Abstract
Trypanosoma cruzi alternates between replicative and nonreplicative life forms, accompanied by a shift in global transcription levels and by changes in the nuclear architecture, the chromatin proteome and histone posttranslational modifications. To gain further insights into the epigenetic regulation that accompanies life form changes, we performed genome-wide high-resolution nucleosome mapping using two T. cruzi life forms (epimastigotes and cellular trypomastigotes). By combining a powerful pipeline that allowed us to faithfully compare nucleosome positioning and occupancy, more than 125 thousand nucleosomes were mapped, and approximately 20% of them differed between replicative and nonreplicative forms. The nonreplicative forms have less dynamic nucleosomes, possibly reflecting their lower global transcription levels and DNA replication arrest. However, dynamic nucleosomes are enriched at nonreplicative regulatory transcription initiation regions and at multigenic family members, which are associated with infective-stage and virulence factors. Strikingly, dynamic nucleosome regions are associated with GO terms related to nuclear division, translation, gene regulation and metabolism and, notably, associated with transcripts with different expression levels among life forms. Finally, the nucleosome landscape reflects the steady-state transcription expression: more abundant genes have a more deeply nucleosome-depleted region at putative 5' splice sites, likely associated with trans-splicing efficiency. Taken together, our results indicate that chromatin architecture, defined primarily by nucleosome positioning and occupancy, reflects the phenotypic differences found among T. cruzi life forms despite the lack of a canonical transcriptional control context.
Highlights
Chromatin is the template of essential cellular processes such as transcription, replication and repair, and its structure and organization must be finely regulated
To investigate the possible impact of epigenetic changes based on nucleosome positioning and occupancy on the phenotypic differences observed among life forms, we performed MNase-seq in two life forms of this parasite
Chromatin architecture differs at strategic genomic regions in Trypanosoma cruzi In short, chromatin from epimastigotes and tissue cultured derived trypomastigote (TCT) was digested with the micrococcal nuclease enzyme, which preferentially cleaves the linker DNA, leaving mononucleosomes (S1A Fig)
Summary
Chromatin is the template of essential cellular processes such as transcription, replication and repair, and its structure and organization must be finely regulated. Nucleosome positioning (where nucleosomes are located with respect to the genomic DNA sequence) and occupancy (local nucleosome density in a cell population) are important features of chromatin organization and influence crucial aspects of epigenetic regulation [1]. Their positioning is governed both by intrinsic DNA sequences and by trans-factors such as ATP-dependent remodelers, transcription factors and the RNA Polymerase elongation [2,3]. A unique nucleosome architecture around the transcription start sites (TSSs) is found, comprising a nucleosome-depleted region (NDR) just upstream of the TSS, followed by at least three wellpositioned nucleosomes downstream [6]. Single-cell analysis of MNase-seq data has started to clarify the association of nucleosome occupancy and transcription levels: uniformly spaced but poorly positioned nucleosomes were found at silent genes, whereas well-positioned but irregularly spaced nucleosomes were found at active genes [8]
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