Synonymous single nucleotide polymorphism in arsenic (+3) methyltransferase of the Western mosquitofish (Gambusia affinis) and its gene expression among field populations

Abstract

Naturally occurring arsenic is toxic at extremely low concentrations, yet some species persist even in high arsenic environments. We wanted to test if these species show evidence of evolution associated with arsenic exposure. To do this, we compared allelic variation across 872 coding nucleotides of arsenic (+3) methyltransferase (as3mt) and whole fish as3mt gene expression from three field populations of Gambusia affinis, from water sources containing low (1.9 ppb), medium-low (3.3 ppb), and high (15.7 ppb) levels of arsenic. The high arsenic site exceeds the US EPA's Maximum Contamination Level for drinking water. Medium-low and high populations exhibited homozygosity, and no sequence variation across all animals sampled. Eleven of 24 fish examined (45.8%) in the low arsenic population harbored synonymous single nucleotide polymorphisms (SNPs) in exons 4 and/or 10. SNP presence in the low arsenic population was not associated with differences in as3mt transcript levels compared to fish from the medium-low site, where SNPs were noted; however, as3mt expression in fish from the high arsenic concentration site was significantly lower than the other two sites. Low sequence variation in fish populations from sites with medium-low and high arsenic concentrations suggests greater selective pressure on this allele, while higher variation in the low population suggests a relaxed selection. Our results suggest gene regulation associated with arsenic detoxification may play a more crucial role in influencing responses to arsenic than polymorphic gene sequence. Understanding microevolutionary processes to various contaminants require the evaluation of multiple populations across a wide range of pollution exposures.

Keywords: Arizona; Arsenic; Cyt19; Gambusia affinis; as3mt; mosquitofish.

Fig. 1

Diagrammatic representation of arsenic metabolic pathways. After reduction of arsenate (iAs^V) into arsenite (iAs^III), it is metabolized to monomethylarsonic acid (MMA^III or MMA^V) and subsequently to dimethylarsinic acid (DMA^III or DMA^V), a less toxic chemical, via methylation by arsenite (+3) methyltransferase (as3mt) and oxidation. The diagram is based on Khairul et al. (2017) and Minatel et al. (2018)

Fig. 2

Map of Arizona and collection sites for Western mosquitofish. BP Bubbling Pond fish-hatchery, WL Willow Lake, SR Salt River

Fig. 3

Gene expression of as3mt in Western mosquitofishes (Gambusia affinis) normalized to actb transcript levels. A as3mt transcript levels in single-nucleotide polymorphic WL fish (WLM) relative to normal WL fish compared by an independent sample t test; (B) and as3mt transcript levels in SR and BP fish relative to WL compared by a Kruskal–Wallis test with post-hoc multiple comparisons. Mean and standard deviation are plotted. P values are shown above the horizontal lines