Retrotransposons Down- and Up-Regulation in Aging Somatic Tissues

Abstract

The transposon theory of aging hypothesizes the activation of transposable elements (TEs) in somatic tissues with age, leading to a shortening of the lifespan. It is thought that TE activation in aging produces an increase in DNA double-strand breaks, contributing to genome instability and promoting the activation of inflammatory responses. To investigate how TE regulation changes in somatic tissues during aging, we analyzed the expression of some TEs, as well as a source of small RNAs that specifically silence the analyzed TEs; the Drosophila cluster named flamenco. We found significant variations in the expression levels of all the analyzed TEs during aging, with a trend toward reduction in middle-aged adults and reactivation in older individuals that suggests dynamic regulation during the lifespan.

Keywords: Idefix; ZAM; gypsy; longevity; transposons.

Figure 1

gypsy, ZAM and Idefix retrotransposons are under the regulation of flamenco in head somatic tissues. (A,B) Gene expression analysis in RNAs isolated from female head tissues from Canton S (CS), flam^A /flam^A, w^IR6RevII7/w^IR6RevII7 and flam^BG /flam^BG strains. The fold changes indicate the expression levels relative to calibrator (gypsy in CS). (A) Expression analysis of gypsy, ZAM and Idefix. (B) Expression analysis of flamenco. Shown are average levels (n = 3), and error bars indicate SD. Significance was determined by one-way ANOVA with Tukey post hoc test.

Figure 2

Analysis of the expression of the selected retrotransposons and of flamenco in aging. (A) Schematic representation of the genetic cross performed to obtain individuals permissive and non-permissive for the activation of gypsy. (B) Analysis of the expression of gypsy in RNAs isolated from 3-days-old ovaries of flam^A/Df(1)l11 and flam^A/flam^FM7 adult females. The fold changes indicate the expression levels relative to calibrator (flam^A/flam^FM7) (C–F) Gene expression analysis in RNAs isolated from head tissues from flam^A/Df(1)l11 and flam^A/flam^FM7 adult females on Day 1 (young), Day 30 (middle-aged) and Day 50 (old). The fold changes indicate the expression levels relative to calibrator (gypsy in 1 day-old flam^A/flam^FM7) (C) gypsy expression. A two-way ANOVA reveals a significant effect of genotype (p = 0.015) and of age (p = 0.001). (D) ZAM expression. A two-way ANOVA reveals a significant effect of genotype (p = 0.002) and of age (p = 0.001). (E) Idefix expression. A two-way ANOVA reveals a significant effect of age (p = 0.009). (F) flam1 expression. A two-way ANOVA reveals a significant effect of genotype/age (p = 0.011) and of age (p = 0.001). (G) flam5 expression. A two-way ANOVA reveals a significant effect of genotype/age (p = 0.007), of genotype (p = 0.004) and of age (p = 0.034). Shown are average levels (n = 3), and error bars indicate SD. Comparison of two groups with different ages was done by applying a LSD post hoc analysis. Pairwise comparisons between genotypes of the same age were done using a two tails unpaired t-test. (H) Kaplan-Meier survival curves of Df(1)l11/flam^A and flam^FM7C/flam^A adult females.

Figure 3

A speculative model for gypsy regulation during the aging process. flamenco is highly enriched in antisense TE sequences, comprising gypsy fragments. The flamenco primary transcript can form double strand RNAs (dsRNAs) with gypsy sense transcripts that serve as substrates for the production of endo-siRNAs specific for the silencing of gypsy. In young flies, gypsy expression level depends on its transcriptional rate and on post-transcriptional silencing induced by specific endo-siRNAs (gypsy-antisense). In middle age, reduction in gypsy and flamenco expression can be explained by an increased production of endo-siRNAs (sense and antisense) specific for the silencing of gypsy and gypsy fragments that are part of the flamenco transcript (cosuppression). In old flies, gypsy and flamenco appeared derepressed, and this can be explained by a reduction of silencing, probably due to a reduction in the amount of endo-siRNAs (sense and antisense). Similar mechanisms can be hypothesized to explain down- and up- regulation of ZAM and Idefix in aging. Right boxes: a legend is represented in the upper box; correlation between the abundance of gypsy transcription and mortality in P(permissive) and NP (non-permissive) flies in middle aged individuals (middle age box) and old individuals (old box).