Researchers at Newcastle University are using a conditional mouse mutant made by the Mary Lyon Centre’s Genome Editing Mice for Medicine (GEMM) programme to study the role of an ultra-conserved exon in the splicing regulator gene Tra2b.
Many genes function as instructions for producing proteins, which perform the majority of tasks within our cells. To enable greater complexity without increasing the number of genes needed, multiple versions of a protein can be made from a single gene, via a process called splicing. This involves modules within a gene being included or excluded from the final protein product. In some cases, a module, called a poison exon, can be included to introduce an early “stop” instruction, which prevents a functional protein being made.
The gene Tra2b encodes a splicing regulator that promotes inclusion of its own poison exon, as well as affecting the splicing of other genes. This creates a feedback loop in which increased levels of the functional Tra2β protein block further protein production. Interestingly, the Tra2b poison exon is defined as being ultra-conserved, as it shows 100% sequence identity between human, mouse, and rat, while also retaining 96% identity across 300 million years of evolution between human, chicken, and lizard! This not only suggests that the mouse can provide a useful model for studying the function of this poison exon but also underscores its likely biological importance.
To support his group’s investigation of the role of this poison exon, David Elliott, Professor of Genetics at Newcastle University, applied to the Mary Lyon Centre’s second GEMM call in 2017 to have a mouse generated with the Tra2b poison exon flanked by LoxP sites, using CRISPR/Cas9. This creates a mouse line that, when crossed with different Cre-expressing mice, allows the removal of the poison exon with precise spatial and/or temporal control.
In their new paper, published in the EMBO Journal, David’s group has investigated the role of the Tra2b poison exon in male fertility, prompted by the observation of particularly high levels of its inclusion in the mouse testis. By crossing their GEMM programme-generated line with Vasa-Cre mice to specifically delete the poison exon in embryonic germ cells (cells in the developmental pathway that leads to the production of sperm and egg cells), they found that this exon is essential for male fertility. Importantly, spermatogenesis cannot currently be studied using cell culture approaches, so animal models provide a useful system for increasing our understanding of fertility.
In addition to finding that the Tra2b poison exon is essential for meiotic cell division, the form of cell division used to produce sperm and egg cells, David’s group found that gene expression in cells in which the poison exon was deleted was significantly altered, with 300 genes upregulated and 203 downregulated. Several of those downregulated included genes important in chromosome biology and male fertility, while Tra2b itself was amongst those upregulated due to the loss of the feedback loop provided by the poison exon. Furthermore, they found 157 genes with significant splicing differences.
This work demonstrates that the Tra2b poison exon is essential for sperm production and male fertility. Interestingly, Tra2β is also known to have roles in wider embryonic development, including in the brain, and has been found to be misregulated in cancer, with protein levels increased particularly in cancers of the lung and breast. This means that the conditional Tra2b poison exon allele generated by the GEMM programme could be a useful tool for further investigations in which the poison exon is deleted in different cell types, potentially benefiting our understanding of brain development or identifying new therapeutic targets in lung or breast cancer.
This line and all others generated through the GEMM programme are available through the National Mouse Archive, housed here at MRC Harwell, and through the European Mouse Mutant Archive (EMMA).
