Functional dissection of Tmem95: a sperm cell surface protein essential for mammalian fertilization

在人类中，新生命始于受精，当精子和卵子相互识别时，它们周围的膜融合形成一个新的胚胎。精子和卵子利用细胞表面识别分子相互作用，这些分子特异性结合并使它们融合在一起。虽然受精是一个基本的生物学过程，但我们对所涉及的分子的理解非常差，特别是在哺乳动物中;例如，我们不知道负责将精子和卵子融合在一起的分子。这些知识差距可以部分解释人类由于受精的伦理问题，以及研究细胞表面分子的困难。例如，卵子是一种非常罕见的细胞类型，人类通常在每个生育周期中只释放一个卵子，这限制了可用的生物材料的数量。此外，已知细胞表面分子之间的物理相互作用非常弱（半衰期仅为几秒钟），需要使用专门的方法来检测它们。桑格研究所的细胞表面信号实验室专门研究这些短暂的相互作用，我们已经开发了一套技术来规避这些困难。我们之前已经使用这些技术鉴定了第一个必需的精卵受体对（参见比安奇等人，Nature 2014 v508 p483），这对受精的分子理解做出了重要贡献。我们对一种新的精子表面蛋白感到兴奋，该蛋白被命名为Tmem 95（transmembrane蛋白编号95），它对精子与卵子融合的能力至关重要。Tmem 95是从养牛业的研究中鉴定出来的，其中研究了一例特发性男性不育症（尽管看起来和运动正常，但精子不育）。研究人员使用了数千次对奶牛进行人工授精的结果，并通过仔细记录，确定了TMEM 95基因突变的来源。进一步的研究表明，在Tmem 95基因中有工程突变的雄性小鼠也是不育的，因为正常的精子在IVF试验中无法与卵子融合。人类也有一个TMEM 95基因，它看起来非常相似，我们相信它也会在人类中发挥重要作用;然而，我们对这种蛋白质知之甚少。这项资助申请旨在更多地了解TMEM 95在施肥中的重要作用。具体来说，我们将使用我们的专业技术来确定它与卵子表面受体相互作用的分子身份，描述蛋白质在受精过程中如何在精子中重新分布，确定Tmem 95是否在膜融合中起特定作用，并研究它如何与另一种称为Izumo 1的精子细胞表面蛋白相互作用。这些研究将进一步加深我们对精子细胞表面蛋白的分子理解。基本的生物过程，并提供了一个基础，以开发更好的避孕和生育治疗都在人类和农业。人口的迅速增长（目前超过70亿，预计到2050年将达到100亿）引起了人们的担忧，即地球上有限的资源将无法维持持续的扩张。奇怪的是，不孕症是一个日益严重的问题，特别是在西方国家，近年来夫妇生育第一个孩子的平均年龄有所增加。通过发现新的不育基因，这项研究可以为不育夫妇提供简单而廉价的基因筛查测试，这可能会指导他们的生育治疗，并节省试管婴儿失败的费用和痛苦。这些研究还可以改善畜牧业的生殖技术，并开发避孕疫苗，以控制野生动物种群，并以道德上更可接受的方式对伴侣动物进行绝育，而不是扑杀或绝育。

In humans, new life starts at fertilization when a sperm and an egg recognise each other and their surrounding membranes fuse to form a new embryo. Sperm and egg interact using cell surface recognition molecules which bind specifically and enable them to fuse together. Although fertilization is a fundamental biological process, our understanding of the molecules involved is remarkably poor, especially in mammals; for example, we do not know the molecule/s that are responsible for fusing the sperm and egg together. These knowledge gaps can be partly explained in humans due to the ethical issues surrounding fertilization, and the difficulties in studying cell surface molecules. For example, eggs are a very rare cell type, with humans usually releasing just a single egg in each fertility cycle limiting the amount of biological material available. In addition, the physical interactions between cell surface molecules are known to be very weak (having half-lives of just seconds), requiring the use of specialised methods to detect them. The Cell Surface Signalling Laboratory at the Sanger Institute specializes in identifying these fleeting interactions and we have developed a set of techniques to circumvent these difficulties. We have previously used these techniques to identify the first essential sperm-egg receptor pair (see Bianchi et al. Nature 2014 v508 p483), which made an important contribution to the molecular understanding of fertilization.We are excited about a new sperm surface protein prosaically named Tmem95 (for TransMEMbrane protein number 95) that is essential for the ability of sperm to fuse with eggs. Tmem95 was identified from research in the cattle industry where a case of idiopathic male infertility (sperm that is infertile despite looking and moving normally) was investigated. Researchers used the results of thousands of artificial inseminations performed in cows, and by careful record-keeping, identified the source to a mutation in the TMEM95 gene. Further research has shown that male mice which have an engineered mutation in their Tmem95 gene are also infertile because normal-looking sperm are unable to fuse with eggs in IVF assays. Humans also have a TMEM95 gene which looks very similar and we believe that it will have an important role in humans too; however, we know very little about this protein. This grant application is aimed at understanding more about the important role TMEM95 has in fertilization. Specifically, we will use our specialized technologies to determine the molecular identity the receptor it interacts with on the egg surface, describe how the protein is redistributed in the sperm during fertilization, establish if Tmem95 has a specific role in fusing membranes together, and investigate how it interacts with another essential sperm cell surface proteins called Izumo1.These studies will both further our molecular understanding of a fundamental biological process, and provide a foundation to develop better contraceptives and fertility treatments both in humans and in the farming industry. The rapidly expanding human population (currently over 7 billion and predicted to reach 10 billion by 2050) has raised concerns that the limited resources on the planet will not sustain a continued expansion. Paradoxically, infertility is a growing problem, particularly in Western countries where the average age of couples having their first child has increased in recent years. By discovering new infertility genes, this research could open up the possibility of offering simple and inexpensive genetic screening tests to infertile couples that may guide their fertility treatment and save the expense and pain of failed rounds of IVF. These studies could also improve reproduction technologies in the livestock industry and also in the development of contraceptive vaccines to control wild animal populations and sterilize companion animals in an ethically more acceptable way other than culling or neutering.