Transcriptional activity, chromosomal distribution and expression effects of transposable elements in Coffea genomes

Abstract

Plant genomes are massively invaded by transposable elements (TEs), many of which are located near host genes and can thus impact gene expression. In flowering plants, TE expression can be activated (de-repressed) under certain stressful conditions, both biotic and abiotic, as well as by genome stress caused by hybridization. In this study, we examined the effects of these stress agents on TE expression in two diploid species of coffee, Coffea canephora and C. eugenioides, and their allotetraploid hybrid C. arabica. We also explored the relationship of TE repression mechanisms to host gene regulation via the effects of exonized TE sequences. Similar to what has been seen for other plants, overall TE expression levels are low in Coffea plant cultivars, consistent with the existence of effective TE repression mechanisms. TE expression patterns are highly dynamic across the species and conditions assayed here are unrelated to their classification at the level of TE class or family. In contrast to previous results, cell culture conditions per se do not lead to the de-repression of TE expression in C. arabica. Results obtained here indicate that differing plant drought stress levels relate strongly to TE repression mechanisms. TEs tend to be expressed at significantly higher levels in non-irrigated samples for the drought tolerant cultivars but in drought sensitive cultivars the opposite pattern was shown with irrigated samples showing significantly higher TE expression. Thus, TE genome repression mechanisms may be finely tuned to the ideal growth and/or regulatory conditions of the specific plant cultivars in which they are active. Analysis of TE expression levels in cell culture conditions underscored the importance of nonsense-mediated mRNA decay (NMD) pathways in the repression of Coffea TEs. These same NMD mechanisms can also regulate plant host gene expression via the repression of genes that bear exonized TE sequences.

Figure 1. Comparative proportions of distinct TE families in ESTs from C. arabica and C. canephora (data available in Tables S1 and S2 in File S1).

Figure 2. Expression levels of TE transcripts.

(A) Heatmap showing the relative expression levels of TE transcripts for 31 DNA transposons and 33 retrotransposons. CHX+: C. arabica callus treated with cycloheximide, CHX–: C. arabica callus untreated, I59_I: C. arabica irrigated leaves from drought tolerant cultivar Iapar59, I59_NI: C. arabica non-irrigated leaves from Iapar59, 14_I: C. canephora irrigated leaves from drought tolerant cultivar, 14_NI: C. canephora non-irrigated leaves from drought tolerant cultivar, Rubi_I: C. arabica irrigated leaves from drought sensitive cultivar Rubi, Rubi_NI: C. arabica non-irrigated leaves from Rubi. (B) Overall expression level differences between TEs, genes with TE insertions (TE+Genes, n = 77) and genes without TE insertions (TE–Genes, n = 63) across the conditions measured here. Average expression levels ± standard errors were compared using the Students’ t-test and the Mann-Whitney U test (MWU) as indicated. (C) Average expression levels for retrotransposons versus DNA transposons. (D) Average Manhattan distances between expression profiles within versus between TE families. (E) Individual TE sequences that have significantly up-regulated upon cycloheximide treatment (CHX+).

Figure 3. TE expression level differences for paired cultivar samples.

Overall TE expression levels are compared for cycloheximide treated (CHX+) versus untreated (CHX–) C. arabica callus and irrigated (I) versus non-irrigated (NI) leaves for drought tolerant C. arabica and C. canephora as well as drought sensitive C. arabica. Average expression levels ± standard errors were compared using the Students’ t -test and the Mann-Whitney U test (MWU) as indicated.

Figure 4. Effect exonized TEs on gene expression.

(A) Comparison of overall expression levels of genes with TE cassettes (TE+Genes, n = 77) versus genes with no TE cassettes (TE–Genes, n = 63). Average expression levels ± standard errors were compared using the Students’ t-test and the Mann-Whitney U test (MWU) as indicated. (B) Differences in overall expression levels between CHX+ and CHX– conditions for TE+Genes versus TE–Genes. Average expression levels ± standard errors were compared between CHX+ and CHX– conditions for TE+Genes and TE–Genes individually using the Students’ t-test and the Mann-Whitney U test (MWU) as indicated.

Figure 5. Chromosomal locations of TEs.

FISH using sequences of transposons MuDRA (GI311206994), Tip100 (GI 315896428) and of retrotransposon Del1 (GI 315862857) in the chromosomes of C. arabica var. typica, C. eugenioides and C. canephora. The MuDRA probe hybridized in 14 locations in C. arabica var. typica (A), with terminal, interstitial and proximal signals. Arrows indicate interstitial/proximal sites. This same probe hybridized preferentially clustered signals in C. eugenioides, with scattered signals in two pairs (D) and only clustered terminal signals in C. canephora (G). The Tip100 probe showed 36 hybridization sites in C. arabica var. typica (B), with chromosomes containing three sites (arrows) and two hybridization sites in terminal and proximal/interstitial regions (arrowheads). The same probe showed only eight chromosomes with terminal sites in C. eugenioides (E) and 14 chromosomes with signals in C. canephora (H). Note that four chromosomes exhibit two signals, being terminal and interstitial (arrows) and double terminal (arrowheads). The Del1 probe hybridized in 20 chromosomes in C. arabica var. typica (C). In only eight of them clustered signals were observed (arrows). From 12 chromosomes with signals observed in C. eugenioides (F), only two presented scattered ones (arrows). For C. canephora (I), this probe showed two pairs with scattered signals and evident terminal signals in six chromosomes (arrows). Bar represents 10 µm.