Close

A systematic analysis of Trypanosoma brucei chromatin factors identifies novel protein interaction networks associated with sites of transcription initiation and termination

Desislava P. Staneva, Roberta Carloni, Tatsiana Auchynnikava, Pin Tong, Juri Rappsilber, A. Arockia Jeyaprakash, Keith R. Matthews, Robin C. Allshire

Preprint posted on 10 February 2021 https://doi.org/10.1101/2021.02.09.430399

Article now published in Genome Research at http://dx.doi.org/10.1101/gr.275368.121

A whole new world of chromatin regulation in Trypanosoma brucei. @dpstaneva, @robbiecarloni and others reveal new networks that control transcription in this early-branching eukaryote.

Selected by Tamara Sternlieb

Some context

Gene expression in eukaryotic organisms is regulated at several levels, the first of which is the chromatin state and its effect on transcription. Heterochromatin is compacted and repressive for transcription, while euchromatin is relaxed and transcriptionally active. These chromatin states are regulated by several modifications on DNA and on histones, the proteins around which DNA is wrapped. The dynamics of these modifications and their effect on transcription are well studied in eukaryotic model organisms, such as yeast, plants and animals. However, the Eukarya domain also includes early-branching organisms, such as protozoa parasites like Trypanosoma brucei, which have evolved for over 500 million years, diverging from the most studied organisms.

T. brucei is the causative agent of sleeping sickness in humans and nagana in cattle. Unusually for a eukaryote, most trypanosome genes are transcribed in polycistronic units, which are then resolved by trans-splicing. RNA Polymerase II begins transcription at divergent Transcription Start Regions (TSR) on the chromatin, and it typically terminates at convergent transcription regions called Transcription Termination Regions (TTR). Less frequently, transcription units are arranged head to tail. Transcription termination in such units often coincides with genes transcribed by RNA Polymerase I or RNA Polymerase III. All these regions show distinct histone variant and histone modification enrichment.

Although the general consensus is that trypanosome gene expression is mostly regulated post-transcriptionally via control of RNA stability and translation, trypanosome genomes encode for several chromatin regulator sequences, which haven’t been thoroughly studied and whose function is unknown.

 

Key Findings

The authors interrogated the TriTrypDB trypanosome genome database, searching for writers, readers and erasers of chromatin post-translational modifications (PTMs), specifically histone acetylation and methylation. They identified 68 proteins, of which some had been explored previously. They proceeded with a three-step analysis that revealed the intracellular localization, chromatin enrichment and protein interaction networks of the chromatin-associated factors.

They endogenously tagged the candidate proteins with YFP and first evaluated their cellular localization. Then they performed ChIP-seq to evaluate their chromatin association and enrichment in convergent and divergent TSRs and TTRs. They used genomic annotations and H2A.Z histone ChIP enrichement to identify these regions.

15 of the candidate proteins were specifically enriched at TSRs, of which 10 had not been shown to act at this region before. These proteins revealed two enrichment patterns, which the authors named Class I and Class II TSR-associated factors, characterized by sharp peaks and a broader pattern around TSRs, respectively. Class I and Class II factors may play different roles in RNAPII transcription, as initiators and facilitators of RNA Polymerase processivity, respectively. Using Immunoprecipitation and mass spectrometry, they identified interacting proteins that may participate in the RNAPII regulation networks, including factors characterized in other eukaryotes and new players.

The ChIP-seq analysis also revealed 8 proteins which displayed specific enrichment over a subset of RNAPII TTRs. They were also enriched at small nuclear RNA and tRNA genes, which are transcribed by RNAPIII. The authors’ hypothesis is that these proteins block the passage of RNAPII over genes transcribed by RNAPIII and could facilitate transcription of snRNA and tRNA genes.

While TSR-associated factors showed several reciprocal interactions, none of the eight TTR associated proteins interact with each other. The analysis of their interaction networks confirmed that these might have different functions, including altering nucleosome density, resolving R-loops and dual roles promoting RNAPII Splice Leader and RNAPIII transcription or termination of RNAPII transcription.

 

How I believe this moves the field forward

This is an extensive and thorough exploration of the localization and function of known and unknown proteins potentially regulating T. brucei chromatin organization and regulation. I believe the analysis of association networks for each protein opens up many new research questions to explore and will prime new discoveries that further our understanding of gene expression control in trypanosomes.

Chromatin regulation has been given a very high profile in model eukaryotic organisms and thus has been profoundly studied. Since trypanosome transcription is polycistronic, the levels of transcription regulation have been quite a challenge to explore in these organisms and RNA post-transcriptional modulation took up the protagonist role. I admire this study which shows that we shouldn’t underestimate chromatin regulation in early branching organisms.

 

Open questions

You mention there is some colocalization of TSRs associated factors and TTRs factors on the chromatin, but the significance of this colocalization is unknown. Do you have any new findings or hypothesis regarding these?

Authors response:

We currently don’t know why TSR and TTR factors might be enriched in the same locations. One speculations could be that the TTR factors are necessary to terminate any spurious transcription originating at TSRs, possibly in the wrong direction. That would of course not apply to bidirectional promoters which lead to the production of stable transcripts in both directions. RNAPII TSR factors might also be important for expression of RNAPIII-transcribed tRNAs and snRNAs.

On another note, while we call some proteins ”TTR factors”, we are not sure whether they are important for RNAPII transcription termination or for regulating RNAPIII transcription of tRNAs and snRNAs.

Some of the proteins that presented some form of chromatin enrichment show both nuclear and cytoplasmic localizations when tagged. Do you believe there could be some form of nuclear import/export regulation to their function?

Authors response:

We think it is possible that some proteins are regulated by import in/export out of the nucleus under different conditions, such as developmental transitions. Additionally, it is possible that some proteins have dual independent functions in the nucleus and the cytoplasm. However, at present we do not have data to support or reject these hypotheses.

Tags: chromatin, evolution, parasites

Posted on: 11 March 2021 , updated on: 26 March 2021

doi: https://doi.org/10.1242/prelights.27646

Read preprint (No Ratings Yet)

Have your say

Your email address will not be published.

This site uses Akismet to reduce spam. Learn how your comment data is processed.

Sign up to customise the site to your preferences and to receive alerts

Register here

preLists in the cell biology category:

Fibroblasts

The advances in fibroblast biology preList explores the recent discoveries and preprints of the fibroblast world. Get ready to immerse yourself with this list created for fibroblasts aficionados and lovers, and beyond. Here, my goal is to include preprints of fibroblast biology, heterogeneity, fate, extracellular matrix, behavior, topography, single-cell atlases, spatial transcriptomics, and their matrix!

 



List by Osvaldo Contreras

EMBL Synthetic Morphogenesis: From Gene Circuits to Tissue Architecture (2021)

A list of preprints mentioned at the #EESmorphoG virtual meeting in 2021.

 



List by Alex Eve

FENS 2020

A collection of preprints presented during the virtual meeting of the Federation of European Neuroscience Societies (FENS) in 2020

 



List by Ana Dorrego-Rivas

Planar Cell Polarity – PCP

This preList contains preprints about the latest findings on Planar Cell Polarity (PCP) in various model organisms at the molecular, cellular and tissue levels.

 



List by Ana Dorrego-Rivas

BioMalPar XVI: Biology and Pathology of the Malaria Parasite

[under construction] Preprints presented at the (fully virtual) EMBL BioMalPar XVI, 17-18 May 2020 #emblmalaria

 



List by Dey Lab, Samantha Seah

1

Cell Polarity

Recent research from the field of cell polarity is summarized in this list of preprints. It comprises of studies focusing on various forms of cell polarity ranging from epithelial polarity, planar cell polarity to front-to-rear polarity.

 



List by Yamini Ravichandran

TAGC 2020

Preprints recently presented at the virtual Allied Genetics Conference, April 22-26, 2020. #TAGC20

 



List by Maiko Kitaoka et al.

3D Gastruloids

A curated list of preprints related to Gastruloids (in vitro models of early development obtained by 3D aggregation of embryonic cells). Updated until July 2021.

 



List by Paul Gerald L. Sanchez and Stefano Vianello

ECFG15 – Fungal biology

Preprints presented at 15th European Conference on Fungal Genetics 17-20 February 2020 Rome

 



List by Hiral Shah

ASCB EMBO Annual Meeting 2019

A collection of preprints presented at the 2019 ASCB EMBO Meeting in Washington, DC (December 7-11)

 



List by Madhuja Samaddar et al.

EMBL Seeing is Believing – Imaging the Molecular Processes of Life

Preprints discussed at the 2019 edition of Seeing is Believing, at EMBL Heidelberg from the 9th-12th October 2019

 



List by Dey Lab

Autophagy

Preprints on autophagy and lysosomal degradation and its role in neurodegeneration and disease. Includes molecular mechanisms, upstream signalling and regulation as well as studies on pharmaceutical interventions to upregulate the process.

 



List by Sandra Malmgren Hill

Lung Disease and Regeneration

This preprint list compiles highlights from the field of lung biology.

 



List by Rob Hynds

Cellular metabolism

A curated list of preprints related to cellular metabolism at Biorxiv by Pablo Ranea Robles from the Prelights community. Special interest on lipid metabolism, peroxisomes and mitochondria.

 



List by Pablo Ranea Robles

BSCB/BSDB Annual Meeting 2019

Preprints presented at the BSCB/BSDB Annual Meeting 2019

 



List by Dey Lab

MitoList

This list of preprints is focused on work expanding our knowledge on mitochondria in any organism, tissue or cell type, from the normal biology to the pathology.

 



List by Sandra Franco Iborra

Biophysical Society Annual Meeting 2019

Few of the preprints that were discussed in the recent BPS annual meeting at Baltimore, USA

 



List by Joseph Jose Thottacherry

ASCB/EMBO Annual Meeting 2018

This list relates to preprints that were discussed at the recent ASCB conference.

 



List by Dey Lab, Amanda Haage
Close