CHAPTER 3: Results
4.1. TEs accumulation on Sb
Among total 260 genes with copy number differences between SB and Sb-carrying individuals in the fire ant, we found around half (50.4%, n
=131) to be TEs. When using a more relaxed threshold to identify CNV genes (log2-fold ≥0.6 or ≤-0.6 (~1.51 fold)), we found a similar proportion of TEs (53.9%, n = 256, out of 475 CNV genes).
This accumulation of TEs is consistent with theoretical predictions for non-recombining genomic regions (Bachtrog, 2013; Bartolomé et al., 2002; Bergero & Charlesworth, 2009; Dolgin & Charlesworth, 2008; Liu et al., 2004; Peichel et al., 2004).
We found 124 TEs with greater copy number in Sb, and only 7 in SB, confirming a general trend of accumulation of repetitive elements on the Sb genome (Wang et al., 2013). Since TEs accumulate in regions with suppressed recombination, finding of TEs on Sb is most probably a consequence of reduced recombination between the two supergene alleles in heterozygous SB/Sb queens coupled with no (or potentially infrequent) recombination between Sb alleles due to the essential absence of functional Sb/Sb queens. In the invasive range Sb/Sb queens are extremely rare (<1%) (Fritz, Vander Meer, & Preston, 2006; K. G. Ross, Krieger, Shoemaker, Vargo, & Keller, 1997) while in the native range (South America) none have been reported but plausibly could occur (~20% Sb/Sb polygyne queens have been observed for a related species, Solenopsis richteri) (Hallar et al., 2006; K G Ross, 1997). Since normal
accumulation of TEs on SB.
Two models can explain the presence of TEs with greater copy number in Sb in relation the reduced recombination between SB and Sb variants of the supergene:
1) More TEs were able to proliferate after the formation of the supergene, due to lack of recombination and consequent reduced purging of deleterious repeats insertion.
2) Both SB and Sb possessed similar levels of TEs at the time of the supergene formation, and these TEs were lost on SB though recombination, but retained in Sb.
Both processes may have occurred, perhaps on different TEs.
Most of the TE duplications (and deletions) were likely already present in the native range. First, the introduction of S. invicta into the Southern United States was less than 100 years ago (Ascunce et al., 2011;
Tschinkel, 2006), which seems too little time for the observed TE patterns. Second, while our bioinformatics analysis was for a population from Georgia, USA, we validated CNV TEs with a Taiwan population (10 out of 12 were confirmed). Finding similar TE duplications in both Georgia and Taiwan supports the idea that most of the Sb-linked TE likely predated invasion into the USA.
Similar cases of inversions and lack of recombination, such as the animal Y or W sex chromosomes, show accumulation of TEs and deleterious mutation (Bachtrog, 2006; Peichel et al., 2004; Vicoso, Emerson, Zektser, Mahajan, & Bachtrog, 2013; Yoshida, Makino, & Kitano, 2016). In the case of the fire ant Sb allele, most of the TE insertions presumably have no fitness effect, i.e., they are neutral. However, some TE insertions may contribute to the deleterious phenotypes present in Sb-carrying
individuals (e.g., lower sperm production in males (Huang & Wang, 2013;
Lawson, Vander Meer, & Shoemaker, 2012), and low viability of Sb/Sb individuals (Hallar et al., 2006)). Another possibility is that some of the TEs contribute adaptively to Sb phenotypes by providing cis-regulatory elements or promoting host gene duplication. For example, in Drosophila miranda, helitron transposable elements mediate dosage compensation by creating MSL (male specific lethal) complex binding sites (Ellison &
Bachtrog, 2013). Similarly, cis-regulatory elements from endogenous retrovirus and TEs have been co-opted to regulate the immune pathway and pregnancy-related genes in mammals (Edward B Chuong et al., 2016;
Lynch et al., 2015). Furthermore, host genes can be captured by TEs, and in this way TEs can promote gene transduction (the process of transferring DNA to a new genetic location) and duplication (Jiang, Bao, Zhang, Eddy, & Wessler, 2004; Moran, DeBerardinis, & Kazazian, 1999).
Along these lines, it is interesting to note that the BEL transposon family, with 18% (n =22) of the CNV TEs, seems to have disproportionately proliferated on the Sb supergene. TE from the Mariner family (DNA transposons), were under-represented among CNV genes when using a relaxed threshold of ~1.52 fold. TEs from the Mariner family, one of the first identified in the fire ant (M. J. B. Krieger & Ross, 2003), have shown expansion in ant genomes (Bonasio et al., 2010) and have high horizontal gene transfer rates in both insects and mammals (Lee & Wang, 2018; Oliveira, Bao, Martins, & Jurka, 2012; Peccoud, Loiseau, Cordaux,
& Gilbert, 2017). One possible explanation for their under-representation among CNV genes is that the majority of the fire ant Mariner expansions predates the supergene, hence most of the Mariner elements were already inactive at the time of the supergene birth.
The TEs that have accumulated in the Sb genome might have been
expected to have higher gene expression, and conversely, increased copy number might have driven higher gene expression. While we found a weak positive correlation for DNA TEs between the ratio of Sb:SB copy number (i.e., the increase in copy number in Sb) with DE in SB/Sb versus SB/SB virgin queens, and with absolute gene expression in SB/Sb virgin and reproductive queens, we found none for LTR and LINE elements.
Overall, this suggests that gene expression is not the only (or even main) driver of copy number and vice versa. Rather, more factors (e.g., age of TE in genome) need to be considered to explain the difference in TE copy number between SB and Sb.
Many high copy number transposons had low gene expression levels.
One possibility is suppression by the host. As we found no correlation of gene expression with methylation levels, we suggest that suppressed gene expression of some TEs is likely by other mechanisms, such as piRNAs (Brennecke et al., 2007). Alternatively, some of the TE copies may not be expressed because they lack a promoter or have partial protein sequence.
Another potential explanation is that some TEs were co-opted in the soma, so their high expression would be uncorrelated with their germline expression level (E B Chuong et al., 2016).