Integrated genome and transcriptome sequencing identifies a noncoding mutation in the genome replication factor DONSON as the cause of microcephaly-micromelia syndrome

Genome Res. 2017 Aug;27(8):1323-1335. doi: 10.1101/gr.219899.116.

Gilad D Evrony ¹ ² ³, Dwight R Cordero ⁴, Jun Shen ⁵ ⁶, Jennifer N Partlow ¹ ² ³, Timothy W Yu ¹ ² ³, Rachel E Rodin ¹ ² ³, R Sean Hill ¹ ² ³, Michael E Coulter ¹ ² ³, Anh-Thu N Lam ¹ ² ³, Divya Jayaraman ¹ ² ³, Dianne Gerrelli ⁷, Diana G Diaz ⁷, Chloe Santos ⁷, Victoria Morrison ⁷, Antonella Galli ⁸, Ulrich Tschulena ⁹, Stefan Wiemann ⁹, M Jocelyne Martel ¹⁰, Betty Spooner ¹¹, Steven C Ryu ¹ ² ³, Princess C Elhosary ¹ ² ³, Jillian M Richardson ¹ ² ³, Danielle Tierney ¹ ² ³, Christopher A Robinson ¹², Rajni Chibbar ¹², Dana Diudea ¹², Rebecca Folkerth ⁵, Sheldon Wiebe ¹³, A James Barkovich ¹⁴, Ganeshwaran H Mochida ¹ ² ³ ¹⁵, James Irvine ¹¹ ¹⁶, Edmond G Lemire ¹⁷, Patricia Blakley ¹⁷, Christopher A Walsh ¹ ² ³

Affiliations

1 Division of Genetics and Genomics, Manton Center for Orphan Disease, and Howard Hughes Medical Institute, Boston Children's Hospital, Boston, Massachusetts 02115, USA.
2 Departments of Neurology and Pediatrics, Harvard Medical School, Boston, Massachusetts 02115, USA.
3 Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.
4 Department of Obstetrics, Gynecology and Reproductive Biology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
5 Department of Pathology, Brigham and Women's Hospital and Harvard Medical School, Boston, Massachusetts 02115, USA.
6 Laboratory of Molecular Medicine, Partners Personalized Medicine, Cambridge, Massachusetts 02139, USA.
7 Institute of Child Health, University College London, London WC1N 1EH, United Kingdom.
8 Wellcome Trust Sanger Institute, Cambridge CB10 1SA, United Kingdom.
9 Division of Molecular Genome Analysis, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany.
10 Department of Obstetrics and Gynecology, University of Saskatchewan College of Medicine, Saskatoon, Saskatchewan S7N 5E5, Canada.
11 Northern Medical Services, University of Saskatchewan College of Medicine, Saskatoon, Saskatchewan S7K 0L4, Canada.
12 Department of Pathology, Royal University Hospital, University of Saskatchewan, Saskatoon, Saskatchewan S7N 0W8, Canada.
13 Department of Medical Imaging, Royal University Hospital, University of Saskatchewan, Saskatoon, Saskatchewan S7N 0W8, Canada.
14 Department of Radiology, University of California San Francisco, San Francisco, California 94143, USA.
15 Pediatric Neurology Unit, Department of Neurology, Massachusetts General Hospital, Boston, Massachusetts 02114, USA.
16 Population Health Unit, Mamawetan Churchill River and Keewatin-Yatthé Health Regions, and Athabasca Health Authority, La Ronge, Saskatchewan S0J 1L0, Canada.
17 Department of Pediatrics, Royal University Hospital, University of Saskatchewan, Saskatoon, Saskatchewan S7N 0W8, Canada.

PMID: 28630177 DOI: 10.1101/gr.219899.116

Abstract

While next-generation sequencing has accelerated the discovery of human disease genes, progress has been largely limited to the "low hanging fruit" of mutations with obvious exonic coding or canonical splice site impact. In contrast, the lack of high-throughput, unbiased approaches for functional assessment of most noncoding variants has bottlenecked gene discovery. We report the integration of transcriptome sequencing (RNA-seq), which surveys all mRNAs to reveal functional impacts of variants at the transcription level, into the gene discovery framework for a unique human disease, microcephaly-micromelia syndrome (MMS). MMS is an autosomal recessive condition described thus far in only a single First Nations population and causes intrauterine growth restriction, severe microcephaly, craniofacial anomalies, skeletal dysplasia, and neonatal lethality. Linkage analysis of affected families, including a very large pedigree, identified a single locus on Chromosome 21 linked to the disease (LOD > 9). Comprehensive genome sequencing did not reveal any pathogenic coding or canonical splicing mutations within the linkage region but identified several nonconserved noncoding variants. RNA-seq analysis detected aberrant splicing in DONSON due to one of these noncoding variants, showing a causative role for DONSON disruption in MMS. We show that DONSON is expressed in progenitor cells of embryonic human brain and other proliferating tissues, is co-expressed with components of the DNA replication machinery, and that Donson is essential for early embryonic development in mice as well, suggesting an essential conserved role for DONSON in the cell cycle. Our results demonstrate the utility of integrating transcriptomics into the study of human genetic disease when DNA sequencing alone is not sufficient to reveal the underlying pathogenic mutation.

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