Ectopic Expression of a Maize Gene Is Induced by Composite Insertions Generated Through Alternative Transposition

Abstract

Transposable elements (TEs) are DNA sequences that can mobilize and proliferate throughout eukaryotic genomes. Previous studies have shown that in plant genomes, TEs can influence gene expression in various ways such as inserting in introns or exons to alter transcript structure and content, and providing novel promoters and regulatory elements to generate new regulatory patterns. Furthermore, TEs can also regulate gene expression at the epigenetic level by modifying chromatin structure, changing DNA methylation status and generating small RNAs. In this study, we demonstrated that Ac/fAc transposable elements are able to induce ectopic gene expression by duplicating and shuffling enhancer elements. Ac/fAc elements belong to the hAT family of Class II TEs. They can undergo standard transposition events, which involve the two termini of a single transposon, or alternative transposition events which involve the termini of two different, nearby elements. Our previous studies have shown that alternative transposition can generate various genome rearrangements such as deletions, duplications, inversions, translocations and Composite Insertions (CIs). We identified over 50 independent cases of CIs generated by Ac/fAc alternative transposition and analyzed 10 of them in detail. We show that these CIs induced ectopic expression of the maize pericarp color 2 (p2) gene, which encodes a Myb-related protein. All the CIs analyzed contain sequences including a transcriptional enhancer derived from the nearby p1 gene, suggesting that the CI-induced activation of p2 is effected by mobilization of the p1 enhancer. This is further supported by analysis of a mutant in which the CI is excised and p2 expression is lost. These results show that alternative transposition events are not only able to induce genome rearrangements, but also generate Composite Insertions that can control gene expression.

Transposon-induced inversions activate gene expression in Maize pericarp

“…We were surprised to see ears carrying p1-wwB54 produced red kernel pericarp sectors of varying sizes (Figure 1). We hypothesized that the p2 gene, which is normally not expressed in kernel pericarp, could be activated by inversions generated by Reversed Ends Transposition (RET) (Zhang and Peterson 2004;Zhang, J. and Peterson 2005;Zhang et al 2009Zhang et al , 2013Huang and Dooner 2008;Yu et al 2011;Su et al 2020). A diagram of this model showing an inversion with breakpoints in the p2 promoter region is shown in Figure 2.…”

“…Stock J (p1ww[4Co63] r1-m3::Ds) (described in Zhang et al 2003) was used as common genetic background and to detect the presence of Ac by excision of Ds from r1-m3. The occurrence of red kernel pericarp in p1-wwB54 was used as a visual screen for p2 activation in the pericarp (see Materials and Methods in Su et al 2020). p1-wwB54 has white kernels, but approximately 1 in ears were found to have a single red kernel, and ~1 in 40 ears had a multi-kernel red sector (Figure 1, F86).…”

“…Genomic DNA preparations from these lines were tested for structural rearrangements by PCR using sets of primer pairs (Table S1) that can amplify the Ac and fAc junctions in p1-wwB54: primer set 1 detects the p1/3' Ac junction, and primer set 2 detects the 5' Ac/p1/3' fAc segment (Figure 3A). Simple Ac transposition or RET-induced deletion (Yu et al, 2011) would result in negative PCR for both sets 1 and 2; while the formation of Composite Insertions (Su et al, 2020) results in retention of both junctions. Whereas, RET-induced inversion would result in retention of the p1/3' Ac junction (Set 1 positive), and loss of the 5' Ac/p1/3' fAc segment (Set 2 negative).…”

“…The markedly different proportion of inversions recovered here (14%) compared to Yu et al 2011 (53%) is most likely due to the different screens used to detect RET events. The 2011 study began with a functional p1 gene and selected for loss-of-function events, yielding mostly deletions and inversions; most Composite Insertions would not be detected because they leave the original donor locus intact (Zhang et al 2014;Su et al 2018Su et al , 2020. Whereas, this study began with a non-functional p1 allele, and required gain-offunction (red pericarp sectors).…”

“…When two transposons are in direct orientation, the internal-facing termini are present in a reversed orientation compared to the termini of a single transposon. In this configuration, the two facing termini can undergo Reversed-Ends Transposition (RET) (Huang and Dooner 2008;Zhang and Peterson 2004;Zhang et al 2009) to induce deletions (Zhang, J. and Peterson 2005;, duplications (Zhang et al 2013), Composite Insertions (Zhang et al 2014;Su et al 2018Su et al , 2020, inversions (Zhang and Peterson 2004;Yu et al 2011) and reciprocal translocations (Pulletikurti et al 2009;Zhang et al 2009). For example, Zhang et al 2009 described 17 reciprocal translocations and two large pericentric inversions derived by RET from a progenitor allele containing Ac and fAc insertions in the maize p1 gene.…”