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Title: Discovery of candidate disease genes in ENU-induced mouse mutants by large-scale sequencing, including a splice-site mutation in nucleoredoxin

Author
item BOLES, MELISSA - Baylor College Of Medicine
item WILKINSON, BONNEY - Baylor College Of Medicine
item WILMING, LAURENS - Wellcome Trust Sanger Institute
item LIU, BIN - Baylor College Of Medicine
item PROBST, FRANK - Baylor College Of Medicine
item HARROW, JENNIFER - Wellcome Trust Sanger Institute
item GRAFHAM, DARREN - Wellcome Trust Sanger Institute
item HENTGES, KATHRYN - University Of Manchester
item WOODWARD, LANETTE - Baylor College Of Medicine
item MAXWELL, ANDREA - Baylor College Of Medicine
item MITCHELL, KAREN - University Of Manchester
item RISLEY, MICHAEL - University Of Manchester
item JOHNSON, RANDY - Md Anderson Cancer Center
item HIRSCHI, KAREN - Children'S Nutrition Research Center (CNRC)
item LUPSKI, JAMES - Baylor College Of Medicine
item FUNATO, YOSUKE - University Of Osaka
item MIKI, HIROAKI - University Of Osaka
item MARIN-GARCIA, PABLO - Wellcome Trust Sanger Institute
item MATTHEWS, LUCY - Wellcome Trust Sanger Institute
item COFFEY, ALISON - Wellcome Trust Sanger Institute
item PARKER, ANNE - Wellcome Trust Sanger Institute
item HUBBARD, TIM - Wellcome Trust Sanger Institute
item ROGERS, JANE - Wellcome Trust Sanger Institute
item BRADLEY, ALLAN - Wellcome Trust Sanger Institute
item ADAMS, DAVID - Wellcome Trust Sanger Institute
item JUSTICE, MONICA - Baylor College Of Medicine

Submitted to: PLoS Genetics
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/9/2009
Publication Date: 12/11/2009
Citation: Boles, M.K., Wilkinson, B.M., Wilming, L.G., Liu, B., Probst, F.J., Harrow, J., Grafham, D., Hentges, K.E., Woodward, L.P., Maxwell, A., Mitchell, K., Risley, M.D., Johnson, R., Hirschi, K., Lupski, J.R., Funato, Y., Miki, H., Marin-Garcia, P., Matthews, L., Coffey, A.J., Parker, A., Hubbard, T.J., Rogers, J., Bradley, A., Adams, D.J., Justice, M.J. 2009. Discovery of candidate disease genes in ENU-induced mouse mutants by large-scale sequencing, including a splice-site mutation in nucleoredoxin. PLoS Genetics. 5(12):e1000759.

Interpretive Summary: We analyzed the defects in heart and blood vessel formation in mouse mutant embryos that were generated in a large scale screen to discover genes that regulate development. Since many genes involved in development are regulated by nutrients, such as retinoic acid, we may have discovered new nutrient-regulated genes that are important for embryonic development. We are now studying the functions of the genes we discovered in utero, and will determine whether they are regulated by maternal diet.

Technical Abstract: An accurate and precisely annotated genome assembly is a fundamental requirement for functional genomic analysis. Here, the complete DNA sequence and gene annotation of mouse Chromosome 11 was used to test the efficacy of large-scale sequencing for mutation identification. We re-sequenced the 14,000 annotated exons and boundaries from over 900 genes in 41 recessive mutant mouse lines that were isolated in an N-ethyl-N-nitrosourea (ENU) mutation screen targeted to mouse Chromosome 11. Fifty-nine sequence variants were identified in 55 genes from 31 mutant lines. 39% of the lesions lie in coding sequences and create primarily missense mutations. The other 61% lie in noncoding regions, many of them in highly conserved sequences. A lesion in the perinatal lethal line l11Jus13 alters a consensus splice site of nucleoredoxin (Nxn), inserting 10 amino acids into the resulting protein. We conclude that point mutations can be accurately and sensitively recovered by large-scale sequencing, and that conserved noncoding regions should be included for disease mutation identification. Only seven of the candidate genes we report have been previously targeted by mutation in mice or rats, showing that despite ongoing efforts to functionally annotate genes in the mammalian genome, an enormous gap remains between phenotype and function. Our data show that the classical positional mapping approach of disease mutation identification can be extended to large target regions using high-throughput sequencing.