Gene editing of the multi-copy H2A.B gene and its importance for fertility

Anuar, Nur Diana, Kurscheid, Sebastian, Field, Matt, Zhang, Lei, Rebar, Edward, Gregory, Philip, Buchou, Thierry, Bowles, Josephine, Koopman, Peter, Tremethick, David J., and Soboleva, Tatiana A. (2019) Gene editing of the multi-copy H2A.B gene and its importance for fertility. Genome Biology, 20. 23.

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Abstract

Background: Altering the biochemical makeup of chromatin by the incorporation of histone variants during development represents a key mechanism in regulating gene expression. The histone variant H2A.B, H2A.B.3 in mice, appeared late in evolution and is most highly expressed in the testis. In the mouse, it is encoded by three different genes. H2A.B expression is spatially and temporally regulated during spermatogenesis being most highly expressed in the haploid round spermatid stage. Active genes gain H2A.B where it directly interacts with polymerase II and RNA processing factors within splicing speckles. However, the importance of H2A.B for gene expression and fertility are unknown.

Results: Here, we report the first mouse knockout of this histone variant and its effects on fertility, nuclear organization, and gene expression. In view of the controversy related to the generation of off-target mutations by gene editing approaches, we test the specificity of TALENs by disrupting the H2A.B multi-copy gene family using only one pair of TALENs. We show that TALENs do display a high level of specificity since no off-target mutations are detected by bioinformatics analyses of exome sequences obtained from three consecutive generations of knockout mice and by Sanger DNA sequencing. Male H2A.B.3 knockout mice are subfertile and display an increase in the proportion of abnormal sperm and clogged seminiferous tubules. Significantly, a loss of proper RNA Pol II targeting to distinct transcription–splicing territories and changes to pre-mRNA splicing are observed.

Conclusion: We have produced the first H2A.B knockout mouse using the TALEN approach.

Item ID: 57058
Item Type: Article (Research - C1)
ISSN: 1474-760X
Keywords: chromatin; histone variants; H2A.B; RNA polymerase II; Pre-mRNA splicing; splicing speckles; genome editing; TALENs
Copyright Information: Copyright © The Author(s) 2019.
Additional Information:

This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

Funders: National Health and Medical Research Council (NHMRC)
Projects and Grants: NHMRC grant number 1058941
Date Deposited: 11 Feb 2019 23:48
FoR Codes: 06 BIOLOGICAL SCIENCES > 0604 Genetics > 060403 Developmental Genetics (incl Sex Determination) @ 50%
06 BIOLOGICAL SCIENCES > 0601 Biochemistry and Cell Biology > 060102 Bioinformatics @ 50%
SEO Codes: 92 HEALTH > 9201 Clinical Health (Organs, Diseases and Abnormal Conditions) > 920114 Reproductive System and Disorders @ 100%
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