DISCUSSION
Signatures of selection detection in both wild and domesticated species
is considered a critical step toward understanding the molecular basis
of adaptive evolution. This information can be used to identify genomic
regions and underlying biological mechanisms affecting their essential
traits. There are different methods of detecting selection signatures.
However, two complementary approaches (composite likelihood ratio test
(CLR) and integrated haplotype score (iHS)) that were found to have
power > 70% to detect selection signatures were applied in
this study. Unfortunately, there is currently no information on the
effects of selection on the wild African harlequin quail, despite the
incessant harvesting by rural smallholder farmers, effects of habitat
destruction, and climate change, among other challenges. Moreover, there
is limited information on selection signatures detected in the domestic
Japanese quail, the most common domesticated quail species reared in
Kenya. It has experienced constant evolutionary and genetic changes in
its phenotype and genome architecture due to modern breeding methods.
Such information could have been a foundation for understanding
biological pathways governing important traits in wild quails and how
modern breeding methods will likely affect them if breeding initiatives
were to be made.
The application of CLR and iHS in detecting selective sweeps in wild
African harlequin and domestic Japanese quail genomes was successful.
Several complementary methods for signatures of selection detection
allow for improved testing as each statistical test captures patterns in
data differently (Fariello et al., 2013). The linkage disequilibrium
(LD)-based iHS approach, which requires haplotypes per individual of one
population to detect selection, was anticipated to be more reliable for
this study due to the single populations that were sampled and the lack
of a good reference population about quail (Voight et al., 2006). In
addition, the CLR method was also adopted for this study as it uses
allele frequency data to compare a neutral and a selective sweep model,
and it is not highly sensitive to assumptions about the underlying
recombination rate or recombination hotspots (Williamson et al., 2007).
Furthermore, the CLR method can detect a beneficial mutation that spread
in the entire population. In contrast, methods based on extended
haplotype length and high linkage disequilibrium only detect the
beneficial mutation that has yet to spread throughout the entire
population (Voight et al., 2006; Wang et al., 2006; Williamson et al.,
2007).
Both signatures of selection detection methods identified candidate
genes associated with crucial biological, molecular, and cellular
processes such as immune response, growth, reproduction, and
morphological and behavioral traits. Some essential candidate genes
involved in various immune response processes and pathways include
MAPK1, MAPK13, MAPK14, CREB1, DYNLL2, RAC1, and ITGB3. The MAPK
signaling pathway genes were enriched in the wild African harlequin and
domestic Japanese quail. Mitogen-activated protein kinases (MAPKs) are a
group of serine/threonine protein kinases that are highly conserved and
have essential roles in cellular processes, such as proliferation,
stress responses, apoptosis, and immune response regulation (Liu et al.,
2007). The MAPK13 and MAPK14 (p38 MAPK pathway) genes, which play an
important role in inflammatory responses, were positively selected in
the wild African harlequin quail. The p38 pathway participates in the
innate and adaptive immune response process by controlling the
production of inflammatory cytokines (TNFα, interleukin (IL)-1, IL-10,
and IL-12) by specialized dendritic and macrophage cells, CD40-induced
gene expression, and proliferation of B cells, antigen processing in
CD8+ conventional dendritic cells, and T cell
homeostasis and function (Arthur & Ley, 2013; Soares-Silva et al.,
2016; Han et al., 2020).
In addition to the p38 pathway, the extracellular signal-regulated
kinase (ERK1/2) is also a MAPK pathway involved in positive selection
and the differentiation of DP thymocytes to either CD4 or CD8 T cells
(Fischer et al., 2005). The MAPK1, a key component of the ERK1/2
pathway, was positively selected in domestic Japanese quails. It was
shown to participate in adaptive immune responses during bacterial
infection in Nile tilapia (Wei et al., 2020). MAPK1, through the
MAPK/ERK pathway, facilitates the activation and proliferation of immune
cells, including T cells, B cells, and macrophages (Sun et al., 2015).
Bacterial pathogens such as Salmonella , Shigella ,Yersinia and Escherichia species are known to manipulate
pathways like the MAPK in their hosts to propagate infection (Krachler
et al., 2011). Another positively selected candidate gene in domestic
Japanese quails associated with Salmonella infection was the
DYNLL2 gene, which is predicted to be involved in cytoskeletal motor
activity and cytoskeletal protein binding. An observed increase in
DYNLL2 protein in gamma delta T-lymphocytes (γδ T-lymphocytes) of
chickens infected with Salmonella Enteritidis showed its role in
regulating immune response (Sekelova et al., 2017). Like commercial
chicken, domestic Japanese quails are reared on farms under different
housing systems such as battery cages, free-range and floor-raised
systems, among others. Choice of the housing system in connection to
other production factors such as infrastructure, stocking density (farm
and flock size), manure collection, disease status of the flock, and
rodent and insect load does determine the risk of Salmonellainfection (Van Hoorebeke et al., 2011).
The ITGB3 gene, which was positively selected in the wild African
harlequin quail, is involved in the blood coagulation pathway through
adrenergic signaling in platelet activation. ITGB3 encodes for
glycoprotein IIIa (GPIIIa) and, along with the alpha IIb chain, forms
the platelet adhesive protein receptor complex glycoprotein IIb/IIIa (GP
IIb/IIIa), which mediates platelet aggregation by acting as a receptor
for fibrinogen (Cerhan et al., 2007). It plays a critical role in many
cellular processes, such as cytoskeletal organization, cell adhesion,
migration, proliferation, and survival (Ridley et al., 1992; Arthur et
al., 2002; Guo et al., 2008). Its immune roles in the T cell development
(Luo et al., 2013), B cell development and signaling (Walmsley et al.,
2003), epidermal homeostasis resulting in wound healing (Winge &
Marinkovich, 2019), and phagocytosis ( Lee et al., 2000; Han et al.,
2019) are continuously being studied. Phagocytosis is a necessary
component of the innate immune response. It plays an essential role in
host-defense mechanisms by enabling the uptake and destruction of
infectious pathogens through specialized cell types like macrophages,
neutrophils, and monocytes (Lee et al., 2020). Similar to the ITGB3
gene, the RAC1 candidate gene that was detected in domestic Japanese
quail is also associated with wound healing by regulating the innate
immune response in keratinocytes (Bustelo et al., 2007; Pedersen et al.,
2012; Winge & Marinkovich, 2019).
PPP1CA, WNT5A, GRIA1, CREB1 and ADCY8 candidate genes, associated with
adaptation and behavior, were detected in the wild African harlequin
quail. Melanogenesis and ear morphogenesis biological processes are
crucial for wild African harlequin quail breeding behavior, social
interactions, and survival. Melanogenesis involves melanin production,
which plays a significant role in structural plumage in birds (Jeon et
al., 2021). Plumage color and patterns are helpful in camouflage,
mating, and differentiating between male and female wild African
harlequin quails (Mason & Bowie, 2020). Melanin-based patterns are
influenced by melanocyte migration, differentiation, cell death, and/or
interaction with neighboring skin cells at the cellular level (Inaba &
Chuong, 2020). The wild African harlequin quail songs and calls are also
essential to their communication and behavior; hence, ear morphogenesis
is a vital biological process for their survival and reproduction. The
WNT5A gene, among other genes, was implicated in melanogenesis, ear
morphogenesis, sex differentiation, and muscle development. Through
activating multiple intracellular signaling cascades, the WNT gene
family controls cell proliferation, differentiation, apoptosis,
survival, migration, and polarity (Kikuchi et al., 2012). According to
Kikuchi et al. (2012), regulating cellular functions, including
migration and differentiation, makes the WNT5A gene a target for
selection.
The GluA1 subunit has been implicated in the regulation of circadian
rhythms and behavior (Ang et al., 2021). The Glutamate receptor 1
(GluA1), encoded by the GRIA1 gene, forms part of the AMPA receptor that
mediates fast glutamate signaling in the central nervous system
(synaptic plasticity). Positive selection of genes associated with
behavioral response to stress and circadian entrainment (GRIA1, CREB1,
ADCY8) could contribute to the observed timely seasonal migration and
behavioral patterns of wild African harlequin quails (Bossu et al.,
2022). The PPP1CA gene encodes for a protein that is part of the three
catalytic subunits of protein phosphatase 1 (PP1). The PP1 protein was
found to be a key regulator of period and light-induced resetting of the
circadian clock in the common fruit fly (Fang et al., 2007) and
different mammals (Eide et al., 2005; Gallego et al., 2006; Schmutz et
al., 2011). It regulates the circadian period length, in counterbalance
with casein kinase 1δ and ε (CK1δ/ε), through the regulation of the
speed and rhythmicity of period circadian regulator 1 and 2 (PER1 and 2)
phosphorylation (Lee et al., 2011; Schmutz et al., 2011).
Candidate genes associated with growth and reproduction were detected in
the wild African harlequin and domestic Japanese quail. The Wnt–Hippo
signaling pathway-related genes identified in wild African harlequin
quails include SOX2, FZD7, WNT3, NKD1, and WNT4. The Hippo signaling
pathway plays a crucial role in cellular differentiation, tissue, and
organ development by controlling organ size through the regulation of
cell proliferation and apoptosis (Justice et al., 1995; Xu et al., 1995;
Heallen et al., 2011; Wu & Guan, 2021). It has also been implicated in
other diverse roles, such as tissue homeostasis, wound healing and
regeneration, immunity, tumorigenesis, and embryogenesis (Wu & Guan,
2021). The transcription factor SOX2 is involved in osteoblast
differentiation (Seo et al., 2013), inner ear development (Kiernan et
al., 2005), neurogenesis, and the proliferation and/or maintenance of
stem cells (Oesterle et al., 2008), among others. The positive selection
of genes associated with tissue and organ size, among other factors,
could contribute to how the wild African harlequin quails have managed
to maintain their small body size, supporting their growth and survival
in the wild. The wild African harlequin quail is known to fly longer
distances during migration and at faster speeds (Wamuyu et al., 2017).
CREB1 gene is involved in growth hormone synthesis, secretion, and
action. CREB1 encodes a phosphorylation-dependent transcription factor
that mediates the response to various cellular processes, including
regulation of transcription, signal transduction, glucose homeostasis,
and growth-factor-dependent cell survival, proliferation, and memory
(Kinjo et al., 2005). CREB1 stimulates transcription upon binding to the
DNA cAMP response element (CRE) located in the promoter region of target
genes, leading to the recruitment of transcriptional coactivators and
the initiation of gene transcription (Shankar et al., 2005). In humans,
the CREB/CREB1 gene is involved in immune function (Cerhan et al., 2007)
as it promotes proliferation and survival and differentially regulates
Th1, Th2, and Th17 responses (Wen et al., 2010). It has also been shown
to be a critical driver of vaccine efficacy in non-human primates
(Tomalka et al., 2021).
Several candidate genes associated with lipid and cholesterol transport
and metabolism in domestic Japanese quails, such as the APOA (APOA1 and
APOA4) and ABCA (ABCA2, ABCA5, ABCA7) gene families, were identified
(Albrecht & Viturro, 2007; Dominiczak & Caslake, 2011). Additionally,
muscle cell differentiation, tissue, and structure development genes
(COL6A3, SLC9A1, SMARCD3, MSX2, and PRF1) were identified. The
positively selected DYNLL2 gene, apart from its role in immune response,
was also found to be a regulator of chicken myogenesis, providing
insights into breast muscle development in chickens and other birds (Li
et al., 2022). VIPR2, one of the positively selected cAMP signaling
pathway genes identified in domestic Japanese quails, has been
implicated in the regulation of egg production as it has been associated
with brooding behavior in chicken and geese (Luan et al., 2014; Huang et
al., 2022).
In this study, only two genes (CBFB and RET) overlapped between the CLR
and iHS tests in the domestic Japanese quail, in contrast to the wild
African harlequin quail, where no overlapped genes were observed. CBFB
protein is mainly associated with definitive hemopoiesis (Speck et al.,
1999). In contrast, RET is known for its protein kinase activity,
leading to the activation of signaling pathways involved in cell growth,
differentiation, and survival, such as MAPK and AKT (Schuchardt et al.,
1994; Taraviras et al., 1999).