Targeting of P-Element Reporters to Heterochromatic Domains by Transposable Element 1360 in Drosophila melanogaster

Abstract

Heterochromatin is a common DNA packaging form employed by eukaryotes to constitutively silence transposable elements. Determining which sequences to package as heterochromatin is vital for an organism. Here, we use Drosophila melanogaster to study heterochromatin formation, exploiting position-effect variegation, a process whereby a transgene is silenced stochastically if inserted in proximity to heterochromatin, leading to a variegating phenotype. Previous studies identified the transposable element 1360 as a target for heterochromatin formation. We use transgene reporters with either one or four copies of 1360 to determine if increasing local repeat density can alter the fraction of the genome supporting heterochromatin formation. We find that including 1360 in the reporter increases the frequency with which variegating phenotypes are observed. This increase is due to a greater recovery of insertions at the telomereassociated sequences (50% of variegating inserts). In contrast to variegating insertions elsewhere, the phenotype of telomere-associated sequence insertions is largely independent of the presence of 1360 in the reporter. We find that variegating and fully expressed transgenes are located in different types of chromatin and that variegating reporters in the telomere-associated sequences differ from those in pericentric heterochromatin. Indeed, chromatin marks at the transgene insertion site can be used to predict the eye phenotype. Our analysis reveals that increasing the local repeat density (via the transgene reporter) does not enlarge the fraction of the genome supporting heterochromatin formation. Rather, additional copies of 1360 appear to target the reporter to the telomere-associated sequences with greater efficiency, thus leading to an increased recovery of variegating insertions. KEYWORDS heterochromatin; transposable elements; position-effect variegation; chromatin; Drosophila I N vivo regulation of gene expression occurs in the context of chromatin, the complex structure formed by DNA, histones, and a variety of associated proteins. Two basic types of chromatin were originally distinguished, euchromatin and heterochromatin, based on cytological staining behavior during the cell cycle (Heitz 1928). Euchromatin and heterochromatin differ in a number of characteristics, including biochemical makeup and their effect on the expression of transgene reporters. Euchromatin is generally accessible for transcription, contains the majority of the genes, and is characterized by biochemical marks associated with transcription, such as high levels of histone acetylation, transcriptional activators, and RNA polymerase II. In contrast, heterochromatin contains few genes and has a high repeat density. Heterochromatin is characterized by low levels of histone acetylation, high levels of histone 3 lysine 9 (H3K9) methylation, and the presence of "silencing" proteins such as heterochromatin protein 1 (HP1) (Beisel and Paro 2011). While this model of two basic chromatin types has been very successful...

Short and long-term evolutionary dynamics of subtelomeric piRNA clusters in Drosophila

“…Such similarities between TAS-R and TAS-L families have been suggested recently. 9 , 10 In this study, we specify these similarities showing that TAS-L repeats consists of six tandemly-repeated sequences of 147, 53, 29, 44, 45 and 20 bp in length that share over 80% sequence similarity with the 158 bp region identified in the T3 domain of X-TAS ( Fig. 1E ).…”

“… 18 TAS-R, especially X-TAS , appear to be piRNA clusters, into which P elements (as well as P -derived transgenes) can insert. 1 , 10 , 19 Once inserted, P sequences are included in the piRNA produced and these piRNA will participate to repression of euchromatic P copies in the germline leading to the so-called TSE . 20–23 …”

EvoChromo: towards a synthesis of chromatin biology and evolution

“…The formation of nucleosomes, the presence of specific histone marks and the binding of transcription factors have been found to affect the incidence of DNA lesions, as well as DNA repair efficacy, leading to chromatin-dependent variability in mutation rates across the genome, which can also be observed as local differences in variation at the population level (Makova and Hardison, 2015). Along similar lines, chromatin affects the insertion probabilities of TEs, thus influencing how selfish genetic elements spread across the genome and also perhaps how genes spread horizontally across genomes (Huisinga et al, 2016;Lesage and Todeschini, 2005;Quadrana et al, 2018 preprint). In a likely related manner, the nature of chromatin strongly affects recombination landscapes (Székvölgyi et al, 2015).…”

The Impact of Genetic Background and Cell Lineage on the Level and Pattern of Position Effect Variegation

“…are common to most variegating lines [10], although individual heterochromatic domains can show differences in sensitivity to a subset of the known suppressors of variegation [12][13][14]. A major exception are insertions into the TAS (Telomere Associated Sequences) sequences, just proximal to HeT-A and TART; these lines exhibit a PEV phenotype that is sensitive to mutations in the Polycomb silencing machinery [15], while ChIP analysis shows association with Pc [16]. Here we have used as our test locus a reporter in the fourth chromosome, a largely heterochromatic domain that for the most part mimics pericentric heterochromatin, a chromatin structure dependent on H3K9 methylation and HP1a [10,12].…”