4.4. Signature of genomic diversification of P. lilfordipopulations
Local adaptation in effectively closed populations is a trade-off between genetic drift and selective pressures (Savolainen et al., 2013). The two processes are intrinsically associated with the island surface and its exposure to open sea, both affecting the impact of environmental stochasticity (stronger in smaller islets) and resource availability (reduced in smaller islands). Particularly, the vegetation cover, highly correlated to the island size, provides a strong cascade effect on lizard resource availability, including pollen and fruits production, insect visitor frequency and diversity, and even seabirds presence, all potential sources of dietary items (Pérez-Mellado et al., 2008; Ruiz De Infante Anton et al., 2013; Salvador, 2009; Santamaría et al., 2020). Along with stochasticity, this gives rise to a heterogeneous landscape across islands, which is expected to drive independent processes of lizard phenotypic diversification.
We explored the signature of genomic diversification that might underpin this lizard phenotypic diversity through outlier analysis. Outlier distribution showed a clearly skewed chromosome representation, with most outliers falling within the sexual chromosome Z, consistently for all datasets analyzed (Figure 7A). Recent studies in birds and lizards showed that closely related species often present high differentiation on the Z chromosome (Kulikova et al., 2022; Rovatsos et al., 2019), a pattern that is typically explained as faster Z evolution and lower recombination rates (Irwin, 2018; Lima, 2014; Mank et al., 2010; Wright et al., 2015). The existence of potential ”islands of differentiation” within the sexual chromosomes has also been hypothesized (Lavretsky et al., 2019). At present, detection of such “islands” in P. lilfordi Z chromosome would require a higher sequencing coverage than the one currently provided by GBS and RADseq (i.e., genome resequencing).
Most outliers fell within protein-coding genes, although the large majority were intronic (Figure 7A). Substantial evidence supports the notion that introns have a crucial and evolutionarily conserved function in controlling gene expression in eukaryotes (Kumari et al., 2022; Rose, 2019), providing a particularly rapid mechanism for increasing variation in proteome variance by production of a diverse array of alternative splicing variants (AS) (Reixachs‐Solé and Eyras, 2022; Wang et al., 2015). Moreover, most genes with outliers were associated with protein binding and catalytic activities and were involved in metabolic and growth processes (Figure S3), critical molecular functions for gene expression regulation and phenotypic diversification (Van Nostrand et al., 2020). In small, isolated populations, extensive modulation of gene expression conferring phenotypic plasticity could represent a major mechanism to counteract the loss of genetic diversity (Fulgione et al., 2023; Sherpa et al., 2023), a hypothesis that needs to be validated by whole transcriptome and epigenome data (Chapelle and Silvestre, 2022; Fulgione et al., 2023).
We found a subset of protein-coding genes targets of multiple outliers according to both independent sequencing methods, GBS and RADSeq (Figure 7B), which might represent interesting candidates for further exploration of population genomic diversification in P. lilfordi . Of these proteins, few represented potassium and calcium channels, which could hint to an important regulatory role of osmotic pressure in lizards (Dantzler and Bradshaw, 2008). We also highlight the cullin-1 protein, with a shared outlier SNP detected by all datasets. The protein is known to be involved in ubiquitination and subsequent proteasomal degradation of target proteins (Duan et al., 2011; Gao et al., 2011; Scott et al., 2016). Recent evidence indicates a critical role of this protein in modulation of the transcription factor c-MYC, a major regulator of gene expression and cell proliferation (Sweeney et al., 2020). While there are no specific studies on cullin-1 protein in lizards, c-MYC was linked to the cellular regenerative response after tail amputation (Alibardi, 2017). The gradual loss of tail autotomy ability in insular lizards is a hallmark of their reduced antipredator response following insular adaptation (part of the island syndrome) (Cooper et al., 2004; Pafilis et al., 2009; Perez-Mellado, 1997).
Exploration of the major variables driving outlier genotypes primarily recovered a signature of past geographic separation between Menorca and Cabrera/Mallorca, with a large proportion of loci presenting fixed allelic differences between archipelagos (Figure 7C). These SNPs are putatively derived from a past founder effect that occurred during initial colonization of the Menorca archipelago (Brown et al., 2008; Terrasa et al., 2009) or the result of unclear selective forces, including potential selective sweeps (Brown et al., 2023; Campagna et al., 2022; Stephan, 2019). In all cases, they support no recent secondary contact between Archipelagos, in line with previous mtDNA-based phylogeographic reconstructions (Terrasa et al., 2009). We note that given the major confounding effect of archipelago, the significance/impact of other environmental variables in driving genome diversification cannot be reliably assessed with the current sampling design (Bassitta et al. 2021). Additional sampling, along with phenotypic data are required to clarify the putative adaptive role of these genomic changes.