HAT2 mediates histone H4K4 acetylation and affects micrococcal nuclease sensitivity of chromatin in Leishmania donovani

Abstract

Histone post-translational modifications (PTMs) such as acetylation and methylation are known to affect chromatin higher order structures. Primary targets of these modifications include basic residues present at N-terminus tail region of core histones. Four histone acetyltransferase (HAT) genes have been identified in trypanosomatids. HAT1, HAT3 and HAT4 of Leishmania donovani have been partially characterized. However, there is no report about HAT2 of Leishmania donovani. Lysine residues present on the N-terminal tail of Leishmania donovani histone H4 are conserved in other trypanosomatids and humans. PTMs of lysines modulate various functions at chromatin level. The four histone acetyltransferases encoded in Leishmania genome were over-expressed to analyse their functional activity. All four HATs were found actively acetylating core histones H3/H4. Similar to L. donovani HAT3 and HAT4, HAT2 was found to be a member of MYST family protein and have SAS2 type domain. Over-expression of HAT2 significantly increases acetylation of H4K4. To analyse the effect of HAT2 over-expression on chromatin accessibility, micrococcal nuclease digestion assay was performed. MNase digestion resulted in a higher proportion of the mononucleosomes and dinucleosomes in HAT2 over-expressing cells as compared to WT L. donovani cells. Acetylation of lysine-4 neutralizes the amino terminal region of histone H4. This weakens its interaction with neighbouring nucleosomes and the linker DNA. HAT2 over-expression in L. donovani resulted in highly accessible chromatin suggesting chromatin decondensation. HAT2 may have an important role to play in global regulation of transcription in L. donovani. Better understanding of these epigenetic determinants of parasite would help in designing novel therapeutic strategies.

Diverse DNA Sequence Motifs Activate Meiotic Recombination Hotspots Through a Common Chromatin Remodeling Pathway

“…, Figure 3B, 0 hr), might be due to the removal of nucleosomes in a subset of cells, or by looser wrapping of DNA around nucleosomes, or some combination of the two. For example, histone PTMs (such the acetylation of lysine residues) can affect how tightly DNA is wrapped around nucleosomes, and, consequently, sensitivity of that DNA to MNase (Zhao et al 2014; Lorzadeh et al 2016; Jha et al 2017; Kurup et al 2019). Notably, at least 23 different combinations of histone PTMs are enriched preferentially around the ade6-M26 hotspot at one or more time points of meiosis, relative to basal recombination control (Storey et al 2018).…”

Application of Fluorescence Lifetime Imaging Microscopy of DNA Binding Dyes to Assess Radiation-Induced Chromatin Compaction Changes

“…Due to this widespread impact in biology and particularly in many human diseases, improving the methods to study chromatin compaction is highly relevant. So far, various approaches have been utilized to address chromatin organization, e.g., nuclease digestion assays [ 37 , 38 ], DNA denaturation dependent staining using Acridine Orange in fixed cells [ 39 ], different forms of electron or super-resolution microscopy utilizing DNA dyes or antibodies [ 40 , 41 , 42 , 43 , 44 ], or fluorescence intensity based measurements [ 45 ]. Recently, also FLIM has been applied to get insights into the chromatin structure and its reaction upon different stimuli [ 23 , 24 , 25 , 26 , 27 , 28 , 29 , 31 ].…”

Oxidative Stress‐Mediated Overexpression of Uracil DNA Glycosylase in Leishmania donovani Confers Tolerance against Antileishmanial Drugs

“…The LdUNG ORF was also cloned into pLpneo2, which is a suitable expression vector for leishmania, and it was transfected into sensitive laboratory strain (Ag83) by electroporation at 25 μ F, 1500 V (3.75 kV/cm) with a gap of 10 seconds between pulses using Gene Pulser X-Cell (Bio-Rad) [ 21 ]. Parasites were transfected with pLpneo2 alone for control.…”