Doctoral Degrees (Biochemistry)
Permanent URI for this collection
Browse
Browsing Doctoral Degrees (Biochemistry) by Subject "African trypanosomiasis"
Now showing 1 - 1 of 1
Results Per Page
Sort Options
- ItemIdentification and genomic distribution of select combinatorial post-translational modification patterns of Trypanosoma brucei histones(Stellenbosch : Stellenbosch University, 2020-03) Maree, Johannes Petrus; Patterton, H. G.; Louw, Ann; ; Stellenbosch University. Faculty of Science. Dept. of Biochemistry.ENGLISH ABSTRACT:Trypanosoma brucei is a dixenous parasitic protozoan, and the causative agent of African Sleeping sickness, or Human African Trypanosomiasis. Trypanosomes diverged very early on from the main eukaryotic lineage and display some very unusual genomic features. Pol II transcribed genes are arranged in long, non-overlapping polycistronic transcription units (PTUs) containing up to a hundred functionally unrelated genes. These PTUs are separated by strand switching regions (SSRs) which can either be divergent (dSSR) or convergent (cSSR), depending on the direction of transcription. The entire housekeeping portion on the genome is constitutively transcribed with pol II initiating from poorly defined promoters. Putative transcription start sites are epigenetically demarcated by histone post-translational modifications (PTMs) and variant deposition, with transcription initiating bi-directionally from divergent SSRs as well as internal stop/start sites (ISS) at PTU head-to-tail regions. Over the past two decades it has become increasingly clear that in Trypanosoma the epigenome has a vital role in gene regulation and transcriptional control. T. brucei lacks pol II promoter and other regulatory sequences and employs epigenetic marks to demarcate and control genomic function. More than a decade has passed since the last survey of T. brucei histone PTMs, and improvements in mass spectrometry (MS) technology and software now allows us to map PTMs with great accuracy. Application of these technologies revealed numerous histone PTMs in T. brucei, with some forming intricate combinatorial PTM patterns. The C-terminal tail of histone H2A was found to be hyperacetylated. Half of all H2A peptides observed were hyperacetylated, with up to 5 acetylated lysines occurring simultaneously. On histone H3 the lysine 10 residue was found to be trimethylated, present as a binary PTM with hydroxylated H3P40. H3HyPro40 and H2AHyPro26 were the only hydroxylated residues detected, and further analysis supports the validity of this PTM in T. brucei. Investigation of the genome-wide distribution of hyperacetylated H2A PTMs via MNase-ChIP-seq revealed that hyperacetylation of the C-terminal tail in specific forms served distinct functions and displayed different genomic localization. It was further determined that one of these states of hyperacetylation, H2AK125ac, was enriched at dSSRs and ISSs. The H2AK125ac pattern was seen to co-localize with H2A.V and RPB9, indicators of putative pol II transcription start sites. Immunofluorescence microscopy (IF) microscopy showed that different H2A hyperacetylation patterns localized to distinct nuclear foci, further emphasizing distinct functionality. The data and findings of this study provides further insight into the epigenetic mechanisms employed by T. brucei and hopefully leads to development of targeted epigenetic therapies.