Role of the RNA-binding protein ZC3H41 in the regulation of ribosomal protein messenger RNAs in trypanosomes

Background: Trypanosomes are single-celled eukaryotes that rely heavily on post-transcriptional mechanisms to regulate gene expression. RNA-binding proteins play essential roles in regulating the fate, abundance and translation of messenger RNAs (mRNAs). Among these, zinc finger proteins of the cysteine3histidine (CCCH) class have been shown to be key players in cellular processes as diverse as differentiation, regulation of the cell cycle and translation. ZC3H41 is an essential zinc finger protein that has been described as a component of spliced leader RNA granules and nutritional stress granules, but its role in RNA metabolism is unknown. Methods: Cell cycle analysis in ZC3H41- and Z41AP-depleted cells was carried out using 4′,6-diamidino-2-phenylindole staining, microscopic examination and flow cytometry. The identification of ZC3H41 protein partners was done using tandem affinity purification and mass spectrometry. Next-generation sequencing was used to evaluate the effect of ZC3H41 depletion on the transcriptome of procyclic Trypanosoma brucei cells, and also to identify the cohort of mRNAs associated with the ZC3H41/Z41AP complex. Levels of 5S ribosomal RNA (rRNA) species in ZC3H41- and Z41AP-depleted cells were assessed by quantitative reverse transcription-polymerase chain reaction. Surface sensing of translation assays were used to monitor global translation. Results: We showed that depletion of the zinc finger protein ZC3H41 resulted in marked cell cycle defects and abnormal cell morphologies. ZC3H41 was found associated with an essential protein, which we named Z41AP, forming a stable heterodimer, and also with proteins of the poly(A)-binding protein 1 complex. The identification of mRNAs associated with the ZC3H41/Z41AP complex revealed that it is primarily composed of ribosomal protein mRNAs, and that binding to target transcripts is diminished upon nutritional stress. In addition, we observed that mRNAs encoding several proteins involved in the maturation of 5S rRNA are also associated with the ZC3H41/Z41AP complex. Finally, we showed that depletion of either ZC3H41 or Z41AP led to the accumulation of 5S rRNA precursors and a decrease of protein translation. Conclusions: We propose that ZC3H41 and Z41AP play important roles in controlling the fate of ribosomal components in response to environmental cues. Graphical Abstract: [Figure not available: see fulltext.]Open Access funding provided thanks to the CRUE-CSIC agreement with Springer Nature. This work was funded by the Spanish National Research Council (CSIC) (grant 201920E114 to AME)

Trypanosomiasis and Leishmaniasis Symposium: Advances in Basic and Applied Research

About the British Society for Parasitology. About.Today many researchers and students are very passionate about the fascinating world of infectious diseases, parasites, their complex lifestyles and their associated impact on people and livelihoods.To draw attention to the unique importance of parasitology as a distinct discipline within biology, The British Society for Parasitology was formed in 1962 from the Parasitological Section of the Institute of Biology. Today the Society is the central networking and meeting point for many professional and amateur parasitologists throughout the UK and across the world.Did you know that the UK leads Europe – and Europe leads the world – in parasitology research? No European country publishes more parasitology research than the UK, and UK papers were cited more than those from any other country in the past 5 years (2011-2016, data from Elsevier SciVal, see News item for more details.)As the leading academic society for a country preeminent in parasitology, the remit of the BSP is broad. It promotes and supports the academic study of parasitology in all its many guises. This can be from experimental to theoretical approaches as applied to infection biology and disease research, or from ecological to medical and veterinary studies in global health and international aid. Each year students are given financial support to attend BSP meetings and scholarship schemes are in place to support fieldwork and training events.The membership of the BSP stands at around 1000 in number. Approximately a third of members are from overseas locations. Highlights of the annual BSP calendar include the annual residential meetings in spring and autumn which are focused upon general and specialist aspects of parasitology. The BSP has a close relationship with Cambridge University Press that prints a special issue on our autumn meeting.The BSP Society is a Charitable Incorporated Organisation and managed by the BSP Council. This comprises a President, Honorary Officers and ordinary Council Members, who together act as Company Trustees. Co-opted members of Council also include representation from the student, early career membership and other learned societies where clear synergies are apparent

Identification of elusive sequence-specific promoters of RNA polymerase II polycistronic transcription in African trypanosomes

Kinetoplastids have evolved in isolation for one billion years resulting in several divergent molecular and cellular processes. One example is protein-coding genes transcribed polycistronically by a typical RNA polymerase II (RNA pol II). Transcription most likely starts at divergent Strand Switch Regions (dSSRs), long sequences between divergently oriented polycistronic transcription units (PTUs). The lack of regulation in trypanosome transcription has become the paradigm in our field. Previous work suggests that changes in chromatin structure over broad SSR regions drives unregulated and dispersed transcription initiation. We investigate such an exceptional feature in trypanosomes by first identifying RNA pol II-enriched regions using ChIP-Seq, as potential promoter sequences. The high resolution of this technique allowed us to accurately determine peaks of RNA pol II accumulation in the dSSRs. To functionally investigate pol II-enriched sequences unbiasedly, the peaks on chromosome VII were assayed for their ability to direct transcription using transient transfection. This analysis suggests that two unidirectional short sequence specific promoters within each dSSR make up the general structure. Primer extension analysis of nascent RNA allowed us to identify precise transcription start sites (TSS) of promoters inserted in a chromosome. Detailed analysis of one of these promoters defined 75bp as sufficient to fully drive transcription and identified essential nucleotides for precise initiation around the TSS. In addition, mutations to internal and downstream boxes led to dramatic decreased activity. In summary, we show that sequence-specific unidirectional RNA pol II promoters with proper TSS transcription initiation are present in the T. brucei genome. Our results challenge the currently accepted hypothesis that trypanosomes lack true promoters with transcription initiation control