Introduction
Genetic variation is the raw material for natural selection to act upon. Hence, there is a long history of evolutionary studies on how genetic variation is maintained in natural populations (Dobzhansky, 1982), either invoking selection or neutral processes. One of the most dramatic cases of genetic variation concerns polymorphism at Major Histocompatibility Complex (MHC) genes, which are commonly the most variable genes in vertebrate genomes (Sommer, 2005). Surprisingly, despite decades of research on MHC evolution, the evolutionary processes sustaining MHC polymorphism remain unclear, with inconsistent and often ambiguous support for competing hypotheses (Radwan, Babik, Kaufman, Lenz, & Winternitz, 2020). Here, we evaluate predictions of multiple competing MHC evolution models (adaptive and neutral), using an exceptionally large dataset of genetic variation and macroparasite infection in a metapopulation of threespine stickleback (Gasterosteus aculeatus ) inhabiting lakes on Vancouver Island.
MHC genes play a key role in the adaptive immune system of vertebrates. The main function of these genes is to encode for cell surface proteins, which are used to detect foreign molecules. These molecules are then presented to T cells to initiate appropriate immune responses. The MHC gene family consists of two classes of genes, MHC I and MHC II, each of which can be represented by multiple paralog copies. MHC class I genes are expressed in all nucleated cells and typically bind to peptides derived from intracellular molecules, such as virus protein (Jensen, 2007). MHC class II genes (our focus here) are expressed only in antigen presenting cells, such as macrophages, dendritic cells, and B cells. These cells phagocytize extracellular material, digest the proteins, and if the MHC II proteins bind to the resulting fragments, these are presented on the cell surface to T cells, to possibly initiate an adaptive immune response (Jensen, 2007). The second exon of MHC II gene is often used to characterize the variability of MHC II genes, because it contains the antigen-binding domain (Sommer, 2005).
The maintenance of MHC diversity is often attributed to pathogen-mediated selection, although intraspecific processes, such as mate choice (Penn & Potts, 1999), are also potential sources of selection. As reviewed in Spurgin & Richardson (2010), there are three non-mutually exclusive pathogen-mediated selection mechanisms to explain MHC diversity: (1). Heterozygote advantage . Individuals with more MHC alleles might be able to recognize a more diverse set of parasite species, thus have fewer parasites. This advantage may lead to directional selection for ever-increasing diversity, but some widely-cited studies have presented evidence for intermediate optima (K. Mathias Wegner, Kalbe, Kurtz, Reusch, & Milinski, 2003). (2).Negative frequency-dependent selection . In a given population, common MHC alleles that protect against parasites impose selection on those parasites to change their antigens and evade recognition. Once parasites evolve strategies to evade these common alleles, individuals carrying the common alleles may be at a selective disadvantage. By contrast, a rare MHC allele imposes little selection on the parasites it recognizes (because it is rare), and so may tend to be more protective. For this negative frequency-dependence to work in the long run, it must on average hold that rare alleles are more protective (and hence more fit) than common alleles. (3) Fluctuating selection . Parasite community and parasite genotypes may vary in space and time. Given this, different host MHC alleles may be selectively favored in different locations and at different times. Gene flow between locations subject to divergent selection can sustain MHC diversity, as can temporally fluctuating selection.
Although many studies have examined the roles of the above-mentioned mechanisms in maintaining MHC diversity, the results are often mixed or even contradictory. For example, heterozygote advantage was supported in one experimental infection study with inbred mice (Penn, Damjanovich, & Potts, 2002), but the effect was not found in another study with outbred mice (Ilmonen et al., 2007). Theoretically, negative frequency dependent selection is more likely to explain the extraordinary diversity in MHC genes than heterozygote advantage (Borghans, Beltman, & De Boer, 2004). However, empirically demonstrating negative frequency dependent selection is more challenging, as the pattern is often also consistent with other mechanisms. For example, in house sparrows, population-specific MHC alleles were linked to host resistance or susceptibility to malaria (Bonneaud, Pérez‐Tris, Federici, Chastel, & Sorci, 2006), but this pattern could be explained either by negative frequency dependent selection or spatially varying selection. A previous study of stickleback from three lake-stream population pairs (Stutz & Bolnick, 2017) tried to distinguish between negative frequency dependent selection or spatially varying selection. The study took advantage of migration between parapatric populations with different parasite communities, which in principle would allow them to partition benefits of rarity within populations, from costs of being a (rare) immigrant to a foreign habitat. Yet, no overall trend was found to support either mechanism. Such inconsistent or ambiguous results are typical in the MHC evolution literature. Therefore, a consensus regarding the relative importance of these mechanisms in maintaining MHC diversity has not been reached (Radwan et al., 2020).
In contrast to these adaptive hypotheses, whether and how MHC diversity is shaped by neutral processes is less studied. In small and/or isolated populations, MHC diversity is often heavily influenced by genetic drift, rather than balancing selection (Hedrick, Parker, & Lee, 2001; Miller & Lambert, 2004; Seddon & Ellegren, 2004). In large metapopulations, it is unclear what role neutral processes play in shaping MHC diversity.
MHC studies in threespine stickleback have generated some unique insights in the past (Eizaguirre, Lenz, Kalbe, & Milinski, 2012; Matthews, Harmon, M’Gonigle, Marchinko, & Schaschl, 2010; Mccairns, Bourget, & Bernatchez, 2011; Milinski et al., 2005; K. Mathias Wegner et al., 2003). For example, Wegner et al. (2003) conducted a study on 8 stickleback populations to examine the association between MHC diversity and 15 parasite species. They found populations exposed to a wider range of parasites had higher MHC allelic diversity, though MHC diversity also positively, albeit weakly, correlated with neutral diversity. Moreover, at the individual level, parasite burden was minimized in fish with an intermediate, rather than maximal, number of MHCIIβ alleles. This result supports the theoretical model of stabilizing selection on MHC heterogeneity (Nowak, Tarczy-Hornoch, & Austyn, 1992). The logic is that low MHC-diversity individuals do not have the capacity to recognize diverse parasites, but high MHC-diversity individuals deplete their T cells via negative selection on self-reactive T cells. The resulting paucity of T cell receptor diversity also limits their ability to recognize diverse parasites. However, this intermediate-advantage hypothesis, while widely cited, has not been replicated in many systems and remains controversial (Borghans, Noest, & De Boer, 2003). Another example of interesting insights from stickleback is the involvement of MHC alleles in mate recognition. It was found that female stickleback could choose their mate based on odor to optimize the number of MHC alleles in their offspring, though the molecular mechanism is unknown (Milinski et al, 2005).
In this study, we set out to test predictions associated with three main models of MHC polymorphism invoking parasite-mediated selection, using an extensive field survey of parasites in a metapopulation of threespine stickleback. We did not find evidence for stabilizing or positive selection for MHC heterozygosity, contrary to Wegner et al. (2003). Nor did we observe fluctuating selection across populations. Although we found significant associations between specific MHC alleles and parasite species both within and across populations, neutral processes in our dataset best explained within- and between-population MHC diversity in our data set.