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Am J Hum Genet. 2019 Sep 5;105(3):573-587. doi: 10.1016/j.ajhg.2019.07.013. Epub 2019 Aug 22.

Pathogenic Abnormal Splicing Due to Intronic Deletions that Induce Biophysical Space Constraint for Spliceosome Assembly.

Author information

1
Kids Neuroscience Centre, Kids Research, The Children's Hospital at Westmead, Sydney, NSW 2145, Australia; Discipline of Child and Adolescent Health, Sydney Medical School, University of Sydney, Sydney, NSW 2006, Australia.
2
Kids Neuroscience Centre, Kids Research, The Children's Hospital at Westmead, Sydney, NSW 2145, Australia.
3
Kids Neuroscience Centre, Kids Research, The Children's Hospital at Westmead, Sydney, NSW 2145, Australia; Discipline of Child and Adolescent Health, Sydney Medical School, University of Sydney, Sydney, NSW 2006, Australia; The Children's Medical Research Institute, 214 Hawkesbury Road, Sydney, NSW 2145, Australia.
4
Max-Planck-Institute for Biophysical Chemistry, Department of Cellular Biochemistry, Am Fassberg 11, 37077 Göttingen, Germany.
5
Kids Neuroscience Centre, Kids Research, The Children's Hospital at Westmead, Sydney, NSW 2145, Australia; Discipline of Child and Adolescent Health, Sydney Medical School, University of Sydney, Sydney, NSW 2006, Australia; Royal Adelaide Hospital, Department of Neurology, Adelaide, SA 5000, Australia.
6
Centre for Medical Research, The University of Western Australia, Harry Perkins Institute of Medical Research, Nedlands, WA 6009, Australia.
7
Analytic and Translational Genetics Unit, Massachusetts General Hospital, Boston, MA 02114, USA; Center for Mendelian Genomics, Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, MA 02142, USA; Medical and Population Genetics, Broad Institute of Massachusetts Institute of Technology and Harvard, Cambridge, MA 02142, USA.
8
Department of Histopathology, The Children's Hospital at Westmead, Sydney Children's Hospital Network, Sydney, NSW 2145, Australia.
9
Neurogenetics Clinic, The Children's Hospital at Westmead, Sydney Children's Hospital Network, Sydney, NSW 2145, Australia.
10
Neurogenetic Unit, Department of Diagnostic Genomics, PathWest Laboratory Medicine, QEII Medical Centre, Nedlands, WA 6009, Australia.
11
Kids Neuroscience Centre, Kids Research, The Children's Hospital at Westmead, Sydney, NSW 2145, Australia; Discipline of Child and Adolescent Health, Sydney Medical School, University of Sydney, Sydney, NSW 2006, Australia; The Children's Medical Research Institute, 214 Hawkesbury Road, Sydney, NSW 2145, Australia. Electronic address: sandra.cooper@sydney.edu.au.

Abstract

A precise genetic diagnosis is the single most important step for families with genetic disorders to enable personalized and preventative medicine. In addition to genetic variants in coding regions (exons) that can change a protein sequence, abnormal pre-mRNA splicing can be devastating for the encoded protein, inducing a frameshift or in-frame deletion/insertion of multiple residues. Non-coding variants that disrupt splicing are extremely challenging to identify. Stemming from an initial clinical discovery in two index Australian families, we define 25 families with genetic disorders caused by a class of pathogenic non-coding splice variant due to intronic deletions. These pathogenic intronic deletions spare all consensus splice motifs, though they critically shorten the minimal distance between the 5' splice-site (5'SS) and branchpoint. The mechanistic basis for abnormal splicing is due to biophysical constraint precluding U1/U2 spliceosome assembly, which stalls in A-complexes (that bridge the 5'SS and branchpoint). Substitution of deleted nucleotides with non-specific sequences restores spliceosome assembly and normal splicing, arguing against loss of an intronic element as the primary causal basis. Incremental lengthening of 5'SS-branchpoint length in our index EMD case subject defines 45-47 nt as the critical elongation enabling (inefficient) spliceosome assembly for EMD intron 5. The 5'SS-branchpoint space constraint mechanism, not currently factored by genomic informatics pipelines, is relevant to diagnosis and precision medicine across the breadth of Mendelian disorders and cancer genomics.

KEYWORDS:

5′ splice site; abnormal splicing; branchpoint; intronic deletion; non-coding variant; pathogenic splice variant; pre-mRNA splicing; spliceosome assembly

PMID:
31447096
PMCID:
PMC6731365
[Available on 2020-03-05]
DOI:
10.1016/j.ajhg.2019.07.013

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