Discussion:
Our results provide one of the very few human examples (
47) where the effects of a common genetic variant are sex and environment dependent. In that regard, we suggest that
GHRd3 has important ramifications for metabolic disorders, such as obesity and diabetes, but only for males within particular environmental contexts. For example, it was reported that
GHRd3 has a preventative impact on type 2 diabetes. However, in the small number of diabetes patients who are homozygous for
GHRd3, a significant increase in further metabolic complications was observed (
26). This parallels our observation that a significant size difference was observed between wt/wt and d3/d3 mice only under calorie restriction for males. Similarly, we expect the reported effects on birth and placental size, time to menarche, and longevity to vary considerable for different dietary and endocrinological contexts. Following this thread, we expect that
GHRd3 may have other important roles that would be visible only when specific sexes, environmental conditions, and developmental stages are investigated. Our study has characterized the effect of homozygous
Ghrd3. Given that this gene codes for a protein that self-dimerizes for it to function properly, the functional impact of
Ghrd3 in heterozygous individuals remains a fascinating question. Previous studies suggest that the heterozygous
GHRd3 may contribute to different traits in an additive, recessive, or dominant manner. Our mouse model makes it possible to test all these different perspectives in future studies.
The sex-specific effects of
GHRd3 raise additional evolutionary considerations, especially because the GH pathway is a major driver of sexual dimorphism in mammals (
48). Moreover, multiple genes in this pathway were recently implicated in sexually antagonistic transmission distortions in humans (
49). In this regard,
GHRd3 provides an interesting case. The traditional understanding of the evolution of sex-specific effects of genetic variation first considers sexual conflict where the functional effect of the variant leads to increased fitness in one sex but decreased fitness in the other (
50). This conflict leads to maintenance of the variation in the population through balancing selection until the conflict is resolved by additional genetic variation that moderates the effect of the variant in a sex-specific manner.
GHRd3 does not fit this scenario. Instead, the sex-specific effect of
GHRd3 is instantaneous because the GH pathway is already operating in a sex-specific manner. This observation raises questions about the extent of the evolutionary effects of genetic variation in pathways that already operate in a sex-specific manner.
The evolutionary history of
GHRd3 is complex. Our model suggests that geography-specific adaptive pressures over time have shaped the allele frequency of
GHRd3 across human evolutionary history, effectively maintaining this variation in human populations over 700 ka. In this regard,
GHRd3 evolutionary history fits a very broad definition of balancing selection, i.e., maintenance of advantageous variation over long periods of time, overcoming fixation due to varying strengths of selective advantage through time, including drift and potentially negative selection. However, the signatures of
GHRd3’s evolutionary history do not fit the expectations of more narrowly defined balancing selection models, such as heterozygous advantage or frequency-dependent selection (
51). Instead,
GHRd3 evidences the existence of loci under selective pressures that do not appear to conform with classical sweep or balancing selection models, and future work should investigate the prevalence of such loci across the genome (
52–
57).
Our insights into this genetic variation at the
GHR locus bring forth additional questions concerning recent human evolution. The initial allele frequency increase in
GHRd3 across evolutionary time coincides with unstable environments that mark recent hominin evolution (
58). Given its enhanced effect under calorie restriction, it is plausible that
GHRd3 provided a fitness advantage to early hominins, perhaps facilitating adaptation to new environments during
Homo migrations out of Africa starting ~1 million years ago. Our finding that
Ghrd3 is protective against the severe consequences of malnutrition in male children supports this hypothesis. The sex-specific nature of this adaptation may be better understood by considering times of nutritional stress and assuming that the increased size in males is a derived trait. During these periods, survival may be a stronger adaptive force than the fitness benefits of increased size, causing smaller sizes in males to be favored (
28). Thus,
GHRd3 may be favored under environmental stress because it increases survival although it reduces sexual dimorphism and thus reduces competitiveness for mate choice among males.
The marked reduction in
GHRd3 allele frequencies coincides with the emergence of technologically advanced material culture, such as bone tools, fish hooks, and composite weapons, along with a concurrent population expansion, seemingly occurring independently in different parts of the world between 90 and 30 ka ago (
59). Emerging technological innovations that may have allowed these populations to adapt to diverse environments (
60) may also have acted as a buffer against the effects of fluctuating resource levels, thereby changing the selective pressures acting on the
GHR locus. Note that the reduction in
GHRd3 allele frequency coincides with major demographic and ecological changes including the Last Glacial Maximum (
61). Thus, the combined effect of cultural and climate change could explain the rapid adaptive decrease in
GHRd3 allele frequencies in all human populations, the most marked of which was observed in East Asia.
