PDF(Pubmed)
Abstract：
CRISPR-Cas9 technology has facilitated development of strategies that can potentially provide more humane and effective methods to control invasive vertebrate species, such as mice. One promising strategy is X chromosome shredding which aims to bias offspring towards males, resulting in a gradual and unsustainable decline of females. This method has been explored in insects with encouraging results. Here, we investigated this strategy in Mus musculus by targeting repeat DNA sequences on the X chromosome with the aim of inducing sufficient DNA damage to specifically eliminate X chromosome-bearing sperm during gametogenesis. We tested three different guide RNAs (gRNAs) targeting different repeats on the X chromosome, together with three male germline-specific promoters for inducing Cas9 expression at different stages of spermatogenesis. A modest bias towards mature Y-bearing sperm was detected in some transgenic males, although this did not translate into significant male-biasing of offspring. Instead, cleavage of the X chromosome during meiosis typically resulted in a spermatogenic block, manifest as small testes volume, empty tubules, low sperm concentration, and sub/infertility. Our study highlights the importance of controlling the timing of CRISPR-Cas9 activity during mammalian spermatogenesis and the sensitivity of spermatocytes to X chromosome disruption.
摘要：
CRISPR-Cas9技术促进了战略的发展，这些战略可能提供更人性化和有效的方法来控制入侵脊椎动物物种。比如老鼠。一种有希望的策略是X染色体切碎，旨在使后代偏向雄性，导致女性逐渐和不可持续的衰落。已在昆虫中探索了这种方法，并取得了令人鼓舞的结果。这里,我们通过靶向X染色体上的重复DNA序列，在小家鼠中研究了这种策略，目的是诱导足够的DNA损伤，以在配子发生过程中特异性消除带有X染色体的精子。我们测试了三种不同的引导RNA（gRNA），靶向X染色体上的不同重复序列，与三个雄性种系特异性启动子一起在精子发生的不同阶段诱导Cas9表达。在一些转基因雄性中检测到对成熟Y精子的适度偏见，尽管这并没有转化为后代的显著男性偏见。相反，减数分裂期间X染色体的裂解通常会导致生精阻滞，表现为小睾丸体积，空小管,低精子浓度，和亚/不孕症。我们的研究强调了在哺乳动物精子发生过程中控制CRISPR-Cas9活性的时机和精母细胞对X染色体破坏的敏感性的重要性。
