Direct measurement of the male germline mutation rate using sequential sperm samples

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PROJECT SUMMARY/ABSTRACT Most new genetic mutations in humans arise in the male germline. While most such de novo mutations are benign, collectively they drive human evolution and are responsible for an immense burden of disease— contributing to disorders such as autism, intellectual disability, and a wide range of developmental disorders. But remarkably, the rate at which mutations accumulate in the male germline, one of the most fundamental processes in human genetics, has never been directly measured. Indirect estimates of germline mutation rates have been based on family studies that identify de novo mutations in children that are not detected in their parents’ blood. These studies have shown that approximately 80% of de novo mutations arise in the male germline, and moreover, that the number of mutations transmitted by fathers via their sperm increases with paternal age. However, these ”family”-based measurements are limited by the number of children borne by any single individual, so they are only able to estimate the average mutation rate across a large number of individuals. They are also blind to mutations in sperm incapable of fertilization or viable pregnancies. Reliance on family-based measurements has therefore precluded quantification of how germline mutations accumulate with paternal age at the level of single individuals, the degree to which germline mutation rates vary across the population, and discovery of genetic and environmental factors that modify and control mutation rates. We hypothesize that directly sequencing sequential sperm samples of single individuals will define individual germline mutation rates and their inter-individual variability. In this project, we will develop a novel methodology to directly quantify, for the first time, the male germline mutation rate at the level of individuals and its variability among individuals. Aim 1 will develop a novel DNA sequencing technology (HiDEF-seq) for ultra-high fidelity detection of mosaic mutations at significantly lower cost than current methods. Aim 2 will then use HiDEF-seq to measure germline mutation rates in a novel approach whereby we will obtain sperm samples collected > 10 years apart from the same individuals in a collaboration with two of the largest sperm banks in the world. We will also develop data collection and analytic methods for use in future larger studies to define the relationships between measured mutation rates and genetic factors (e.g., variants in DNA repair/replication genes), health history, environmental exposures, fertility, and sperm quality. This work will provide a novel, scalable platform to directly measure the male germline mutation rate and, in future larger cohorts, to discover its genetic and environmental modifiers, all fundamental unknowns in human genetics. Answering these questions will provide key insights into the relationships between mutation, aging, health, and disease. This work will also provide an approach for developing germline mutation rate as a potential novel biomarker for aging, offspring disease risk, and fertility.