Phenotype- and gene-driven Speciation under Environment-mediated
Natural Selection
The differentiation of floral characteristics is considered the core of
prezygotic isolation in plants and plays an important role in speciation
(Celep et al., 2020; Xu et al., 2011). In the present study, the four
identified groups of A. viridiflora were located in different
climate types and had continuous phenotypic variation. The areas where
NE and NW were located showed large mean diurnal temperature ranges,
while the areas where EL and CN were located showed small mean diurnal
temperature ranges. Populations located in environments with a wide mean
diurnal temperature range exhibit longer spurs and pistils than those
located in environments with a narrow mean diurnal temperature range.
The differences in the spur and pistil lengths may promote the formation
of reproductive isolation (Brothers & Delph, 2017; Kramer, 2009; Sun,
Schlüter, Gross, & Schiestl, 2015). Additionally, plant growth is
greatly affected by the environment; therefore, adaptive adjustments to
different environments are necessary (Anderson & Song, 2020). Among the
adaptable traits, flower size (corolla diameter, petal length, and spur
length) was significantly negatively correlated with the inflorescence
number and leaf area (Table 1 and Figure S5). NE and NW had larger and
fewer flowers and smaller leaves, while CN and EL had smaller flowers, a
greater number of flowers and larger leaves. This phenomenon has also
been reported in Pedicularis siphonantha populations (Dai,
Amboka, Kadiori, Wang, & Yang, 2017). The wide mean diurnal temperature
range limits the activities of pollinators. To ensure the production of
offspring, A. viridiflora uses more energy for reproductive
growth and less energy for vegetative growth. Additionally, the
populations in this area (NW and NE) used the limited energy available
for reproductive growth to form a small number of flowers, thus
enhancing their ability to attract pollinators. By contrast, abundant
pollinators were found in areas (EL and CN) with a
narrow
mean diurnal temperature range, and A. viridiflora populations in
such areas use more energy for vegetative growth to ensure their
long-term survival. The groups in these areas use their reproductive
growth energy to form more flowers and produce more seeds.
According to Wu’s
“Genic
view of the process of speciation”, the key genes or sites of
speciation determine the adaptive differentiation of the population to
the environment in which it is located (Wang, He, Shi, & Wu, 2020; Wu,
2001). Among selected and differentially expressed genes related to the
environment, Aqcoe5G182600 (Cytokinin oxidase 5) might have
played an important role in the early speciation of A.
viridiflora . Cytokinin oxidase 5 and its orthologous genes play an
important role in plant grain yield.Os CKX2knockout results in increases in the numbers of both panicle branches
and grains per plant under stress conditions in Oryza sativa(Ashikari et al., 2005). Accordingly, Aqcoe5G182600 may regulate
the activity of flower primordia and control the number of
inflorescences by controlling the levels of cytokinin. Furthermore, five
candidate genes were related to resistance to environmental stress,Aqcoe1G340100 (Seong et al., 2007), Aqcoe1G488100(Koussevitzky et al., 2008), Aqcoe3G242600 (Magadlela et al.,
2019), Aqcoe5G182600 (S. Li et al., 2019) andAqcoe7G135600 (He et al., 2019), indicating that the differences
in the growth environment drive the genetic differentiation of different
groups of A. viridiflora .
The appearance of nectar spurs is a critical morphological
characteristic of highly diversified Aquilegia . Nosil et al.
proposed that a ‘speciation gene’ is defined as any gene contributing to
the evolution of reproductive isolation (Nosil & Schluter, 2011).
Because Aqcoe5G459400(OBP3-responsive gene 1) was shown to be related to nectar spurs by
integrating the results of WGCNA, and its expression level was
negatively correlated with the spur length, as well as the selected SNP
in Aqcoe5G459400 was differentiated and almost fixed in NE and
EL, Aqcoe5G459400 could be considered a key gene in the early
stage of A. viridiflora speciation. Previous studies have shown
that OBP3-responsive gene 3 (ORG3 ) controls the size of petals by
controlling the number of cells (Omidbakhshfard et al., 2018).
Additionally, A. ecalcarata has short or no spurs because cell
division ceased and cell differentiation began earlier (Ballerini,
Kramer, & Hodges, 2019). Therefore, Aqcoe5G459400 may control
the spur size by controlling the number of cells in A.
viridiflora . Additionally, the ABI3/VP1 transcription factor located
upstream of Aqcoe5G459400 and its orthologs are key genes
involved in regulating abscisic acid (ABA), which not only plays a role
in the dormancy of seeds and buds but also affects the flowering time of
plants (Riboni, Robustelli Test, Galbiati, Tonelli, & Conti, 2016; Shu
et al., 2018; Shu et al., 2016; Shu et al., 2013). In summary, a cascade
reaction between ABI3/VP1 and ORG3 likely exists, controlling the
spur size in A. viridiflora . The expression level ofAqcoe5G459400 was lower in the lineages from regions with a wide
mean diurnal temperature range, causing A. viridiflora to produce
larger flowers to attract more pollinators and increasing the chance of
successful reproduction. By contrast, its expression level is higher in
the lineages from regions with a narrow mean diurnal temperature range,
and A. viridiflora produces smaller flowers in such areas to
increase the number of seeds produced. Thus, the differentiation ofAqcoe5G49500 enables A. viridiflora to develop different
adaptive evolution mechanisms in response to different environments and
promotes speciation. During speciation of A. viridiflora , except
for the differentiation of Aqcoe5G49500 , the gene flow between
populations may increase at other sites because of the genetic variation
mediated by natural selection, which accelerates the divergence ofA. viridiflora into different species (X. Wang et al., 2020).
Additionally, Aqcoe2G056600 , Aqcoe2G172400 ,Aqcoe3G439700 , Aqcoe7G244300 and Aqcoe7G395200interacting with Aqcoe5G459400 in the module are related to
histone methylation and ion transport across membranes and therefore may
improve the ability of plants to resist insect pests and adverse
stresses (Fedoreyeva, Vanyushin, & Baranova, 2020; Liao et al., 2018;
Philippe, Ralph, Külheim, Jancsik, & Bohlmann, 2010; Rodríguez-Celma,
Chou, Kobayashi, Long, & Balk, 2019; Surya, 2020).