Periodic Reporting for period 3 - XChromosome (Functions of the X chromosome in the mammalian germ line)
Reporting period: 2018-11-01 to 2020-04-30
Infertility is common in humans, affecting one in seven couples. In at least one quarter of cases, the cause cannot be found. The sex chromosomes are a special pair of chromosomes that differ between men (XY) and women (XX), and that are thought to be particularly important for making the sex cells, i.e. sperm and eggs. Historically, a lot of research on infertility has focused on the Y chromosome, and it has been shown that mutations of Y chromosome genes are a common cause of male infertility. However, in comparison, the X chromosome has been largely overlooked. In this project, we aim to study the functions of X chromosome genes in the formation of the male and female gonads and in the formation of sperm. The principle technique we are using for this is CRISPR genome editing, a novel way in which the sequence of genes can be changed in a rapid and inexpensive manner. For our experiments, we use both mice, which have sex chromosomes similar to humans, and a marsupial, the opossum, which represents a more distant cousin of humans but that is very useful to understand how the sex chromosomes evolved. Our overall aim is to address whether genetic problems affecting the X chromosome could be responsible for infertility in patients for which a cause cannot currently be found. The findings will be of great importance to the treatment of infertility and will help us better understand why the sex chromosomes are so important for the formation of sperm and eggs.
Work performed from the beginning of the project to the end of the period covered by the report and main results achieved so far
So far, our work has focused on setting up the CRISPR genome editing technique for both mice and opossums. The mutations that we need to make in mice are quite complex, involving deletions of many genes in one go, and this is not straightforward. To overcome this issue, we are refining our experiments in mouse embryonic stem cells, a special cell that can be grown in a dish and that can later be used to make live mice. Genome editing has never been performed in an opossum, or indeed in any marsupial, so this is even more challenging. We have however succeeded in several of the steps required to set this technique up. For example, we can successfully perform editing in opossum embryos cultured in a dish. We are now at a stage where we need to be able to transplant these embryos back into the uterus of a mother, in order for the embryos to develop to live offspring that we can study. This will require further experimental refinements that are the subject of the next period of the grant.
Progress beyond the state of the art and expected potential impact (including the socio-economic impact and the wider societal implications of the project so far)
The main way in which our work so far has gone beyond the state-of-the-art is in the successful editing of a marsupial genome. Marsupials form the second largest class of mammals, second to the common “placental” mammals, e.g. humans, mice. They are a excellent model system for studying lots of areas of biology, including spinal cord regeneration, immunity to infections and skin cancer. However, to fully realise the potential of marsupials as a model system, we need to be able to introduce genetic modifications into them. This project aims to create such a genetic modification system using CRISPR genome editing. If successful, the project could benefit lots of areas of human biology, including understanding nervous diseases and cancer.