Gene Interactions and
t h e Origin of Species
NORMAN A . JOHNSON
Many authors (e.g., Mayr 1991; Coyne 1992; Dennett 1995; O r r 1995)
have noted that, despite the title of his book, Darwin (1859) did not solve the
origin of species. That problem was solved in outline by Dobzhansky, Mayr,
Muller, and the other architects of the modern synthesis. In contrast to Darwin,
they saw biodiversity being broken into "a multitude of separate, discrete,
distributions" (Dobzhansky 1937, p. 4), with species as the fundamental, discontinuous units of this diversity. This view of species is inherent in the biological
species concept (BSC), wherein species are defined both by their ability to
interbreed inter se and by their reproductive isolation from other such groups
(Dobzhansky 1935; Mayr 1942). While many other species concepts have been put
forth since (for a sample, see Table 1 of Coyne 1994), the BSC remains one of the
most widely used (Coyne 1992, 1994; Orr 1995). Moreover, many evolutionary
biologists who do not support the BSC still recognize the importance of
reproductive isolation in the creation of biodiversity (e.g., Templeton 1989; Frost
and Hillis 1990). In a recent attempt to forge a conciliation between the BSC and
phylogenetic species concepts, Avise and Wollenberg (1997, p. 7754) argue that
"to cleanse from species concepts all references to reproductive isolation would be
to leave an unduly sterile epistemological foundation for the origin and
maintenance of the biotic discontinuities so evident to Dobzhansky 60 years ago.'
The reproductive barriers to gene flow are diverse but can be placed into two
general categories according to when they occur: prezygotic barriers reduce the
probability of the formation of hybrid zygotes, and postzygotic barriers reduce the
fitness of those hybrid zygotes (Dobzhansky 1937; Mayr 1963; Futuyma 1998). In
this chapter, I will discuss the relationship between genetic interactions and
198 Genetic Differentiation: From Populations to Species
Gene Interactions and the Origin of Species 199
postzygotic reproductive isolation. With few exceptions, I will not discuss
prezygotic reproductive isolation, as this subject is discussed in Meffert's chapter
(chap. 11) in this volume. I will also not explicitly discuss hybrid zones; this subject
is discussed by Gardner et al, (chap. 16) in this volume.
The Dobzhansky-Muller Model
Dobzhansky (1937) and Muller (1940, 1942) each recognized the importance of
gene interaction in the evolution of postzygotic reproductive isolation (also called
hybrid fitness reduction). In the allopatric divergence model of speciation, microevolutionary forces (mutation, random genetic drift, and natural selection) act
upon geographically isolated populations. Without the homogenizing force of gene
flow, these populations will diverge genetically. With increasing divergence, the
genomes of the two nascent species become increasingly incompatible with each
other. If the geographic barriers were removed to allow for the production of
hybrids between diverged populations, the hybrids would possess combinations of
genes that do not work well together (i.e., are incompatible). These genetic
incompatibilities would result in the hybrids being (partially or completely) inviable
or sterile and thus would produce reproductive isolation.
E.
Fitnesses of genotypes
Figure 12.1
Figure 12.1 The Dobzhansky-Muller mode1 of evolution of postzygotic reproductive
isolation. Assume that A and B represent different loci, each with two alleles (subscripted 0
and l), and that individuals with any combination of A l and Bl have greatly reduced
fitness. (A) Stage I: the population is fixed for alleles An and Bo. ( B ) Stage 11: a geographic
barrier splits the population and gene flow ceases. (C) Stage 111: the variant Al appears in
the population on the left and the variant Bl appears in the population on the right. (D)
Stage IV: the Al allele becomes fixed in the left population and Bl becomes fixed in the
right population. (E) Hybrids from a cross between the left and right populations would be
AoA,BoB, and would thus have reduced fitness. Note that neither population passes
through an adaptive valley.
(continued)
Nonadditive interactions of variants at different loci (epistasis) are presumed to
be the genetic basis for these incompatibilities (Fig. 12.1). In principle, the simpler
case of single-locus underdominance could lead to incompatibility if populations
could fix different homozygotes at each locus (hybrids would be heterozygotes and
thus less fit). This underdominance model, however, is very unlikely due the
exceedingly low probabilities of fixing different homozygotes in the different
populations. In order for this to occur, one or both of the populations would have
to go through an adaptive valley (where less-fit heterozygotes were transiently
common). In contrast, reproductive isolation could occur without the need for
either population to cross an adaptive valley if the incompatibility were caused by
alleles of at least two loci that interact epistastically (Dobzhansky 1937; Mayr 1942;
Walsh 1982; Barton 1989b; Orr 1991; Bateson in Orr 1996). Thus, in the allopatric
divergence model, the evolution of hybrid incompatibility does not require the
parental populations to transverse adaptive valleys, and reduces to the formation
of incompatible sets of loci (Orr 1995).
Although alternatives to this model have been proposed (e.g., White 1978; Diehl
and Bush 1989), evolutionary biologists widely support the allopatric divergence
model (Futuyma and Mayer 1980; Coyne 1992; Wu and Palopoli 1994; Futuyma
Gene interactions and the Origin of Species 20
Box 12.1 (continued)
Box 12.1 Allelic effects in negative and positive phenotypes
MIelic variants at "hybrid incompatibility loci" have t w o distinct phenotypic effects: one
~ p o the
n fitness and othertraits of hybrids (the negative ~henotype),and one upon the fitless and other traits of the pure species individuals that possess it (the positive phenotype)
[see Fig. B12.1).
If populations are diverging completely in allopatry, only pure species individuals (and
not hybrids)will becreated ineachgeneration. Hence, evolutionaryforces(naturalselection,
random genetic drift) will operate upon only the positive phenotype and not the negative
phenotype. Incontrast.geneticstudiesofhybrid incompatibility, by necessity, beginwiththe
negative phenotype (the hybrid effects). Forthe most part,thesestudies have not examined
the positive phenotype. Thus, the evolutionary dynamics ofthe hybrid incompatibility loci
arenotwell known.Thechallengeforspeciationgeneticresearcherswillbetounitethesetwo
realms.JohnsonandWade's(1996)model.whichconsiderstheeffectsofasingleallelicsubstitution on purespeciesand hybrid genetic backgrounds, isthe first-steptoward aconceptual
framework of this unification. This model, however, is still an oversimplification of the
complexity involved (seetext).
Conspecific (Positive) Phenotypes
-
Hybrid (Negative) Phenotypes
Fig. 612.1
(continued)
Figure B12.1 illustrates the challenges involved in the unification of these t w o realms.
Here, genetic variants (middle panel) affect both the positive (pure species) phenotype
(top) and the negative (hybrid) phenotype (bottom). In the figure, G l o and GI b are allelic
variants at locus I, and G2a and G2b are allelic variants at locus 2. The traits C l and C a r e
expressed in the positive (pure species) genetic background, and traits HI and H; are
expressed in the negative (hybrid) genetic background. Relative t o a standard variant (not
shown),Glagreatly increases(assh0wnby ++) bothCl andC2, whileitdecreasesHl. Likewise. GI b decreasesCl, while it greatly increases HI. Notethat for locus I,the relationship
between effects on conspecifics and on hybrids is negative ( r <0, in Johnson and Wade's
model). For locus 2, this relationship is positive: G2a greatly increasesboth C2and H2.
Natural selection operates directly on Cl and C;,but only indirectly on H I and Hi.
Selection for the increase of Cl may result in an increase in frequency of Gla. If so, t h e n H I
should decrease. Another possibility is that selection for an increase of C j would result i n
a decrease in frequency of Glb. If so, HI would decrease, and so would H2. Selection that
alters C2 could result in changes in allelic frequency at either o r both loci, and could have
effectsoneither o r both H I and Hi.
Despite the apparent complexity of this figure, it greatly underrepresents t h e complexity of real systems. Many genes (often more than the t w o shown here) can affect a given
trait (either pure species or hybrid). Furthermore, this figure does not depict conspecific
epistasis.
1998). The controversy is not over whether speciation occurs via divergence in
allopatry, but rather to what extent reproductive isolation arises via other means. It
is also clear that, except for certain, quite restrictive conditions (Coyne 1974;
Leibowitz 1994; Johnson and Wade 1995), postzygotic reproductive isolation does
not arise as the result of direct selection (in contrast to the reinforcement model of
prezygotic reproductive isolation; Dobzhansky 1940; Howard 1993; Kelly and
Noor 1996), but only as a pleiotropic by-product of divergence.
Microevolutionary forces that operate within species, coupled with geographical
isolation, thus lead to the formation of intrinsic reproductive barriers and
eventually to biological discontinuities. One can consider the observed hybrid
fitness reduction to be a negative phenotypean epiphenomenon of the underlying
hybrid fitness reduction (Box 12.1). The divergence itself is likely to be
opportunistic. By this, I mean it is likely that many classes of "genes" (in the
broad sense) will be involved, and the relative importance of different types of
genetic systems that contribute to the divergence that underlies hybrid fitness
reduction will probably vary across taxa (Johnson 1998).
In the next two sections, I will present what we know about the genetic architecture of hybrid dysfunction. I will then discuss the consequences of the complexity of the genetic architecture of hybrid incompatibility for the Fisher-Wright
debate, and in general. Next, Iconsider the plausibility that hybrid fitnessreduction
is generated by the drift-induced fixation of intraspecific epistatically interacting
variants. In the final section, I discuss what types of future research will be most
beneficial to our understanding of the role that gene interaction plays in the
evolution of reproductive isolation, and the consequences of such interactions.
202 Genetic Differentiation: From Populations to Species
Detection of Gene Interactions that Underlie
Hybrid Fitness Reduction
Dobzhansky (1936) also pioneered the genetic analysis of hybrid traits. His
method, the backcross analysis, consists of crossing individuals of one species
marked with multiple genetic markers to individuals of a related species. If at
least one sex of the F l hybrids is fertile, then the F l can be backcrossed to one or
both of the parental species. The difficulty of examination of the genetic basis of
the inability to produce viable, fertile hybrids by the production of fertile hybrids
is at least partially alleviated by the large number of species in which one sex is
(at least partially) fertile and the other sex is sterile. Across a wide variety of taxa,
the vast majority of hybridizations with sex-biased asymmetry of hybrid fitness
reduction follow Haldane's (1922) rule, where the heterogametic sex (male in
Drosophila and mammals, female in birds and butterflies) is most severely
affected. Various explanations for Haldane's rule are discussed in the reviews of
Wu et al. (1996), Laurie (1997), and Orr (1997), and references therein. Genetic
analyses that stop after only one or two generations of backcrossing are often
limited in their ability to detect the locations of incompatibility genes, and the
nature of the interactions, because the genotypes of the hybrids are too complex
and ill-defined (Wu and Davis 1993; Wu and Palopoli 1994). With the
introgression method, a refinement of the backcross analysis, researchers have
dramatically increased the resolution with which the genetic architecture of
hybrid fitness reduction can be examined (e.g., Coyne and Charlesworth 1986;
Wu et al. 1993; Wu and Palopoli 1994; see also True et al. 1996). This method
allows for the production of a large number of genetically similar individuals,
each with a small and defined portion of the genome from one species in the
genetic background of another species. Various aspects of the phenotype (e.g.,
sperm motility, progeny production, mating ability) of these individuals then can
be assayed.
A major conclusion of the introgression studies in the Drosophila melanogaster
species group is that dozens-if not h u n d r e d s ~ ogenes,
f
when placed in the genetic
background of a closely related species, have severely deleterious effects on male
fertility (Wu and Palopoli 1994; Palopoli et al. 1996; True et al. 1996). The large
number of these so-called hybrid sterility genes demonstrates that, despite the lack
of morphological differences between these species, substantial functional
divergence has occurred (Wu and Palopoli 1994; Wu et al. 1996; Johnson and
Kliman, submitted).
The types
~.of interactions involved in hybrid fitness reduction are rather diverse,
but some basic patterns are apparent. First, X-autosome interactions appear to be
common (Dobzhansky 1936; Charlesworth et al. 1987; Coyne and Orr 1989a). It
has been previously thought that the X chromosome has larger effects on hybrid
fitness reduction than the autosomes (reviewed in Charlesworth et al. 1987; and
Coyne and Orr 1989a), but recent studies in Drosophila show that the density of
genes that contribute to hybrid sterility on the autosomes is comparable to that on
the X chromosome (Hollocher and Wu 1996; True et al. 1996). Thus, the "large X
effectnis probably an artifact of how the studies were performed. Analyses that stop
Gene Interactions and the Origin of Species 203
after the first generation of backcrossing will typically make heterozygous substitutions of the autosomes, but hemizygous substitutions of the X chromosome (see
Wu and Davis 1993 for further discussion).
The Y chromosome sometimes, but not always, contributes to the sterility of
hybrids between closely related species of Drosophila (see Johnson et al. 1993, and
references within). Negative interactions between heterospecific X and Y chromosomes is a simple hypothesis to explain Haldane's rule (Coyne 1985). There are
cases wherein hybrid dysfunction was thought to be due to X-Y interactions, but
further investigation showed that this was probably not the case. For instance, the
sterility of F l male hybrids between D. simulans and D. sechellia was more likely
due to a combination of X-autosome and Y-autosome interactions than due to
direct X-Y interactions (Johnson et al. 1992). Cointrogression studies, wherein
both the Y chromosomes and portions of different autosomes are introgressed at
the same time, provide strong direct evidence for Y-autosome interactions that
cause hybrid male sterility between closely related species in both the D. repleta
and D. virilis species groups (Pantazidis et al. 1993; Lamnissou et al. 1996).
The cytoplasm of nearly every cell in most metazoans includes numerous
mitochondria, each with their own genome. Divergence could lead to incompatibilities between the mitochondria1 genes (or other cytoplasmic factors) of one
species and the nuclear genes of the other. The cytoplasmically located
endosymbiont bacteria Wolbachia sp. has been implicated in intraspecific
cytoplasmic incompatibility that occurs in many different orders of insects (Wade
and Stevens 1985; Hoffman et al. 1986; Breeuwer and Werren 1990; Giordano et al.
1995). In many hybridization studies, interspecific replacement of the cytoplasm
has no detectable effect on hybrid male sterility (e.g., Zeng and Singh 1993; Davis
et al. 1994; Goulielmos and Zouros 1995). In other hybridizations, cytoplasmic
interactions do play an important role in the reduction of hybrid fitness. For
instance, both nuclear-cytoplasmic and nuclear-nuclear incompatibilities reduce
the fitness of hybrids between two species of wasp in the genus Nasonia (Breeuwer
and Werren 1995). Cage studies in Drosophila have shown instances of more subtle
nuclear-cytoplasmic incompatibility, where the frequency of flies with heterospecific cytoplasm decreases (e.g., Hutter and Rand 1995).
There are a few exceptions to Haldane's rule where, in male heterogametic
taxa, female hybrids are more affected than their male counterparts. Many of these
exceptional cases are believed to be due to incompatibilities between cytoplasmic
(maternal) products and the X chromosome (Sawamura et al. 1993a, 1993b;
Sawamura 1996; Wu et al. 1996; Laurie 1997). In the cross of XAXAfemales to
XB males, the male hybrids receive their X chromosome and their cytoplasm
from their mother, but females receive an additional X chromosome (XB) from
their father. The combination of Xa and cytoplasm from species A may result
in hybrid dysfunction, but only in females. Consistent with cytoplasmic-nuclear
interactions, exceptions to Haldane's rule in Drosophila hybridizations occur
more often for viability than for sterility, because maternal effects are likely to be
less important in the latter case. Probably the best studied example o f nuclearcytoplasmic incompatibilities that lead to an exception of Haldane's rule is the
cross between D. simulans females and D. melanogaster males (Sawamura et al.
1993a, 1993b).
Gene Interactions and the Origin of Species 205
204 Genetic Differentiation: From Populationsto Species
Epistasis Gets Complex
While hybrid fitness reduction inherently requires epistatic interactions under the
allopatric divergence model, it is still possible to consider reproductive isolation
evolving via changes at a single genetic locus in each lineage. Most population
genetics models of allopatric divergence make this assumption, and thus are
essentially one-locus models (e.g., Orr 1995; Johnson and Wade 1996; Orr and Orr
1996). Johnson and Wade (1996) explicitly consider the effects of allelic substitutions in conspecific and hybrid genetic backgrounds. Their model essentially treats
the epistasis involved in hybrid fitness reduction as a form of genotype-byenvironment interaction where the conspecific and hybrid genetic backgrounds are
different environments for variants at a single genetic locus.
Yet, the genetic architecture of hybrid fitness reduction often appears to be more
complicated. In the past few years, it has been discovered that the genes that
contribute to hybrid male sterility in the D. simulans/D. tnuuritiana hybridization
appear to display complex epistasis (Cabot et al. 1994; Palopoli and Wu 1994; Perez
and Wu 1995; Davis and Wu 1996; Masdie et al. 1998). That is, introgressions that
sterilize males can be split into two introgressions, neither of which alone can
sterilize males (Palopoli and Wu 1994). The question is now whether there are any
sufficient genes for hybrid male sterility-ones that will cause sterility if introgressed into the genetic background of the other species without their requiring the
introgression of other "helpern genes. Although Odysseus (Ods)locus first appeared
to be a "major genen that causes sterility when introgressed from D. mauritiana
into D. sechellia (Perez et al. 1993), finer-scale analysis revealed that Ods was not
sufficient because introgressions that contain only Ods (without its helper factors)
are semifertile (Perez and Wu 1995). At present, there is no compelling evidence for
the existence of sufficient hybrid male sterility genes in Drosophila.
While the best evidence for hybrid sterility that involves interactions of
conspecific genes comes from the studies in the D.sirnulans clade (where the genetic
resolution is highest), there is support for complex epistasis from other Drosophila
species groups. For instance, fertility reduction in male hybrids between-species in
the D. buzzatii group is due to many genes of small effect that interact to produce
complex epistasis (Naveria and Fontadevila 1986, 1991; Naveria and Masdie
1998). In addition, at least three loci appear to interact to cause inviability of
hybrids between D. buzzatii and D. koepferae (Carvajal et al. 1996).
Complex Epistasis, the Wright-Fisher Debate, and
the Evolution of Natural Populations
The Issue
Are population subdivision, linkage, and within-species epistasis important in the
evolution of postzygotic reproductive isolation? This question is actually a microcosm of the debate between R. A. Fisher and Sewall Wright about how natural
populations evolve (Brodie, chap. 1, this volume). While Fisher (1918, 1930)
certainly believed that nonadditive genetic interactions could be commonplace, he
generally regarded them as unimportant and similar to environmental variation in
their effects. Instead, he focused on the average effect of an allelic substitution. In
contrast, Wright (1931, 1932, 1977) believed that epistasis was universal a n d played
a crucial role in evolution. In Wright's (1932, 1977) view, populations were often
constrained at local, but not global, optima along the adaptive landscape, unable to
scale higher peaks by mass selection. Wright proposed his shifting balance process
as a mechanism in which these populations could reach higher peaks. This process
requires assumptions about population structure, epistatic genetic variance, and
linkage (Wright 1932, 1965, 1977). Wright's theory had tremendous influence on
founder-effect speciation theory (e.g., Mayr 1963; Carson 1975; Templeton 1980a;
Provine 1989).
Discussions and debate about the shifting balance process, epistasis, and the
nature of adaptive landscapes continue in the current evolutionary biology literature (e.g., Crow et al. 1990; Barton 1992; Wade 1992b; Kauffman 1993; Phillips
1993, 1996; Wagner et al. 1994; Whitlock et al. 1995; Michalakis and Slatkin 1996).
The contrast between two recent perspectives in the journal Evolution (Coyne et al.
1997; Wade and Goodnight 1998) very clearly demonstrates that issues of the
Fisher-Wright debate about the shifting-balance theory per se, and evolution in
natural populations in general, are still contentious.
The Introgression Approach
What do the studies of the genetic architecture of hybrid fitness reduction tell us
about the validity and importance of these peak shift models of speciation? What
do they tell us about the evolutionary dynamics that act upon the positive
phenotype and, specifically, whether epistasis is involved in this process.
First, is epistasis necessarily involved in the negative phenotype? Although
strong epistasis for the proportion of males with motile sperm is observed between
conspecific genes, it is still possible that, on a different phenotypic scale, the effects
of the genes could be additive (discussed in Palopoli and Wu 1994, and references
therein). It is also possible that, at a biochemical level, the conspecific genes are not
interacting with one another. Consider the case where two different loci ( A and B)
from species 1 must be introgressed into species 2 to give sterility, or, in other
words, neither A nor B are sufficient alone (Fig. 12.2). There are several possible
models for the actual biochemical interactions, including the following two simple
scenarios. In the "direct interaction model," A and Bcould both be interacting with
each other and a locus or loci in the genetic background of species 2. Alternatively,
A could be interacting with something in species 2's background (call it c) and B
could be interacting with something else in the background (call it d ) . In this latter
case, the "threshold model," sterility is a threshold-like character: one interaction
cannot cause sterility, but two are sufficient (see also Naveira 1992; Naveira and
Maside 1998).
The finding that complex epistasis underlies hybrid sterility, while consistent
with the Wrightian view of evolution, can still be consistent with a Fisherian view,
as Palopoli and Wu (1994) discuss at length. The different types of complex
epistasis discussed in the previous paragraph have different implications. Suppose
206 Genetic Differentiation: From Populationst o Species
A. Complex Epistasis
a
v
6.Direct Interactions
fertile
C. Threshold Model
Figure 12.2 (A) Different genetic interactions that underlie complex epistasis. Solid lines
between loci represent conspecific interactions, dotted lines heterospecific interactions. (B)
Direct interactions. A and B directly interact with c and d. (C) Threshold model. A
interacts with c, B interacts with d. One interacting pair of genes is not sufficient to cause
hybrid fitness reduction.
that complex epistasis was often due to direct interactions. The fact that the
negative phenotype of hybrid sterility requires biochemical interactions among
conspecific loci makes it more likely that those combinations of genes were under
positive selection. That is, the discovery of genes that directly interact in the negative phenotype implies that they would more likely directly interact in the positive
phenotype. In this model, reproductive isolation would likely accrue from divergence that came about via a Wrightian process, not necessarily the shifting balance
sensu stricto, but a process where population structure, gene interaction, and
linkage were of importance (Wade and Goodnight 1998). In contrast, evidence for
the threshold model, while not contrary to the Wrightian viewpoint, does not lend
support to it either.
Unfortunately, we do not have much direct evidence for either of these models.
Palopoli and Wu (1994) present an indirect test. They argue that if epistasis were
involved in the evolution of the positive phenotype, interacting hybrid male sterility
factors should be linked (within a few centimorgan, cM) more often than random
sets of loci. In practice, the introgression method is usually biased toward detection
l i Wu 1994; Wu and Palopoli 1994). To
of linked interacting factors ( ~ a l o ~ oand
address this question, several studies have combined introgressions from different
regions of the chromosome (> 10cM away) that singly do not cause sterility. The
combinations, however, can result in sterility (Naveira 1992; Masdie et al. 1998;
Y. Xu, M. F. Palopoli, and C.-I. Wu 1994, unpublished data). What is not known is
whether the combinations of linked introgressions are more prone to result in
sterility than combinations of similarly sized unlinked introgressions. Thus, we
cannot yet infer, from the genetic architecture of the negative phenotype, whether
epistasis is important in the positive phenotype.
Gene Interactions and the Origin of Species 207
The Quantitative Genetic Approach
What are the evolutionary processes responsible for the divergence that leads to
hybrid fitness reduction (the positive phenotype), and what are the relative
importances of each of these processes? It is difficult to address these questions,
even in model systems like the D. simulans clade. Outside of these few model
systems, the tools needed for fine-scale genetic mapping and molecular characterization are just not available. Given these limitations, what generalizations can
we make about the evolution of postmating reproductive isolation in general, and,
more specifically, about the role that epistasis may play?
One complementary approach to the study of hybrid traits is the use o f quantitative genetic tools to characterize the underlying genetic variation and covariation
within one species for traits that are expressed only in the hybrids produced by a
cross with another species (Wade and Johnson 1994; Wade et al. 1994,1997, 1999).
The characterization of variation is, in general, instrumental for the study of its
adaptive significance,especially in organisms that do not lend themselves a s easily
to fine-scale genetic analysis. While this approach has thus far been used only in
hybridizations between two closely related species of flour beetles (Tribolium
castaneum and T.freemani), it is quite general; it can be used with any pair of species
that can produce hybrids and whose breeding can be controlled.
Interspecific crosses of T. castaneum and T. freemani generally produce a large
number of viable but sterile hybrid offspring (Wade and Johnson 1994). In the cross
of T. castaneum males to T. freemani females, there are generally more female than
male hybrids (owing to hybrid male inviability), and the surviving hybrid males
often have morphological deformities (leg and antennal) (Wade and Johnson 1994;
Wade et al. 1994, 1997, 1999). In contrast, morphological abnormalities are rare
and sex ratios are near 1 : 1 in conspecific crosses, even under conditions of
inbreeding and temperature extremes (Wade et al. 1997). Upon mating to the same
reference strain of T.freemani, different naturally occurring and laboratory strains
of T. castaneum produce hybrids that differ substantially in these hybrid traits.
Moreover, by use of breeding designs (such as half-sib lineages), we have been able
to detect abundant among-lineage, within-strain genetic variation (Wade et al.
1994, 1997, 1999).
More complicated breeding designs can be used to detect epistatic genetic
variance (M. J. Wade, N. A. Johnson, and J. Santiago-Blay 1999, unpublished
manuscript). If groups of brothers are used as sires, first-cousin families-in addition to half-sib and full-sib families~canbe generated. With these three levels of
grouping, it is possible to detect and estimate the additive x additive epistatic
component of genetic variance for these hybrid traits (Falconer and McKay 1996).
Significant additive x additive epistatic genetic variance for both traits (hybrid sex
ratio and the proportion of hybrid males with deformities) segregates within a
laboratory strain of T. castaneum (M. J. Wade, N. A. Johnson, and J. SantiagoBlay 1999, unpublished manuscript).
What does epistatic genetic variation for these hybrid phenotypes imply for the
evolution of the positive (pure species) phenotype? The question again i s one of
continuity: "What does knowledge about the genetic architecture of the negative
phenotype (hybrid fitness reduction) tell us about the evolution of the positive
Gene Interactions and the Origin of Species 209
208 Genetic Differentiation: From Populationsto Species
phenotype (the underlying divergence)?" These results would appear to support
epistatic genetic variance playing an important role in the evolution of the positive
phenotype. However, until we obtain a better theoretical understanding of the
continuity of gene interaction across these two phenotypic realms, this conclusion
cannot be considered definite.
Epistasis and its Consequences
Given that hybrid incompatibility requires nonadditive genetic interactions, which
are often complex, what can we predict about the patterns of speciation? Orr's
(1995) population-genetic model of the evolution of hybrid incompatibility predicts
that postzygotic reproductive isolation between diverging populations should
accumulate at a rate faster than linear with time (what he calls "snowballing").
Snowballing occurs because of the rapid rise in the number of combinations that
could potentially reduce the fitness of hybrids as the number of genetic differences
increases. Orr (1995) has also shown that it may be easier to evolve reproductive
isolation with incompatibilities that involve three or more loci than those with just
two (see also Cabot et al. 1994). At the initial stages of isolation, however, systems
with only two incompatible loci should evolve reproductive isolation faster than
those with three or more because of the required waiting time for mutation and
fixation at each locus (N. Johnson 1999, unpublished results). Hence, the
"snowballing" effect should be even more pronounced in incompatibility systems
with a large number of loci.
Coyne and Orr (1989b, 1997) and Sasa et al. (1998) have examined the relationship between genetic divergence and indices of reproductive isolation in Drosophila
and frogs, respectively. In both the Drosophila and the frog data sets, the rates of
acquisition of postzygotic reproductive isolation are linear, or even less than linear
(not snowballing), with respect to genetic distance (as measured by Nei's D, which
is believed to increase roughly linear with time). A plausible explanation for this
apparent discrepancy is that the relationship between numbers of genetic incompatibilities and the indices of reproductive isolation used by Coyne and Orr (1989b,
1997) and Sasa et al. (1998) is asymptotic. For instance, there are many more
incompatibility loci found in the D. simulans/D. mauritiana hybridization than in
the D. simulansjD. sechellia hybridization (Johnson et al. 1993; Wu et al. 1993;
Palopoli et al. 1996), but the postzygotic reproductive indices for these hybridizations are the same (0.5) (Coyne and Orr 1989b, 1997).
Dobzhansky (1937) and Muller (1942) realized that their two-locus model of the
evolution of reproductive isolation predicts asymmetry of the location of hybrid
incompatibility loci in reciprocal crosses. That is, the genes from species A that are
incompatible with the genetic background of species B should not be the same as
the genes from species B that are incompatible with the genetic background of
species A. Later researchers extended the theory and presented evidence for initial
asymmetry followed by symmetrical sterility/inviability (Wu and Beckenbach
1983; Zouros 1986; Johnson et al. 1993; Zeng and Singh 1993).
More recent data from high-resolution mapping also support asymmetry. Studies
that introgress D. mauritiana X-chromosome segments into the genetic background
I
I
i
I
of D. simulans map the Odysseus (Ods) hybrid male sterility factor to position 16D
of the polytene chromosome (Perez et al. 1993; Perez and Wu 1995; Ting et al.
1998). Note that while Ods requires "helper factors" to yield complete sterility
(Perex and Wu 1995), it can still be discretely mapped and characterized at the
molecular level (Ting et al. 1998). Reciprocal introgressions-that is, introgression
of the segments of D. simulans into the genetic background of D.mauritianareveal several factors that interact to cause sterility in hybrids (Palopoli and Wu
1994). None of these factors maps to position 16D (Palopoli and Wu 1994). Hybrid
sterility factors are located at different positions in the reciprocal introgressiins.
The genetic architecture of hybrid fitness reduction may also have implications
for models of the evolution ofprezygotic reproductive isolation. In contrast to postzygotic reproductive isolation, prezygotic isolation may evolve because of the direct
action of natural selection according to the reinforcement model (Dobzhansky
1940; Howard 1993). Although reinforcement was previously considered to be
unimportant and perhaps even implausible (e.g., Butlin 1989), in the 1990s several
lines of empirical evidence (Coyne and Orr 1989b, 1997; Howard 1993; Noor 1995;
Butlin and Tregenza 1997; Saerte et al. 1997) have been found that support the
importance of reinforcement. In addition, more recent theoretical models show
that reinforcement is plausible under a fairly wide set of conditions (Liou a n d Price
1994; Kelly and Noor 1996; Servedio and Kirkpatrick 1997; Kirkpatrick and
Servedio 1999).
Two more recent models consider the feasibility of reinforcement, given different
cases of the genetic architecture of hybrid fitness reduction (Kelly and Noor 1996;
Kirkpatrick and Servedio 1999). Kelly and Noor (1996) find that, in cases where
hybrid males are less fit than hybrid females (in accordance with Haldane's rule for
male heterogametic taxa), reinforcement will be facilitated if X-autosome gene
interactions underlie much of the hybrid fitness reduction. In the case where hybrid
females are more adversely affected, X-autosome interactions that cause hybrid
fitness reduction will impede reinforcement. Given the generality of ~ a l d a n e ' rule
i
(Wu et al. 1996; Laurie 1997; Orr 1997), we might expect reinforcement more often
in male heterogametic taxa (e.g., flies and mammals) than in female heterogametic
taxa (e.g., birds and butterflies) if X-autosome interactions are also generally
responsible for much hybrid fitness reduction. For the situation wherein a n island
population receives migrants from the mainland, Kirkpatrick and Sevedio (1999)
show that the likelihood of reinforcement should increase as the number of loci
that interact to cause hybrid incompatibility increases. This effect increases much
less than linearly with the number of loci involved, such that an increase in the
number of loci involved in hybrid incompatibility beyond about 10 has little further
effect on increases of the likelihood of reinforcement (Kirkpatrick and Servedio
1999).
Synthetic Lethals and Reproductive Isolation
Synthetic lethals are genetic systems wherein variants at different loci cause lethality,
but only in combination; individually, their effect on fitness is minimal
(Dobzhansky 1946). Synthetic lethals and the more general case of synthetic
2 10 Genetic Differentiation: From Populations to Species
deleterious loci (SDL) represent an extreme form of synergetic epistasis-a type of
gene interaction that has important implications for the evolution of sex and
inbreeding depression (Phillips and Johnson 1998; Phillips et al., chap. 2, this
volume). While it was a fairly popular topic of study between approximately 1955
and 1975, the subject of synthetics in natural populations has been largely neglected
in the more recent literature (but see Thompson 1986; Phillips and Johnson 1998).
This neglect is unfortunate because an understanding of the role that variation in
SDL systems plays in natural populations can help address the following question:
"To what extent can hybrid dysfunction be generated during the separation of
populations by variation already present in the original population, where there is
no input of new mutations?"
Muller (and the classical school) differed with Dobzhansky (and the balance
school) on this issue. Their differences are highlighted in a passage from Lewontin
(1974, pp. 27-28) who states
[I]f populations are almost entirely homozygous [as held by the classical school], then
speciation must await the occurrence of new mutations that may be advantageous
in a new environment in which an isolated population is found.. . . Unlike the
hypothesis of homozygosity, the balance hypothesis presumes that the genetic
variation for speciation is always present, so that speciation awaits only the
appropriate biogeographical and ecological events.
To address how much hybrid fitness reduction can result from already-present
variation, two factors are of interest. (1) "How much variation is likely to be present
in SDL systems in mutation-selection balance?" (2) "To what extent can a given
amount of variation in SDL systems lead to hybrid fitness reduction upon
vicariance?"
To address the first issue, Phillips and Johnson (1998) examined the expected
frequencies of carriers at SDL at mutation-selection balance. They found that, in
the case where only the double homozygotes (aabb individuals, where a and b are
carrier alleles) are selected against, the frequency of SDL carriers can be rather
high: with free recombination, approximately the quartic root of the mutation rate
(u) divided by the selection coefficient (s), and even higher with tight-linkage. For
instance, given u = l o 6 and s = 0.5, the expected carrier frequency is about 3.7%.
If the extreme synergetic epistasis of SDLs is common, then it is likely that there will
be much SDL variation within populations. Unfortunately, at this time, we know
too little about the degree to which multilocus systems exhibit extreme synergetic
epistasis to judge whether SDL variation should be common (Phillips and Johnson
1998).
Despite the possibility of the existence of much standing variation in SDL
systems, it appears unlikely that this variation can generate much reproductive
isolation upon splitting of the original population and subsequent genetic drift in
the subpopulations (N. A. Johnson 1999, unpublished results). Carrier alleles at
alternative loci in an SDL system must be fixed in each population (i.e., one must
become fixed for AAbb and the other fixed for aaBB). As each fixation event has a
low probability (no greater than the frequency of the carrier allele prior to
subdivision), the probability of carriers fixing at alternative loci will be very low. If
after the vicariance event, however, the fitness matrices changed such that carrier
Gene Interactions and the Origin of Species 2 1 I
genotypes were advantageous, variation at SDL could be a major contributor to
reproductive isolation in the early stages of speciation. Note that this scenario
requires that assumptions are met about both the fitness parameters in new environments and the types of genetic interactions involved. How often such conditions
will occur in nature is not known.
Second, even if alternative alleles at SDL are fixed in different populations, the
amount of reproductive isolation generated upon crossing the two populations is
likely to be low. Given complete epistasis, alternative fixation of SDL carriers
cannot result in fitness reduction in the Fl and will generate only some Fz fitness
reduction (1116 of the Fz will be aabb with free recombination; fewer with linkage).
If there were some selection against AaBb individuals (incomplete epistasis), more
reproductive isolation would be generated by crossing populations fixed for
alternative carriers. In that circumstance, however, the probability of fixation of
alternative carriers would be dramatically reduced because (1) the expected frequency of carrier alleles at mutation-selection balance declines substantially with
departures from complete epistasis (Phillips and Johnson 1998), and (2) there is
more selection in operation against carriers after the populations split.
Perspective and Prospectus
T o use a single term, epistasis, to describe all gene interactions suggests we either do
not care or do not know how to deal with the complexity that interlocus interactions
bring. Different forms of gene interaction have different consequences, and we
should move toward elucidating the nature of these consequences and which forms
of interaction are most prevalent.
(Phillips 1998, p. 1169)
With respect to the role of epistasis, postzygotic reproductive isolation is different
from other areas of evolutionary genetics. While gene interaction may or may not
be of general importance for the evolution of natural populations, it need not be.
Adaptation can work perfectly well in an additive world. In contrast, postzygotic
reproductive isolation requires gene interaction, at least between heterospecific loci.
Because of the ubiquity and necessity of gene interaction in postzygotic
reproductive isolation, researchers in this field need to pay special attention to
Patrick Phillips' words quoted above.
Simple documentiation of the regions of the chromosome (or even the specific
genetic loci) that interact to cause hybrid fitness reduction does not allow us to
easily understand the processes by which reproductive isolation has accumulated
between the diverging species. We expect genetic interaction to be responsible for
the reduction of fitness in hybrids, regardless of the evolutionary processes and
forces responsible for the divergence that has hybrid fitness reduction a s a byproduct. Further, just to show that the interactions are complex provides little information about whether gene interaction was responsible for the divergence, and
what types of gene interactions were involved. Different kinds of gene interaction
212 Genetic Differentiation: From Populations to Species
could be consistent with the data shown in the complex epistasis studies, and the
implications of different types of complex epistasis could be quite different.
We need to continue to use the introgression approach (where feasible) to better
characterize the exact nature of what has been called complex epistasis. At the very
least, studies such as those of Naveira (1992) and Masdie et al. (1998). wherein
introgressions from different parts of the genome are combined, need to be performed more systematically. Unfortunately, these fine-scale genetic dissections are
currently feasible in only a few model systems, such as the Drosophila simulans
clade, and require an enormous amount of time and resources. We also need more
empirical work on speciation genetics outside of model systems. While highresolution genetic mapping studies are impossible at the moment, and probably will
remain so for the foreseeable future, quantitative-genetic approaches such as the
one outlined above, in connection with developing theory, can be very useful.
Despite some recent progress in this area (e.g., Orr 1995; Johnson and Wade
1996;Orr and Orr 1996), neither population- nor quantitative-genetic theory is well
integrated with speciation genetics. We need to establish a conceptual framework in
order to understand how the negative phenotype seen in the hybrid genetic
background is affected by the evolutionary forces that act upon the positive
phenotype in the pure species genetic background. Johnson and Wade's (1996)
theoretical consideration of the allelic effects at a single locus under two different
types of genetic backgrounds (conspecific and hybrid) is an important first-step in
the formation of this conceptual foundation. Yet this study, too, is an oversimplification as it considered only single-locus changes in one species. We need to
know the expected signature of different processes that operate upon the
conspecific phenotype that would appear by observation of the hybrid phenotype.
Computer simulations of the divergence of genetic pathways in allopatric populations provide one promising approach to address this issue (N. A. Johnson and
A. H. Porter 1999, unpublished data).
Acknowledgments My thoughts about the role of gene interactions in evolutionary
biology and speciation, in general, have been shaped to a great extent from my countless
discussions with Mike Palopoli, Patrick Phillips, Mike Wade, and Chung-I. Wu. For this,
I am very thankful. Discussions with Mohamed Noor about reinforcement have been also
very useful. Sally Otto, Patrick Phillips, Adam Porter, the editors, and anonymous
reviewers provided many helpful suggestions that improved the readability of this chapter.
Epistasis as a Genetic Constraint
within Populations and an
Accelerant of Adaptive Divergence
amongThem
MICHAEL J . W A D E
Genetic Architecture: Complexity, Additivity, and Epistasis
The "genetic architecture" of a phenotype is its characterization in terms of the
direct-effects of genes and environments, as well as the suite of genetic and
environmental interactions that affect it. In principle, with controlled breeding
experiments, genes can be identified by patterns of Mendelian segregation
associated with function or phenotypic value, and then mapped to a position in a
genome relative to other segregating factors. Associations between genotypic and
phenotypic value can be decomposed (Table 13.1) into. additive (a and b),
dominance (DAand DB), additive-by-dominance (KAaBand KABb),dominance-byepistatic effects. Environmental
dominance (JAaBb),and additive-by-additive (IAB)
factors generally cannot be so easily defined, distinguished from one another, or
mapped colinearly with respect to other environmental factors. Major environmental factors that affect phenotypes in nature can sometimes be identified by the
use of geographic or temporal correlations, but, in the absence of experimental
controls, it is rare that a single environmental factor can be causally isolated with
respect to its phenotypic effects. After an environmental factor has been identified
as being important, its distribution in nature as experienced by the organism is
rarely known, so that most experimental studies must study environmental factors
as fixed, rather than random, effects. The characterization of genotype-byenvironment interactions (G x E) suffers from many of these same limitations.
In this chapter, I will discuss the relationship between gene-by-gene (g x g or
intragenornic epistatic), genotype-by-genotype (G x G), and the more familiar
genotype-by-environment interactions (G x E). (In keeping with the usage elsewhere