Transcriptionally Driven DNA Replication Program of the Human Parasite Leishmania major

Abstract

Faithful inheritance of eukaryotic genomes requires the orchestrated activation of multiple DNA replication origins (ORIs). Although origin firing is mechanistically conserved, how origins are specified and selected for activation varies across different model systems. Here, we provide a complete analysis of the nucleosomal landscape and replication program of the human parasite Leishmania major, building on a better evolutionary understanding of replication organization in Eukarya. We found that active transcription is a driving force for the nucleosomal organization of the L. major genome and that both the spatial and the temporal program of DNA replication can be explained as associated to RNA polymerase kinetics. This simple scenario likely provides flexibility and robustness to deal with the environmental changes that impose alterations in the genetic programs during parasitic life cycle stages. Our findings also suggest that coupling replication initiation to transcription elongation could be an ancient solution used by eukaryotic cells for origin maintenance.

Well-positioned nucleosomes punctuate polycistronic pol II transcription units and flank silent VSG gene arrays in Trypanosoma brucei

“…A similar pattern of nucleosomal distribution surrounding pol II TSS has been observed in a related kinetoplastid, Leishmania major [57]. A high nucleosome occupancy (defined as the average nucleosome dyad density per base pair in a specified region), but little positioning (defined as the number of co-aligned dyads at a specific base pair), was observed across the constitutively transcribed pol II PTUs [57]. The nucleosomes downstream of the first SAS appeared less organized in the PF life cycle stage (Additional file 1: Fig.…”

“…This nucleosomal arrangement is very similar to that seen in human and yeast genomes, where nucleosomes are also depleted on polyadenylation sites [61, 62]. A similar organization was also seen in L. major , although the observed NDR was present immediately upstream of the PAS [57]. However, the relatively small number of terminal PAS elements ( n  = 35) precluded any statistically meaningful analysis of the average nucleosomal organization at this genomic position.…”

“…Although this nucleosomal arrangement is reminiscent of nucleosomes that were mapped downstream of the pol II TSSs in S. cerevisiae [55] and Drosophila [56], we note that the pol II PTUs in T. brucei are constitutively expressed, and a well-defined nucleosomal array in this region would be unexpected. A similar pattern of nucleosomal distribution surrounding pol II TSS has been observed in a related kinetoplastid, Leishmania major [57]. A high nucleosome occupancy (defined as the average nucleosome dyad density per base pair in a specified region), but little positioning (defined as the number of co-aligned dyads at a specific base pair), was observed across the constitutively transcribed pol II PTUs [57].…”

“…The nucleosome organization at the SASs and PASs therefore appears evolutionarily ancient and was probably already established before the evolutionary divergence of the Excavata from the other eukaryotic supergroups [1, 80]. The aforementioned is supported by recent findings where comparable nucleosomal patterns were observed in L. major , a related kinetoplastid with genomic synteny to T. brucei [57]. The apparent organization of nucleosomes relative to these SAS and PAS sequence elements is intriguing, as these sequence elements, and notably the oligo-dA runs, are functionally relevant for RNA processing rather than transcription.…”

“…2f). This suggests that there is a well-positioned nucleosome preferentially aligned with each internal SAS in T. brucei , contrary to the NDR observed at internal SASs in L. major [57]. When studying the nucleosomal arrangement at the SAS at the level of individual genes, the consistent, tight positioning of a nucleosome in the direct vicinity of the SAS is often observed, with a more random distribution of nucleosomes in the regions adjacent to the SAS (Additional file 1: Fig.…”