A novel SNF2 ATPase complex in Trypanosoma brucei with a role in H2A.Z-mediated chromatin remodelling

Abstract

A cascade of histone acetylation events with subsequent incorporation of a histone H2A variant plays an essential part in transcription regulation in various model organisms. A key player in this cascade is the chromatin remodelling complex SWR1, which replaces the canonical histone H2A with its variant H2A.Z. Transcriptional regulation of polycistronic transcription units in the unicellular parasite Trypanosoma brucei has been shown to be highly dependent on acetylation of H2A.Z, which is mediated by the histone-acetyltransferase HAT2. The chromatin remodelling complex which mediates H2A.Z incorporation is not known and an SWR1 orthologue in trypanosomes has not yet been reported. In this study, we identified and characterised an SWR1-like remodeller complex in T. brucei that is responsible for Pol II-dependent transcriptional regulation. Bioinformatic analysis of potential SNF2 DEAD/Box helicases, the key component of SWR1 complexes, identified a 1211 amino acids-long protein that exhibits key structural characteristics of the SWR1 subfamily. Systematic protein-protein interaction analysis revealed the existence of a novel complex exhibiting key features of an SWR1-like chromatin remodeller. RNAi-mediated depletion of the ATPase subunit of this complex resulted in a significant reduction of H2A.Z incorporation at transcription start sites and a subsequent decrease of steady-state mRNA levels. Furthermore, depletion of SWR1 and RNA-polymerase II (Pol II) caused massive chromatin condensation. The potential function of several proteins associated with the SWR1-like complex and with HAT2, the key factor of H2A.Z incorporation, is discussed.

Suramin action in African trypanosomes involves a RuvB-like DNA helicase

“…In eukaryotes, the proteins RuvB-like 1 and RuvBlike 2 are also called pontin and reptin (Jha, 2009), respectively, and they have acquired additional functions including DNA repair and chromatin remodeling (Tammana, 2017;Dauden, 2021). A chromatin remodelling complex from T. brucei, which contains TbRuvBL1 and TbRuvBL2, has been described recently (Vellmer, 2022).There is to our knowledge only one experimental study from trypanosomatids, where LmRUVBL1 and LmRUVBL2 from Leishmania major were recombinantly (co-)expressed. The purified proteins formed an elongated heterodimer with ATPase activity in vitro, but they did not form a ring (Abrahao, 2021).…”

“…In yeast the two distinct chromatin remodelling complexes, INO80 and SWR1, are well characterized (Gerhold, 2014). A SWR1-like remodeller complex in T. brucei was described recently (Vellmer, 2022). Homology models of RuvBL1 and RuvBL2 were built with high confidence using S. cerevisiae orthologs (pdb 6GEN) (Willhoft, 2018) as template.…”

Histone divergence inTrypanosoma bruceiresults in unique alterations to nucleosome structure

“…Most genes are arranged in intron-less polycistronic transcription units 20 and the chromatin context of transcription initiation and termination is in part defined by histones [22][23][24] . T. brucei histones have been found to be highly divergent 25 , and novel trypanosome-specific histone post-translational modifications 24,26 and chromatin interactors [27][28][29][30] have been identified. However, the molecular details of how local nucleosome-level chromatin structure and molecular pathways in the nucleus intersect have been largely unexplored.…”

Deciphering Divergent Trypanosomatid Nuclear Complexes by Analyzing Interactomic Datasets with AlphaFold2 and Genetic Approaches

“…Another approach to this problem was that of Jones et al, who used a proximity labelling methodology to generate an interactomic dataset of BDF5 from Leishmania mexicana ( Lmx BDF5) followed by Co-IP to validate some of the proximity hits as interactions, proposing a novel conserved regulator of the kinetoplastid transcription (CRKT) complex . Also, Vellmer et al, while trying to unveil the identity and composition of a novel SNF2 complex involved in H2A.Z deposition, arrived at the conclusion that some of the proteins they identified were forming two different HAT complexes, involving Tb HAT1 and Tb HAT2. In Staneva and Jones’ interactomic datasets, also, two distinct MORF4 related gene (MRG) domain-containing proteins were identified.…”