Leveraging the T2T Assembly to Resolve Rare and Pathogenic Inversions in Reference Genome Gaps

Authors

Kristine Bilgrav Saether
Jesper Eisfeldt
Jesse D Bengtsson
Ming Yin Lun
Christopher M Grochowski
Medhat Mahmoud
Hsiao-Tuan Chao
Jill A Rosenfeld
Pengfei Liu
Marlene Ek
Jakob Schuy
Adam Ameur
Hongzheng Dai
Undiagnosed Diseases Network
James Paul Hwang
Fritz J Sedlazeck
Weimin Bi
Ronit Marom
Josephine Wincent
Ann Nordgren
Claudia M B Carvalho
Anna Lindstrand

Publication Date

11-20-2024

Journal

Genome Research

DOI

10.1101/gr.279346.124

PMID

39486878

PMCID

PMC11610578

PubMedCentral® Posted Date

11-3-2024

PubMedCentral® Full Text Version

Post-print

Published Open-Access

yes

Keywords

Humans, Chromosome Inversion, Genome, Human, Animals, Pan troglodytes, Whole Genome Sequencing

Abstract

Chromosomal inversions (INVs) are particularly challenging to detect due to their copy-number neutral state and association with repetitive regions. Inversions represent about 1/20 of all balanced structural chromosome aberrations and can lead to disease by gene disruption or altering regulatory regions of dosage-sensitive genes in cis. Short-read genome sequencing (srGS) can only resolve ∼70% of cytogenetically visible inversions referred to clinical diagnostic laboratories, likely due to breakpoints in repetitive regions. Here, we study 12 inversions by long-read genome sequencing (lrGS) (n = 9) or srGS (n = 3) and resolve nine of them. In four cases, the inversion breakpoint region was missing from at least one of the human reference genomes (GRCh37, GRCh38, T2T-CHM13) and a reference agnostic analysis was needed. One of these cases, an INV9 mappable only in de novo assembled lrGS data using T2T-CHM13 disrupts EHMT1 consistent with a Mendelian diagnosis (Kleefstra syndrome 1; MIM#610253). Next, by pairwise comparison between T2T-CHM13, GRCh37, and GRCh38, as well as the chimpanzee and bonobo, we show that hundreds of megabases of sequence are missing from at least one human reference, highlighting that primate genomes contribute to genomic diversity. Aligning population genomic data to these regions indicated that these regions are variable between individuals. Our analysis emphasizes that T2T-CHM13 is necessary to maximize the value of lrGS for optimal inversion detection in clinical diagnostics. These results highlight the importance of leveraging diverse and comprehensive reference genomes to resolve unsolved molecular cases in rare diseases.

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