• PMID: 30704500
• Gene Name: H2afb1
• Condition: Male subfertility
• Association: Associated
• Sex: Male
• Infertility type: Male infertility
• Associated genes: H2afb1
• Other associated phenotypes: Male subfertility

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

Anuar ND, Kurscheid S, Field M, Zhang L, Rebar E, Gregory P, Buchou T, Bowles J, Koopman P, Tremethick DJ, Soboleva TA.

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. FAU - Anuar, Nur Diana AU - Anuar ND AD - The John Curtin School of Medical Research, The Australian National University, Canberra, ACT, 2601, Australia. FAU - Kurscheid, Sebastian AU - Kurscheid S AD - The John Curtin School of Medical Research, The Australian National University, Canberra, ACT, 2601, Australia. AD - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia. FAU - Field, Matt AU - Field M AD - The John Curtin School of Medical Research, The Australian National University, Canberra, ACT, 2601, Australia. AD - Present Address: James Cook University, PO Box 6811, Cairns, QLD, 4870, Australia. AD - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia. FAU - Zhang, Lei AU - Zhang L AD - Sangamo Therapeutics, 501 Canal Blvd, Richmond, CA, 94804, USA. FAU - Rebar, Edward AU - Rebar E AD - Sangamo Therapeutics, 501 Canal Blvd, Richmond, CA, 94804, USA. FAU - Gregory, Philip AU - Gregory P AD - Sangamo Therapeutics, 501 Canal Blvd, Richmond, CA, 94804, USA. AD - Present Address: bluebird bio, 60 Binney St, Cambridge, MA, 02142, USA. FAU - Buchou, Thierry AU - Buchou T AD - CNRS UMR 5309, Inserm U1209, Universite' Grenoble Alpes, Institute for Advanced Biosciences, 38700, Grenoble, France. FAU - Bowles, Josephine AU - Bowles J AD - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia. FAU - Koopman, Peter AU - Koopman P AD - Institute for Molecular Bioscience, The University of Queensland, Brisbane, QLD, 4072, Australia. FAU - Tremethick, David J AU - Tremethick DJ AD - The John Curtin School of Medical Research, The Australian National University, Canberra, ACT, 2601, Australia. David.Tremethick@anu.edu.au.