Promoter-intrinsic and local chromatin features determine gene repression in lamina-associated domains

Abstract

It is largely unclear whether genes that are naturally embedded in lamina associated domains (LADs) are inactive due to their chromatin environment, or whether LADs are merely secondary to the lack of transcription. We show that hundreds of human promoters become active when moved from their native LAD position to a neutral context in the same cells, indicating that LADs form a repressive environment. Another set of promoters inside LADs is able to "escape" repression, although their transcription elongation is attenuated. By inserting reporters into thousands of genomic locations, we demonstrate that these escaper promoters are intrinsically less sensitive to LAD repression. This is not simply explained by promoter strength, but by the interplay between promoter sequence and local chromatin features that vary strongly across LADs. Enhancers also differ in their sensitivity to LAD chromatin. This work provides a general framework for the systematic understanding of gene regulation by repressive chromatin. Highlights:• Two promoter transplantation strategies elucidate the regulatory role of LAD chromatin• LADs are generally repressive, but also highly heterogeneous • LADs can impede both promoter activity and transcription elongation • Promoters vary intrinsically in their sensitivity to LAD repressionAlthough the majority of genes in LADs is inactive, about 10% are expressed (Guelen et al., 2008;Wu and Yao, 2017). Analysis of such exceptions to the rule may provide valuable mechanistic insights. Possibly, these genes somehow are able to overrule the (putative) repressive LAD environment, for example if their promoters are extremely strong. Alternatively, LADs may be heterogeneous, and the local chromatin context of these active genes may be less repressive or even stimulatory. It is not known which of these mechanisms -which are not mutually exclusive -allow genes to be active inside LADs.Here, we report a detailed analysis of gene repression mechanisms in LADs. In particular, we studied how promoter activity is controlled by LADs. For this purpose we made use of two orthogonal high-throughput genomic transplantation strategies. We systematically moved promoters from their native LAD location to a more neutral chromatin environment, and we also transplanted them to a wide range of locations including many LAD contexts. Combined with extensive computational analysis, the results reveal that indeed hundreds of promoters inside LADs are repressed by the local chromatin state. Moreover, the data demonstrate that features encoded in the promoter sequence, as well as strong variation in local LAD composition, determine the expression level of genes inside LADs. RESULTS Frequent repression of promoter activity in LADsOne approach to investigate whether a promoter embedded in a LAD in a certain cell type is repressed by the LAD chromatin context, is to insert the promoter into a reporter plasmid and transfect it into the same cells. This transfer to an episomal context may lead to increased activity of the promoter due t...

Identification of regulatory elements from nascent transcription using dREG

“…TIRs in H3K27me3 domains also frequently overlapped active histone marks, especially H3K4me3 and H3K4me1. Taken together, our results are consistent with recent reports that transcription start sites within heterochromatin can escape repression (Leemans et al 2018).…”

The repressive genome compartment is established early in the cell cycle before forming the lamina associated domains

“…Moreover, recent studies have demonstrated that both H3K9me2/3 and H3K27me3 are involved in LAD organization 13,18,19,22 . These findings, along with direct comparisons to Hi-C data and A/B compartment organization, have confirmed that the LADs represent a repressive genome compartment 14,[23][24][25][26] . LADs are immediately flanked by active promoters, highlighting the stark delineation between repressed LAD domains and adjacent active regions.…”

Lamina Associated Domains and Gene Regulation in Development and Cancer

“…In support of this notion, a recent genome-wide study of autonomous promoter activity has shown that LAD promoters indeed display differential sensitivity towards the local chromatin features [25]. This study employed the random integration of LAD promoters into the genome to determine the effect of chromatin context on promoter activity, and identified a class of ‘escaper’ promoters that are relatively insensitive to the LAD chromatin environment [25]. The ability of these elements to escape their repressive surroundings does not directly correlate with promoter strength, but likely depends on inherent DNA sequence features.…”

“…The differences between these and multiple other tethering approaches may relate to intrinsic differences in promoter sensitivity to embedding within the repressive LAD chromatin environment [23,24]. In support of this notion, a recent genome-wide study of autonomous promoter activity has shown that LAD promoters indeed display differential sensitivity towards the local chromatin features [25]. This study employed the random integration of LAD promoters into the genome to determine the effect of chromatin context on promoter activity, and identified a class of ‘escaper’ promoters that are relatively insensitive to the LAD chromatin environment [25].…”