Positive Selection for New Disease Mutations in the Human Germline: Evidence from the Heritable Cancer Syndrome Multiple Endocrine Neoplasia Type 2B

Multiple endocrine neoplasia type 2B (MEN2B) is a highly aggressive thyroid cancer syndrome. Since almost all sporadic cases are caused by the same nucleotide substitution in the RET proto-oncogene, the calculated disease incidence is 100–200 times greater than would be expected based on the genome average mutation frequency. In order to determine whether this increased incidence is due to an elevated mutation rate at this position (true mutation hot spot) or a selective advantage conferred on mutated spermatogonial stem cells, we studied the spatial distribution of the mutation in 14 human testes. In donors aged 36–68, mutations were clustered with small regions of each testis having mutation frequencies several orders of magnitude greater than the rest of the testis. In donors aged 19–23 mutations were almost non-existent, demonstrating that clusters in middle-aged donors grew during adulthood. Computational analysis showed that germline selection is the only plausible explanation. Testes of men aged 75–80 were heterogeneous with some like middle-aged and others like younger testes. Incorporating data on age-dependent death of spermatogonial stem cells explains the results from all age groups. Germline selection also explains MEN2B's male mutation bias and paternal age effect. Our discovery focuses attention on MEN2B as a model for understanding the genetic and biochemical basis of germline selection. Since RET function in mouse spermatogonial stem cells has been extensively studied, we are able to suggest that the MEN2B mutation provides a selective advantage by altering the PI3K/AKT and SFK signaling pathways. Mutations that are preferred in the germline but reduce the fitness of offspring increase the population's mutational load. Our approach is useful for studying other disease mutations with similar characteristics and could uncover additional germline selection pathways or identify true mutation hot spots. Author Summary: Multiple endocrine neoplasia type 2B (MEN2B) is a highly aggressive thyroid cancer syndrome. MEN2B offspring with unaffected parents almost always received a new mutation from the father. Moreover, this mutation is almost always at the same nucleotide in the RET proto-oncogene. Thus MEN2B's incidence should equal the average single nucleotide mutation frequency, but the observed incidence is 100–200 times greater. One explanation is that the mutation rate at the causal nucleotide is significantly elevated above the genome average. Another is that human testis stem cells acquiring this mutation have a selective advantage over non-mutated ones and this advantage increases the mutation's frequency in the testis. Computational analysis of our testis dissection and mutation assay data rejects the hot spot but not the selective advantage explanation. Because the normal RET gene is known to be critical for mouse testis stem cell function, we now have an important insight into what biochemical pathways are altered by the MEN2B mutation to provide this selective advantage in humans. Germline selection explains the unexpectedly high incidence of MEN2B, why the mutation's origin is almost always in the father, and why the probability a child is born with this disease increases with the father's age.