The Drosophila speciation factor HMR localizes to genomic insulator sites

Abstract

Hybrid incompatibility between Drosophila melanogaster and D. simulans is caused by a lethal interaction of the proteins encoded by the Hmr and Lhr genes. In D. melanogaster the loss of HMR results in mitotic defects, an increase in transcription of transposable elements and a deregulation of heterochromatic genes. To better understand the molecular mechanisms that mediate HMR’s function, we measured genome-wide localization of HMR in D. melanogaster tissue culture cells by chromatin immunoprecipitation. Interestingly, we find HMR localizing to genomic insulator sites that can be classified into two groups. One group belongs to gypsy insulators and another one borders HP1a bound regions at active genes. The transcription of the latter group genes is strongly affected in larvae and ovaries of Hmr mutant flies. Our data suggest a novel link between HMR and insulator proteins, a finding that implicates a potential role for genome organization in the formation of species.

Intricate structure of the interphase chromocenter revealed by the analysis of a factor involved in species formation

“…To confirm the previously detected centromeric localization of HMR in Drosophila cells with another antibody, we performed immunofluorescent staining using a FLAG antibody in a cell line where HMR is endogenously tagged with the FLAG epitope at the C-terminus using CRISPR/Cas9 (20). Consistent with our previous results (13) most of the FLAG signals co-localize with the signal we obtained when using an anti-dCenpA antibody, which confirms the close proximity of HMR and centromeric chromatin during interphase (Figure 1A).…”

“…This suggests that the localization of HMR to the centromere is either cell cycle regulated or specific for a subset of centromeres (Figure 1B), which may explain the differences observed when staining HMR in different tissues (13, 15). Another observation that is difficult to reconcile with the clustered staining of HMR in cells is the fact that ChIP-seq data suggest HMR binding along the euchromatic arms often at boundaries between HP1a containing heterochromatin and actively transcribed genes (20). As centromeres are also transcribed (10) and in proximity to pericentromeric heterochromatin, we wondered if HMR might also localize to a putative boundary between these two different chromatin regions.…”

“…HMR interacts with the heterochromatin protein HP1a and co-localizes with the centromere-specific H3 variant dCenpA (CID) in Drosophila cell lines and imaginal disc cells (13). HMR binding is also observed at several euchromatic sites where it colocalizes with known boundary factors (20). Interestingly, the intracellular localization of HMR varies among different tissues.…”

Altered chromatin localization of hybrid lethality proteins inDrosophila

“…Chromatin immunoprecipitation was essentially performed as in Gerland et al, 2017(Gerland et al 2017. For each ChIP reaction, chromatin isolated from 1-2 x 10 6 cells was incubated with rat anti-HMR 2C10 antibody pre-coupled to Protein A/G Sepharose through a rabbit IgG anti-rat.…”

“…Peak calling was performed using HOMER 4.9 with parametersstyle factor -F 2 -size 200 for HMR and -style factor -F 6 -L 6 -size 200 for GFZF. Downstream analysis steps were performed using R. The endogenous HMR binding data set (GSE86106, Gerland et al, 2017) and the GFZF binding data set (GSE105009, Baumann et al, 2017) are derived from NCBI GEO.…”