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Roles of the Pumilio domain protein PUF3 in Trypanosoma brucei growth and differentiation

Published online by Cambridge University Press:  09 June 2020

K. Kamanyi Marucha  and
C. Clayton Open the ORCID record for C. Clayton [Opens in a new window]
K. Kamanyi Marucha
Affiliation:
Heidelberg University Centre for Molecular Biology (ZMBH), Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany
C. Clayton*
Affiliation:
Heidelberg University Centre for Molecular Biology (ZMBH), Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany
*
Author for correspondence: C. Clayton, E-mail: cclayton@zmbh.uni-heidelberg.de
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Abstract

Trypanosomes strongly rely on post-transcriptional mechanisms to control gene expression. Several Opisthokont Pumilio domain proteins are known to suppress expression when bound to mRNAs. The Trypanosoma brucei Pumilio domain protein PUF3 is a cytosolic mRNA-binding protein that suppresses expression when tethered to a reporter mRNA. RNA-binding studies showed that PUF3 preferentially binds to mRNAs with a classical Pumilio-domain recognition motif, UGUA[U/C]AUU. RNA-interference-mediated reduction of PUF3 in bloodstream forms caused a minor growth defect, but the transcriptome was not affected. Depletion of PUF3 also slightly delayed differentiation to the procyclic form. However, both PUF3 genes could be deleted in cultured bloodstream- and procyclic-form trypanosomes. Procyclic forms without PUF3 also grew somewhat slower than wild-type, but ectopic expression of C-terminally tagged PUF3 impaired their viability. PUF3 was not required for RBP10-induced differentiation of procyclic forms to bloodstream forms. Mass spectrometry revealed no PUF3 binding partners that might explain its suppressive activity. We conclude that PUF3 may have a role in fine-tuning gene expression. Since PUF3 is conserved in all Kinetoplastids, including those that do not infect vertebrates, we suggest that it might confer advantages within the invertebrate host.

Keywords

mRNA decayPumiliotranslationTrypanosoma brucei
Type
Research Article
Information
Parasitology , Volume 147 , Issue 11 , September 2020 , pp. 1171 - 1183
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Copyright
Copyright © The Author(s), 2020. Published by Cambridge University Press

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Footnotes

*

Current address: Kisii University Medical Biochemistry, P.O. Box 408, 40200 Kisii, Kenya.

References

Abbasi, N, Park, YI and Choi, SB (2011) Pumilio Puf domain RNA-binding proteins in Arabidopsis. Plant Signaling & Behavior 6, 364–368.CrossRefGoogle ScholarPubMed
Alibu, VP, Storm, L, Haile, S, Clayton, C and Horn, D (2004) A doubly inducible system for RNA interference and rapid RNAi plasmid construction in Trypanosoma brucei. Molecular and Biochemical Parasitology 139, 75–82.CrossRefGoogle Scholar
Alsford, S, Turner, D, Obado, S, Sanchez-Flores, A, Glover, L, Berriman, M, Hertz-Fowler, C and Horn, D (2011) High throughput phenotyping using parallel sequencing of RNA interference targets in the African trypanosome. Genome Research 21, 915–924.CrossRefGoogle ScholarPubMed
Antwi, E, Haanstra, J, Ramasamy, G, Jensen, B, Droll, D, Rojas, F, Minia, I, Terrao, M, Mercé, C, Matthews, K, Myler, P, Parsons, M and Clayton, C (2016) Integrative analysis of the Trypanosoma brucei gene expression cascade predicts differential regulation of mRNA processing and unusual control of ribosomal protein expression. BMC Genomics 17, 306.CrossRefGoogle ScholarPubMed
Archer, SK, van Luu, D, de Queiroz, R, Brems, S and Clayton, CE (2009) Trypanosoma brucei PUF9 regulates mRNAs for proteins involved in replicative processes over the cell cycle. PLoS Pathogens 5, e1000565.CrossRefGoogle ScholarPubMed
Azizi, H, Dumas, C and Papadopoulou, B (2017) The Pumilio-domain protein PUF6 contributes to SIDER2 retroposon-mediated mRNA decay in Leishmania. RNA 23, 1874–1885.CrossRefGoogle ScholarPubMed
Bailey, T (2011) DREME: motif discovery in transcription factor ChIP-seq data. Bioinformatics (Oxford, England) 27, 1653–1659.CrossRefGoogle ScholarPubMed
Benz, C and Urbaniak, M (2019) Organising the cell cycle in the absence of transcriptional control: dynamic phosphorylation co-ordinates the cell cycle post-transcriptionally. PLoS Pathogens 15, e1008129.CrossRefGoogle ScholarPubMed
Clayton, C (2019) Control of gene expression in trypanosomatids: living with polycistronic transcription. Open Biology 9, 190072.CrossRefGoogle ScholarPubMed
Dallagiovanna, B, Perez, L, Sotelo-Silveira, J, Smircich, P, Duhagon, MA and Garat, B (2005) Trypanosoma cruzi: molecular characterization of TcPUF6, a Pumilio protein. Experimental Parasitology 109, 260–264.CrossRefGoogle ScholarPubMed
Dean, S, Marchetti, R, Kirk, K and Matthews, K (2009) A surface transporter family conveys the trypanosome differentiation signal. Nature 459, 213–217.CrossRefGoogle ScholarPubMed
Dean, S, Sunter, J and Wheeler, R (2016) Tryptag.org: a trypanosome genome-wide protein localisation resource. Trends in Parasitology 33, 80–82.CrossRefGoogle ScholarPubMed
Droll, D, Archer, S, Fenn, K, Delhi, P, Matthews, K and Clayton, C (2010) The trypanosome Pumilio-domain protein PUF7 associates with a nuclear cyclophilin and is involved in ribosomal RNA maturation. FEBS Letters 84, 1156–1162.CrossRefGoogle Scholar
Duszenko, M, Mühlstädt, K and Broder, A (1992) Cysteine is an essential growth factor for Trypanosoma brucei bloodstream forms. Molecular and Biochemical Parasitology 50, 269–274.CrossRefGoogle ScholarPubMed
Erben, E, Fadda, A, Lueong, S, Hoheisel, J and Clayton, C (2014) Genome-wide discovery of post-transcriptional regulators in Trypanosoma brucei. PLoS Pathogens 10, e1004178.CrossRefGoogle Scholar
Fadda, A, Ryten, M, Droll, D, Rojas, F, Färber, V, Haanstra, J, Bakker, B, Matthews, K and Clayton, C (2014) Transcriptome-wide analysis of mRNA decay reveals complex degradation kinetics and suggests a role for co-transcriptional degradation in determining mRNA levels. Molecular Microbiology 94, 307–326.CrossRefGoogle ScholarPubMed
Fischer, AD and Olivas, WM (2018) Multiple Puf proteins regulate the stability of ribosome biogenesis transcripts. RNA Biology 15, 1228–1243.CrossRefGoogle ScholarPubMed
Goldstrohm, AC, Hall, TMT and McKenney, KM (2018) Post-transcriptional regulatory functions of mammalian Pumilio proteins. Trends in Genetics 34, 972–990.CrossRefGoogle ScholarPubMed
Gomes-Santos, CS, Braks, J, Prudencio, M, Carret, C, Gomes, AR, Pain, A, Feltwell, T, Khan, S, Waters, A, Janse, C, Mair, GR and Mota, MM (2011) Transition of Plasmodium sporozoites into liver stage-like forms is regulated by the RNA binding protein Pumilio. PLoS Pathogens 7, e1002046.CrossRefGoogle ScholarPubMed
Hendriks, EF, Abdul-Razak, A and Matthews, KR (2003) TbCPSF30 depletion by RNA interference disrupts polycistronic RNA processing in Trypanosoma brucei. Journal of Biological Chemistry 278, 26870–26878.CrossRefGoogle ScholarPubMed
Jha, B, Archer, S and Clayton, C (2013) The trypanosome pumilio domain protein PUF5. PLoS One 8, e77371.CrossRefGoogle ScholarPubMed
Jha, B, Fadda, A, Merce, C, Mugo, E, Droll, D and Clayton, C (2014) Depletion of the trypanosome pumilio domain protein PUF2 causes transcriptome changes related to coding region length. Eukaryotic Cell 13, 664–674.CrossRefGoogle ScholarPubMed
Klein, C, Terrao, M, Inchaustegui Gil, D and Clayton, C (2015) Polysomes of Trypanosoma brucei: association with initiation factors and RNA-binding proteins. PLoS One 10, e0135973.CrossRefGoogle ScholarPubMed
Klein, C, Terrao, M and Clayton, C (2017) The role of the zinc finger protein ZC3H32 in bloodstream-form Trypanosoma brucei. PLoS One 12, e0177901.CrossRefGoogle ScholarPubMed
Kolev, NG, Ramey-Butler, K, Cross, GA, Ullu, E and Tschudi, C (2012) Developmental progression to infectivity in Trypanosoma brucei triggered by an RNA-binding protein. Science (New York, N.Y.) 338, 1352–1353.CrossRefGoogle ScholarPubMed
Liu, B, Marucha, K and Clayton, C (2020) The zinc finger proteins ZC3H20 and ZC3H21 stabilise mRNAs encoding membrane proteins and mitochondrial proteins in insect-form Trypanosoma brucei. Molecular Microbiology 113, 430–451.CrossRefGoogle ScholarPubMed
Love, M, Huber, W and Anders, S (2014) Moderated estimation of fold change and dispersion for RNA-Seq data with DESeq2. Genome Biology 15, 550.CrossRefGoogle ScholarPubMed
Lu, G and Hall, TM (2011) Alternate modes of cognate RNA recognition by human Pumilio proteins. Structure (London, England: 1993) 19, 361–367.CrossRefGoogle ScholarPubMed
Lu, G, Dolgner, S and Tanaka Hall, T (2009) Understanding and engineering RNA sequence specificity of PUF proteins. Current Opinion in Structural Biology 19, 110–115.CrossRefGoogle ScholarPubMed
Lueong, S, Merce, C, Fischer, B, Hoheisel, J and Erben, E (2016) Gene expression regulatory networks in Trypanosoma brucei: insights into the role of the mRNA-binding proteome. Molecular Microbiology 100, 457–471.CrossRefGoogle ScholarPubMed
Luu, VD, Brems, S, Hoheisel, J, Burchmore, R, Guilbride, D and Clayton, C (2006) Functional analysis of Trypanosoma brucei PUF1. Molecular and Biochemical Parasitology 150, 340–349.CrossRefGoogle ScholarPubMed
Miller, M and Olivas, W (2011) Roles of Puf proteins in mRNA degradation and translation. WIREs RNA 2, 471–492.CrossRefGoogle Scholar
Minia, I, Merce, C, Terrao, M and Clayton, C (2016) Translation regulation and RNA granule formation after heat shock of procyclic form Trypanosoma brucei: many heat-induced mRNAs are increased during differentiation to mammalian-infective forms. PLoS Neglected Tropical Diseases 10, e0004982.CrossRefGoogle ScholarPubMed
Mugo, E and Clayton, C (2017) Expression of the RNA-binding protein RBP10 promotes the bloodstream-form differentiation state in Trypanosoma brucei. PLoS Pathogens 13, e1006560.CrossRefGoogle Scholar
Mulindwa, J, Leiss, K, Ibberson, D, Kamanyi Marucha, K, Helbig, C, Melo do Nascimento, L, Silvester, E, Matthews, K, Matovu, E, Enyaru, J and Clayton, C (2018) Transcriptomes of Trypanosoma brucei Rhodesiense from sleeping sickness patients, rodents and culture: effects of strain, growth conditions and RNA preparation methods. PLoS Neglected Tropical Diseases 12, e0006280.CrossRefGoogle ScholarPubMed
Ong, HB, Sienkiewicz, N, Wyllie, S, Patterson, S and Fairlamb, AH (2013) Trypanosoma brucei UMP synthase null mutants are avirulent in mice, but recover virulence upon prolonged culture in Vitro While retaining pyrimidine auxotrophy. Molecular Microbiology 90, 443–455.Google ScholarPubMed
Quenault, T, Lithgow, T and Traven, A (2011) PUF Proteins: repression, activation and mRNA localization. Trends in Cell Biology 21, 104–112.CrossRefGoogle ScholarPubMed
Schumann Burkard, G, Kaser, S, de Araujo, PR, Schimanski, B, Naguleswaran, A, Knusel, S, Heller, M and Roditi, I (2013) Nucleolar proteins regulate stage-specific gene expression and ribosomal RNA maturation in Trypanosoma brucei. Molecular Microbiology. doi:10.1111/mmi.12227.CrossRefGoogle ScholarPubMed
Siegel, T, Hekstra, D, Wang, X, Dewell, S and Cross, G (2010) Genome-wide analysis of mRNA abundance in two life-cycle stages of Trypanosoma brucei and identification of splicing and polyadenylation sites. Nucleic Acids Research 38, 4946–4957.CrossRefGoogle ScholarPubMed
Silvester, E, Ivens, A and Matthews, KR (2018) A gene expression comparison of Trypanosoma brucei and Trypanosoma congolense in the bloodstream of the mammalian host reveals species-specific adaptations to density-dependent development. PLoS Neglected Tropical Diseases 12, e0006863.CrossRefGoogle ScholarPubMed
Szoor, B, Wilson, J, McElhinney, H, Tabernero, L and Matthews, KR (2006) Protein tyrosine phosphatase TbPTP1: a molecular switch controlling life cycle differentiation in trypanosomes. Journal of Cell Biology 175, 293–303.CrossRefGoogle ScholarPubMed
Terrao, M, Kamanyi Marucha, K, Mugo, E, Droll, D, Minia, I, Egler, F, Braun, J and Clayton, C (2018) The suppressive cap-binding-complex factor 4EIP is required for normal differentiation. Nucleic Acids Research 46, 8993–9010.CrossRefGoogle ScholarPubMed
Urbaniak, MD, Martin, D and Ferguson, MA (2013) Global quantitative SILAC phosphoproteomics reveals differential phosphorylation is widespread between the procyclic and bloodstream form lifecycle stages of Trypanosoma brucei. Journal of Proteome Research 12, 2233–2244.CrossRefGoogle ScholarPubMed
Valley, CT, Porter, DF, Qiu, C, Campbell, ZT, Hall, TM and Wickens, M (2012) Patterns and plasticity in RNA-protein interactions enable recruitment of multiple proteins through a single site. Proceedings of the National Academy of Sciences of the United States of America 109, 6054–6059.CrossRefGoogle ScholarPubMed
Vassella, E, Kramer, R, Turner, CM, Wankell, M, Modes, C, van den Bogaard, M and Boshart, M (2001) Deletion of a novel protein kinase with PX and FYVE-related domains increases the rate of differentiation of Trypanosoma brucei. Molecular Microbiology 41, 33–46.CrossRefGoogle ScholarPubMed
Wang, M, Oge, L, Perez-Garcia, MD, Hamama, L and Sakr, S (2018) The PUF protein family: overview on PUF RNA targets, biological functions, and post transcriptional regulation. International Journal of Molecular Sciences 19, 410.CrossRefGoogle ScholarPubMed
Ziegelbauer, K, Quinten, M, Schwarz, H, Pearson, TW and Overath, P (1990) Synchronous differentiation of Trypanosoma brucei bloodstream to procyclic forms in vitro. European Journal of Biochemistry 192, 373–378.CrossRefGoogle ScholarPubMed
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