4 | Discussion
Adaptive introgression is known in some plant taxa, often resulting in
divergent ecotypes colonizing novel habitats (Suarez-Gonzalez et al.
2018c). In European and American oaks, divergence and introgression of
adaptive traits between interfertile species have been demonstrated. In
drought-averse Q. rubra and drought-tolerant Q.
ellipsoidalis (section Lobatae ), parapatric populations of both
species are fixed for alternate alleles at a CONSTANS -like gene,
which affected seedling survival in common gardens, and the Q.
ellipsoidalis allele is introgressed into the Q. rubra genome
depending on soil moisture conditions in their sympatric populations
(Lind-Riehl et al. 2014; Lind-Riehl and Gailing 2016; Khodwekar and
Gailing 2017). In Q. robur and Q. petraea (sectionQuercus ), introgression from Q. robur contributes to
divergence among locally adapted Q. petraea populations along a
gradient of climatic conditions (Leroy et al. 2020). In Asian oaks,
introgressed genomic regions were investigated in admixture betweenQ. acutissima and Q. variabillis (section Cerris ),
and admixed populations occupying similar habitats tended to share the
same introgressed regions (Fu et al. 2022). We also found a candidate
system for adaptive introgression, which consists of Q. mongolicavar. crispula (Qc ) in inland habitats and Q.
dentata (Qd ) in coastal habitats in northern Japan (Nagamitsu et
al. 2019; Nagamitsu et al. 2020). In this system, we tried to verify
intensive introgression of Qd alleles at loci associated with
traits adaptive to coastal environment.
Our previous study demonstrated environment-dependent introgression fromQd to Qc in coastal habitats, resulting in Q. ×angustilepidota (Qa ), which are hybrids after the first
generation of backcross to Qc (Nagamitsu et al. 2020). Our
present study using a comprehensive tree collection in common gardens
revealed a hybrid zone around the northern distributional limit ofQd , including both northern-edge Qd populations admixed
with Qc and coastal Qa populations backcrossed toQc . The observed genomic compositions indicated random admixture
between Qc and Qd , suggesting frequent gene flow and weak
genetic drift. The genotypic PCA indicated that some northern-edgeQd populations had unique genetic properties in some PCs with
minor contributions to genetic variations, suggesting the presence of
weak genetic structure in coastal habitats, probably due to
fragmentation of sand dunes and coastal cliffs in the distributional
range margins. This hybrid zone is likely to be maintained by the
tension zone process (Barton and Hewitt 1985). Near the northern
distributional limit of Qd , gene flow between Qc andQd seems relatively frequent, because their flowering synchrony
increases at higher latitudes probably due to shorter periods of growing
season (Shimizu et al. 1995). Lower fitness of Qd at higher
latitudes in comparison to fitness of Qc coupled with both lower
fitness of Qc in coastal habitats and lower fitness of Qdin inland habitats may be responsible for natural selection that
maintains the tension zone.
Common garden experiments have revealed local adaptation in various
traits of oaks (Cavender-Bares and Ramírez-Valiente 2017). Local
adaptation of Qc to climatic conditions was evident at regional
scales (Nagamitsu and Shuri 2021). The size of 30-year-old trees in our
common gardens indicated that the size was larger in Qc trees
than in Qd trees in the inland site but larger in Qd trees
than in Qc trees in the coastal site. This result demonstrates
both adaptation of Qc to inland habitats and adaptation ofQd to coastal habitats. Although the plantation density and
forest management differed between the sites, smaller tree size in the
coastal site than in the inland site suggests higher stress in coastal
habitats than in inland habitats. In the coastal site, which was located
in north to the northern distributional limit of Qd , the size ofQd trees seemed to be suppressed and to become similar to the
size of Qa trees. This result implies that environmental
conditions at higher latitudes are maladaptive for Qd , which is
distributed at lower latitudes than Qc . These findings of local
adaptation are consistent with natural selection expected from the
tension zone process.
Morphological traits of leaves and shoots observed in each site
indicated more Qd -like phenotypes as the Qd ancestry
increased, suggesting that these traits have the genetic basis derived
from Qd . Artificially crossed F1 hybrids betweenQc and Qd exhibited pubescent shoots and intermediate
phenotypes of morphological traits of leaves and fruits (Ubukata et al.
1996). These findings are consistent with our observed phenotypes of
admixed individuals. We categorized into observed traits into
taxon-specific and habitat-specific traits based on the results of
phenotypic PCA. The habitat-specific traits, of which phenotypes ofQa and Qd trees in coastal habitats mainly differed from
those of Qc trees in inland habitats, included the density of
stellate hairs (trichomes) on the lower leaf surface, leaf mass per area
(LMA), shoot diameter, and the number of axillary buds at bud-scale
scars. These traits seem relevant to the stress tolerance in coastal
habitats. Higher water absorption capacity of oak leaves with dense
trichomes is advantageous in soils with lower water storage capacity
(Fernández et al. 2014). Increased LMA in oaks are commonly observed in
response to higher intensity of drought and light (Sancho-Knapik et al.
2021). More buds at the lower part of shoots, such as bud-scale scars,
can compensate for bud mortality at the upper part of shoots caused by
salt spray and harsh wind in winter (Asai et al. 1986). These lines of
evidence suggest that Qd -like phenotypes of the habitat-specific
traits are more adaptive in coastal environment. Thus, we expected thatQd alleles at loci associated with these traits are more
intensively introgressed than those in the genomic background.
The observed shift in phenotypic variation to more Qd -like
phenotypes from inland to coastal habitats suggests phenotypic
plasticity in response to coastal stress. This phenotypic shift was
larger in Qa trees than in Qd trees, indicating lower
plasticity in more stress-tolerant Qd trees. This result is
consistent with our previous findings that phenotypic plasticity in
natural populations in response to coastal stress, which was measured by
bud mortality, decreased with higher Qd ancestry estimated from
nuclear microsatellite genotypes (Nagamitsu et al. 2019). Similar
features in phenotypic plasticity were evident in Mediterranean oaks,Q. faginea , which showed lower morphological and physiological
plasticity and more conservative resource-use strategy in populations
from drier and colder provenances than in those from mesic and milder
provenances (Solé-Medina et al. 2022). The phenotypic shift to moreQd -like phenotypes of Qa trees may result from induced
expression of alleles introgressed from stress-tolerant Qd in
response to coastal stress.
Introgression from Qd to Qc was evident from a positive
genome-wide estimate of the Patterson’s D statistic in our
previous study (Nagamitsu et al. 2020), and genome-wide patterns of
introgression in our present study using more loci indicated that the
extent of introgression were heterogeneous along chromosomes. However,
extensively introgressed regions were not always consistent with local
ancestry, probably due to incomplete linage sorting. To determine
introgressed genomic regions precisely, denser SNP loci mapped to
reference sequences of both parental taxa are necessary (Ai et al. 2022;
Wang et al. 2023). In spite of the low resolution in our present study,
the heterogeneity along chromosomes and various admixture patterns among
individuals may make our GWA analysis feasible owing to a diverse array
of recombinants.
We detected few loci consistently associated with traits in both SNP
genotypes and local ancestry, probably due to sparse distribution of the
available ddRAD loci, polygenic regulation of the examined traits, and
confounding factors with the measured phenotypes. In a model plant,Oryza sativa , for example, major genes of some agronomic traits
with alleles introgressed between rice varieties were clearly detected
from both association mapping using SNP genotypes and admixture mapping
using local ancestry (Zhao et al. 2010). In poplar, various traits were
measured in a common garden of Populus trichocarpa , and
associations of SNP genotypes with these traits were examined using 29k
genome-wide loci (Mckown et al. 2014a; Mckown et al. 2014b).
Introgression of P. balsamifera alleles to P. trichocarpagenome at some candidate genes around these trait-associated loci were
detected using denser loci around these candidate genes (Suarez-Gonzalez
et al. 2016). Furthermore, using 1169k loci, additional introgressed
regions were found from local ancestry inference and admixture mapping
(Suarez-Gonzalez et al. 2018a; Suarez-Gonzalez et al. 2018b). These
studies imply that more loci are necessary to detect genes responsible
for adaptive traits in our oak system. In spite of the limited power of
our present study using 27k loci, we found several loci potentially
associated with one taxon-specific and three habitat-specific traits.
Two of these loci were located at intensively introgressed regions in
chromosomes 9 and 11.
GO terms of genes around the loci potentially associated with the four
traits suggest that these genes are related to signaling processes and
stress responses with functions of DNA binding and protein binding. Some
proteins of those genes were related to catabolic pathways of abscisic
acid (ABA) in response to drought stress (Kushiro et al. 2004) and ABA
transduction cascades in dormant seeds (Lorenzo et al. 2002),
immunophilin responses to bacterial invasion (Mokryakova et al. 2014)
and homoserine accumulation triggering pathogen resistance (Van Damme et
al. 2009), and catabolism of lysine during leaf senescence (Engqvist et
al. 2011). These findings imply that these genes can be related to the
response to coastal stress. In introgression between Q.
acutissima and Q. variabilis in China, not only genes involved
in signaling processes such as responses to abiotic stimulus, stress,
hormone, and pathogens but also cis -regulatory elements in
untranslated regions were enriched in introgressed genomic regions (Fu
et al. 2022). To detect candidate genes responsible for theQd -like phenotypes in response to coastal stress, not only
morphological and physiological traits but also gene expression patterns
should be investigated to select essential transcription factors and
regulatory elements underlying adaptive phenotypes.
In summary, we revealed a hybrid zone with random admixture betweenQc and Qd in the northernmost area of Hokkaido. This
hybrid zone is located in coastal habitats around the northern
distributional limit of Qd , where Qd -like phenotypes are
adaptive. We found heterogeneity of introgression along chromosomes and
various admixture patterns among individuals, but failed to detect loci
associated with the Qd -like phenotypes. To elucidate adaptive
introgression suggested in our present study, more SNP genotypes at
denser loci in admixed populations and deeper knowledge of genetic basis
of adaptive traits are necessary.