Conserved RNA Binding Activity of Phosphatidyl Inositol 5-Phosphate 4-Kinase (PIP4K2A)

Plasmodium falciparum is a causative agent for malaria and has a complex life cycle in human and mosquito hosts. During its life cycle, the malarial parasite Plasmodium goes through different asexual and sexual stages, in humans and mosquitoes. Expression of stage-specific proteins is important for successful completion of its life cycle and requires tight gene regulation. In the case of Plasmodium, due to relative paucity of the transcription factors, it is postulated that posttranscriptional regulation plays an important role in stage-specific gene expression. Translation repression of specific set of mRNA has been reported in gametocyte stages of the parasite. A conserved element present in the 3′UTR of some of these transcripts was identified. Phosphatidylinositol 5-phosphate 4-kinase (PIP4K2A) was identified as the protein that associates with these RNA. We now show that the RNA binding activity of PIP4K2A is independent of its kinase activity. We also observe that PIP4K2A is imported into the parasite from the host on Plasmodium berghei and Toxoplasma gondii. The RNA binding activity of PIP4K2A seems to be conserved across species from Drosophila and C. elegans to humans, suggesting that the RNA binding activity of PIP4K may be important, and there may be host transcripts that may be regulated by PIP4K2A. These results identify a novel RNA binding role for PIP4K2A that may not only play a role in Plasmodium propagation but may also function in regulating gene expression in multicellular organisms.

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Crystal structure of a Y-box binding protein 1 (YB-1)–RNA complex reveals key features and residues interacting with RNA

Journal of Biological Chemistry ◽

10.1074/jbc.ra119.007545 ◽

2019 ◽

Vol 294 (28) ◽

pp. 10998-11010 ◽

Cited By ~ 15

Author(s):

Xiao-Juan Yang ◽

Hong Zhu ◽

Shi-Rong Mu ◽

Wen-Juan Wei ◽

Xun Yuan ◽

...

Keyword(s):

Gene Expression ◽

Posttranscriptional Regulation ◽

Rna Binding ◽

Binding Protein ◽

Consensus Sequence ◽

Regulation Of Gene Expression ◽

Binding Activity ◽

Core Motif ◽

Variable Exon ◽

Nucleotide Resolution

The Y-box binding protein 1 (YB-1) is a member of the cold shock domain (CSD) protein family and is recognized as an oncogenic factor in several solid tumors. By binding to RNA, YB-1 participates in several steps of posttranscriptional regulation of gene expression, including mRNA splicing, stability, and translation; microRNA processing; and stress granule assembly. However, the mechanisms in YB-1–mediated regulation of RNAs are unclear. Previously, we used both systematic evolution of ligands by exponential enrichment (SELEX) and individual-nucleotide resolution UV cross-linking and immunoprecipitation coupled RNA-Seq (iCLIP-Seq) analyses, which defined the RNA-binding consensus sequence of YB-1 as CA(U/C)C. We also reported that through binding to its core motif CAUC in primary transcripts, YB-1 regulates the alternative splicing of a CD44 variable exon and the biogenesis of miR-29b-2 during both Drosha and Dicer steps. To elucidate the molecular basis of the YB-1–RNA interactions, we report high-resolution crystal structures of the YB-1 CSD in complex with different RNA oligos at 1.7 Å resolution. The structure revealed that CSD interacts with RNA mainly through π–π stacking interactions assembled by four highly conserved aromatic residues. Interestingly, YB-1 CSD forms a homodimer in solution, and we observed that two residues, Tyr-99 and Asp-105, at the dimer interface are important for YB-1 CSD dimerization. Substituting these two residues with Ala reduced CSD's RNA-binding activity and abrogated the splicing activation of YB-1 targets. The YB-1 CSD–RNA structures presented here at atomic resolution provide mechanistic insights into gene expression regulated by CSD-containing proteins.

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A Trypanosoma cruzi zinc finger protein that is implicated in the control of epimastigote-specific gene expression and metacyclogenesis

Parasitology ◽

10.1017/s0031182020002176 ◽

2020 ◽

pp. 1-15

Author(s):

Thais S. Tavares ◽

Fernanda L. B. Mügge ◽

Viviane Grazielle-Silva ◽

Bruna M. Valente ◽

Wanessa M. Goes ◽

...

Keyword(s):

Gene Expression ◽

Life Cycle ◽

Trypanosoma Cruzi ◽

Zinc Finger ◽

Rna Binding ◽

Environmental Changes ◽

Negative Regulator ◽

Specific Gene ◽

Genes Encoding ◽

Transcriptome Analyses

Abstract Trypanosoma cruzi has three biochemically and morphologically distinct developmental stages that are programmed to rapidly respond to environmental changes the parasite faces during its life cycle. Unlike other eukaryotes, Trypanosomatid genomes contain protein coding genes that are transcribed into polycistronic pre-mRNAs and have their expression controlled by post-transcriptional mechanisms. Transcriptome analyses comparing three stages of the T. cruzi life cycle revealed changes in gene expression that reflect the parasite adaptation to distinct environments. Several genes encoding RNA binding proteins (RBPs), known to act as key post-transcriptional regulatory factors, were also differentially expressed. We characterized one T. cruzi RBP, named TcZH3H12, which contains a zinc finger domain and is up-regulated in epimastigotes compared to trypomastigotes and amastigotes. TcZC3H12 knockout (KO) epimastigotes showed decreased growth rates and increased capacity to differentiate into metacyclic trypomastigotes. Transcriptome analyses comparing wild type and TcZC3H12 KOs revealed a TcZC3H12-dependent expression of epimastigote-specific genes such as genes encoding amino acid transporters and proteins associated with differentiation (PADs). RNA immunoprecipitation assays showed that transcripts from the PAD family interact with TcZC3H12. Taken together, these findings suggest that TcZC3H12 positively regulates the expression of genes involved in epimastigote proliferation and also acts as a negative regulator of metacyclogenesis.

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Gene expression profiles of dicyemid life-cycle stages may explain how dispersing larvae locate new hosts

Zoological Letters ◽

10.1186/s40851-019-0146-y ◽

2019 ◽

Vol 5 (1) ◽

Author(s):

Tsai-Ming Lu ◽

Hidetaka Furuya ◽

Noriyuki Satoh

Keyword(s):

Gene Expression ◽

Life Cycle ◽

Expression Profiles ◽

Gene Expression Profiles ◽

Molecular Transport ◽

Complex Life Cycle ◽

Sensory Response ◽

Specific Gene ◽

Life Cycle Stages ◽

New Hosts

Abstract Metazoans have evolved a great variety of life histories in response to environmental conditions. A unique example is encountered in dicyemid mesozoans. In addition to a highly simplified adult body comprising only ~ 30 cells, dicyemids exhibit a parasitic lifestyle that includes nematogens (asexual reproductive adults), rhombogens (sexual reproductive adults), vermiform larvae generated by nematogens, and infusoriform larvae generated by rhombogens. However, due to the difficulties of observing microscopic endoparasites, the complex life cycle and biological functions of life-cycle stages of dicyemids have remained mysterious. Taking advantage of the recently decoded genome of Dicyema japonicum, we examined genes that undergird this lifestyle. Using stage-specific gene expression profiles, we found that biological processes associated with molecular transport, developmental regulation, and sensory response are specified at different stages. Together with the expression of potential neurotransmitters, we further suggest that apical cells in infusoriform larva probably serve sensory functions, although dicyemids have no nervous system. Gene expression profiles show that more genes are expressed in free-living infusoriform larvae than in the other three stages, and that some of these genes are likely involved in locating new hosts. These data provide molecular information about the unique lifestyle of dicyemids and illustrate how an extremely simplified endoparasite adapted and retained gene sets and morphological characters to complete its life cycle.

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The ApiAP2 factor PfAP2-HC is an integral component of heterochromatin in the malaria parasite Plasmodium falciparum

10.1101/2020.11.06.370338 ◽

2020 ◽

Author(s):

Eilidh Carrington ◽

Roel H. M. Cooijmans ◽

Dominique Keller ◽

Christa G. Toenhake ◽

Richárd Bártfai ◽

...

Keyword(s):

Gene Expression ◽

Plasmodium Falciparum ◽

Life Cycle ◽

Dna Binding ◽

Regulation Of Gene Expression ◽

Complex Life Cycle ◽

Parasite Development ◽

Specific Gene ◽

Mosquito Vector ◽

Malaria Parasites

AbstractMalaria parasites undergo a highly complex life cycle in the human host and the mosquito vector. The ApiAP2 family of sequence-specific DNA-binding proteins plays a dominant role in parasite development and life cycle progression. Of the ApiAP2 factors studied to date, most act as transcription factors regulating stage-specific gene expression. Here, we characterised a new ApiAP2 factor in Plasmodium falciparum (PF3D7_1456000) that we termed PfAP2-HC. Via detailed investigation of several single or double genetically engineered parasite lines, we demonstrate that PfAP2-HC specifically binds to heterochromatin throughout the genome. Intriguingly, PfAP2-HC does not bind DNA in vivo and recruitment of PfAP2-HC to heterochromatin is independent of its DNA-binding domain but strictly dependent on heterochromatin protein 1. Furthermore, our results suggest that PfAP2-HC functions neither in the regulation of gene expression nor in heterochromatin formation or maintenance. In summary, our findings reveal that PfAP2-HC constitutes a core component of heterochromatin in malaria parasites. They furthermore identify unexpected properties of ApiAP2 factors and suggest substantial functional divergence among the members of this important family of regulatory proteins.

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STAU2 protein level is controlled by caspases and the CHK1 pathway and regulates cell cycle progression in the non-transformed hTERT-RPE1 cells

BMC Molecular and Cell Biology ◽

10.1186/s12860-021-00352-y ◽

2021 ◽

Vol 22 (1) ◽

Author(s):

Lionel Condé ◽

Yulemi Gonzalez Quesada ◽

Florence Bonnet-Magnaval ◽

Rémy Beaujois ◽

Luc DesGroseillers

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Cell Proliferation ◽

Dna Damage ◽

Steady State ◽

Posttranscriptional Regulation ◽

Cell Cycle Progression ◽

Rna Binding ◽

Regulation Of Gene Expression ◽

Cycle Progression

AbstractBackgroundStaufen2 (STAU2) is an RNA binding protein involved in the posttranscriptional regulation of gene expression. In neurons, STAU2 is required to maintain the balance between differentiation and proliferation of neural stem cells through asymmetric cell division. However, the importance of controlling STAU2 expression for cell cycle progression is not clear in non-neuronal dividing cells. We recently showed that STAU2 transcription is inhibited in response to DNA-damage due to E2F1 displacement from theSTAU2gene promoter. We now study the regulation of STAU2 steady-state levels in unstressed cells and its consequence for cell proliferation.ResultsCRISPR/Cas9-mediated and RNAi-dependent STAU2 depletion in the non-transformed hTERT-RPE1 cells both facilitate cell proliferation suggesting that STAU2 expression influences pathway(s) linked to cell cycle controls. Such effects are not observed in the CRISPR STAU2-KO cancer HCT116 cells nor in the STAU2-RNAi-depleted HeLa cells. Interestingly, a physiological decrease in the steady-state level of STAU2 is controlled by caspases. This effect of peptidases is counterbalanced by the activity of the CHK1 pathway suggesting that STAU2 partial degradation/stabilization fines tune cell cycle progression in unstressed cells. A large-scale proteomic analysis using STAU2/biotinylase fusion protein identifies known STAU2 interactors involved in RNA translation, localization, splicing, or decay confirming the role of STAU2 in the posttranscriptional regulation of gene expression. In addition, several proteins found in the nucleolus, including proteins of the ribosome biogenesis pathway and of the DNA damage response, are found in close proximity to STAU2. Strikingly, many of these proteins are linked to the kinase CHK1 pathway, reinforcing the link between STAU2 functions and the CHK1 pathway. Indeed, inhibition of the CHK1 pathway for 4 h dissociates STAU2 from proteins involved in translation and RNA metabolism.ConclusionsThese results indicate that STAU2 is involved in pathway(s) that control(s) cell proliferation, likely via mechanisms of posttranscriptional regulation, ribonucleoprotein complex assembly, genome integrity and/or checkpoint controls. The mechanism by which STAU2 regulates cell growth likely involves caspases and the kinase CHK1 pathway.

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Quantitative proteomics revealed C6orf203/MTRES1 as a factor preventing stress-induced transcription deficiency in human mitochondria

Nucleic Acids Research ◽

10.1093/nar/gkz542 ◽

2019 ◽

Vol 47 (14) ◽

pp. 7502-7517 ◽

Cited By ~ 6

Author(s):

Anna V Kotrys ◽

Dominik Cysewski ◽

Sylwia D Czarnomska ◽

Zbigniew Pietras ◽

Lukasz S Borowski ◽

...

Keyword(s):

Gene Expression ◽

Rna Binding ◽

Mitochondrial Gene ◽

Binding Activity ◽

Stress Conditions ◽

Mitochondrial Rna ◽

Mitochondrial Gene Expression ◽

Compensatory Mechanisms ◽

Temporary Reduction ◽

Mitochondrial Transcription

AbstractMaintenance of mitochondrial gene expression is crucial for cellular homeostasis. Stress conditions may lead to a temporary reduction of mitochondrial genome copy number, raising the risk of insufficient expression of mitochondrial encoded genes. Little is known how compensatory mechanisms operate to maintain proper mitochondrial transcripts levels upon disturbed transcription and which proteins are involved in them. Here we performed a quantitative proteomic screen to search for proteins that sustain expression of mtDNA under stress conditions. Analysis of stress-induced changes of the human mitochondrial proteome led to the identification of several proteins with poorly defined functions among which we focused on C6orf203, which we named MTRES1 (Mitochondrial Transcription Rescue Factor 1). We found that the level of MTRES1 is elevated in cells under stress and we show that this upregulation of MTRES1 prevents mitochondrial transcript loss under perturbed mitochondrial gene expression. This protective effect depends on the RNA binding activity of MTRES1. Functional analysis revealed that MTRES1 associates with mitochondrial RNA polymerase POLRMT and acts by increasing mitochondrial transcription, without changing the stability of mitochondrial RNAs. We propose that MTRES1 is an example of a protein that protects the cell from mitochondrial RNA loss during stress.

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What functional genomics has taught us about transcriptional regulation in malaria parasites

Briefings in Functional Genomics ◽

10.1093/bfgp/elz004 ◽

2019 ◽

Vol 18 (5) ◽

pp. 290-301 ◽

Cited By ~ 3

Author(s):

Christa G Toenhake ◽

Richárd Bártfai

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Life Cycle ◽

Functional Genomics ◽

Expression Patterns ◽

Regulatory Elements ◽

Complex Life Cycle ◽

Malaria Parasites ◽

Gene Regulatory

Abstract Malaria parasites are characterized by a complex life cycle that is accompanied by dynamic gene expression patterns. The factors and mechanisms that regulate gene expression in these parasites have been searched for even before the advent of next generation sequencing technologies. Functional genomics approaches have substantially boosted this area of research and have yielded significant insights into the interplay between epigenetic, transcriptional and post-transcriptional mechanisms. Recently, considerable progress has been made in identifying sequence-specific transcription factors and DNA-encoded regulatory elements. Here, we review the insights obtained from these efforts including the characterization of core promoters, the involvement of sequence-specific transcription factors in life cycle progression and the mapping of gene regulatory elements. Furthermore, we discuss recent developments in the field of functional genomics and how they might contribute to further characterization of this complex gene regulatory network.

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Fitting Pieces into the Puzzle ofPseudomonas aeruginosaType III Secretion System Gene Expression

Journal of Bacteriology ◽

10.1128/jb.00209-19 ◽

2019 ◽

Vol 201 (13) ◽

Cited By ~ 8

Author(s):

Emily A. Williams McMackin ◽

Louise Djapgne ◽

Jodi M. Corley ◽

Timothy L. Yahr

Keyword(s):

Gene Expression ◽

Protein Delivery ◽

Rna Binding ◽

Rna Binding Proteins ◽

Binding Activity ◽

Secretion Systems ◽

Content Type ◽

Global Regulators ◽

Dna Binding Activity ◽

Host Pathogen

ABSTRACTType III secretion systems (T3SS) are widely distributed in Gram-negative microorganisms and critical for host-pathogen and host-symbiont interactions with plants and animals. Central features of the T3SS are a highly conserved set of secretion and translocation genes and contact dependence wherein host-pathogen interactions trigger effector protein delivery and serve as an inducing signal for T3SS gene expression. In addition to these conserved features, there are pathogen-specific properties that include a unique repertoire of effector genes and mechanisms to control T3SS gene expression. ThePseudomonas aeruginosaT3SS serves as a model system to understand transcriptional and posttranscriptional mechanisms involved in the control of T3SS gene expression. The central regulatory feature is a partner-switching system that controls the DNA-binding activity of ExsA, the primary regulator of T3SS gene expression. Superimposed upon the partner-switching mechanism are cyclic AMP and cyclic di-GMP signaling systems, two-component systems, global regulators, and RNA-binding proteins that have positive and negative effects on ExsA transcription and/or synthesis. In the present review, we discuss advances in our understanding of how these regulatory systems orchestrate the activation of T3SS gene expression in the context of acute infections and repression of the T3SS asP. aeruginosaadapts to and colonizes the cystic fibrosis airways.

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