Functions of the X chromosome in the mammalian germ line

Project Information

XChromosome

Grant agreement ID: 647971

DOI

10.3030/647971

Project closed

EC signature date 26 August 2015

Start date 1 November 2015

End date 30 April 2022

Funded under

• EXCELLENT SCIENCE - European Research Council (ERC)

Total cost

€ 1 912 369,00

EU contribution

€ 1 912 369,00

Coordinated by

THE FRANCIS CRICK INSTITUTE LIMITED
United Kingdom

Periodic Reporting for period 5 - XChromosome (Functions of the X chromosome in the mammalian germ line)

Reporting period: 2021-05-01 to 2022-04-30

Summary of the context and overall objectives of the project

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

Thanks to funding from this project, we have succeeded in setting up CRISPR genome editing in marsupials. Using this approach, we have demonstrated that the long non-coding RNA RSX is essential for the initiation of X chromosome inactivation in marsupials. This is important because it sets the stage for RSX to be used as a new paradigm for epigenetics. We have also shown that RSX is dispensable for the maintenance of X-inactivation. This situation is exactly what is observed in mice, and indicates that in marsupials other epigenetic modifications, possibly polycomb proteins, must stabilise the inactivated X after RSX has initiated silencing. We have also initiated genome editing of the RSX antisense RNA XSR, to investigate its role in imprinted X chromosome inactivation in marsupials. As an aside, our work has also demonstrated that genome editing in marsupials leads to an increased incidence of embryonic arrest, most likely due to the creation of large chromosome deletions resulting from faulty repair. These findings mirror those shown by other groups in humans, and suggest that error-prone DNA repair is a shared feature of marsupial and eutherian mammalian embryos. This finding is important when considering the pros and cons of DSB-based genome experiments in embryos.

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)

Our study has progressed beyond the state-of-the-art, by succeeding in optimising genome editing in marsupials. 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, infections and skin cancer. Using our technology, the international research community will now finally be able to carry out functional, in vivo studies to investigate these diverse research areas.

Role of the X chromosome in male fertility in mice (left) and opossums (right)