 10.4049/jimmunol.1101013

• Mouse genomic variation and its effect on phenotypes and gene regulation.

  Keane TM, Goodstadt L, Danecek P, White MA, Wong K, Yalcin B, Heger A, Agam A, Slater G, Goodson M, Furlotte NA, Eskin E, Nellåker C, Whitley H, Cleak J, Janowitz D, Hernandez-Pliego P, Edwards A, Belgard TG, Oliver PL, McIntyre RE, Bhomra A, Nicod J, Gan X, Yuan W, van der Weyden L, Steward CA, Bala S, Stalker J, Mott R, Durbin R, Jackson IJ, Czechanski A, Guerra-Assunção JA, Donahue LR, Reinholdt LG, Payseur BA, Ponting CP, Birney E, Flint J and Adams DJ

  Nature 2011;477;7364;289-94

  PUBMED: 21921910; PMC: 3276836; DOI: 10.1038/nature10413

• Evidence that Cd101 is an autoimmune diabetes gene in nonobese diabetic mice.

  Rainbow DB, Moule C, Fraser HI, Clark J, Howlett SK, Burren O, Christensen M, Moody V, Steward CA, Mohammed JP, Fusakio ME, Masteller EL, Finger EB, Houchins JP, Naf D, Koentgen F, Ridgway WM, Todd JA, Bluestone JA, Peterson LB, Mattner J and Wicker LS

  Journal of immunology (Baltimore, Md. : 1950) 2011;187;1;325-36

  PUBMED: 21613616; PMC: 3128927; DOI: 10.4049/jimmunol.1003523

• Nonobese diabetic congenic strain analysis of autoimmune diabetes reveals genetic complexity of the Idd18 locus and identifies Vav3 as a candidate gene.

  Fraser HI, Dendrou CA, Healy B, Rainbow DB, Howlett S, Smink LJ, Gregory S, Steward CA, Todd JA, Peterson LB and Wicker LS

  Journal of immunology (Baltimore, Md. : 1950) 2010;184;9;5075-84

  PUBMED: 20363978; PMC: 2886967; DOI: 10.4049/jimmunol.0903734

• Genome-wide end-sequenced BAC resources for the NOD/MrkTac() and NOD/ShiLtJ() mouse genomes.

  Steward CA, Humphray S, Plumb B, Jones MC, Quail MA, Rice S, Cox T, Davies R, Bonfield J, Keane TM, Nefedov M, de Jong PJ, Lyons P, Wicker L, Todd J, Hayashizaki Y, Gulban O, Danska J, Harrow J, Hubbard T, Rogers J and Adams DJ

  Genomics 2010;95;2;105-10

  PUBMED: 19909804; PMC: 2824108; DOI: 10.1016/j.ygeno.2009.10.004

• Ly49 cluster sequence analysis in a mouse model of diabetes: an expanded repertoire of activating receptors in the NOD genome.

  Belanger S, Tai LH, Anderson SK and Makrigiannis AP

  Genes and immunity 2008;9;6;509-21

  PUBMED: 18528402; PMC: 2678550; DOI: 10.1038/gene.2008.43

• Interleukin-2 gene variation impairs regulatory T cell function and causes autoimmunity.

  Yamanouchi J, Rainbow D, Serra P, Howlett S, Hunter K, Garner VE, Gonzalez-Munoz A, Clark J, Veijola R, Cubbon R, Chen SL, Rosa R, Cumiskey AM, Serreze DV, Gregory S, Rogers J, Lyons PA, Healy B, Smink LJ, Todd JA, Peterson LB, Wicker LS and Santamaria P

  Nature genetics 2007;39;3;329-37

  PUBMED: 17277778; PMC: 2886969; DOI: 10.1038/ng1958

• Molecular genetic analysis of the Idd4 locus implicates the IFN response in type 1 diabetes susceptibility in nonobese diabetic mice.

  Ivakine EA, Gulban OM, Mortin-Toth SM, Wankiewicz E, Scott C, Spurrell D, Canty A and Danska JS

  Journal of immunology (Baltimore, Md. : 1950) 2006;176;5;2976-90

  PUBMED: 16493056

• Natural genetic variants influencing type 1 diabetes in humans and in the NOD mouse.

  Wicker LS, Moule CL, Fraser H, Penha-Goncalves C, Rainbow D, Garner VE, Chamberlain G, Hunter K, Howlett S, Clark J, Gonzalez-Munoz A, Cumiskey AM, Tiffen P, Howson J, Healy B, Smink LJ, Kingsnorth A, Lyons PA, Gregory S, Rogers J, Todd JA and Peterson LB

  Novartis Foundation symposium 2005;267;57-65; discussion 65-75

  PUBMED: 15999801

• Fine mapping, gene content, comparative sequencing, and expression analyses support Ctla4 and Nramp1 as candidates for Idd5.1 and Idd5.2 in the nonobese diabetic mouse.

  Wicker LS, Chamberlain G, Hunter K, Rainbow D, Howlett S, Tiffen P, Clark J, Gonzalez-Munoz A, Cumiskey AM, Rosa RL, Howson JM, Smink LJ, Kingsnorth A, Lyons PA, Gregory S, Rogers J, Todd JA and Peterson LB

  Journal of immunology (Baltimore, Md. : 1950) 2004;173;1;164-73

  PUBMED: 15210771

• Identification of a structurally distinct CD101 molecule encoded in the 950-kb Idd10 region of NOD mice.

  Penha-Gonçalves C, Moule C, Smink LJ, Howson J, Gregory S, Rogers J, Lyons PA, Suttie JJ, Lord CJ, Peterson LB, Todd JA and Wicker LS

  Diabetes 2003;52;6;1551-6

  PUBMED: 12765969

Useful links

• Diabetes and Inflammation Laboratory
• Children's Hospital Oakland Research Institute
• T1DBase
• BACPAC
• The Jackson Laboratory
• NIAID
• NIDDK
• JDRF
• ITN