scholarly journals Post-translational epigenetics: PRMT7 regulates RNA-binding capacity and protein stability to controlLeishmaniaparasite virulence

2019 ◽  
Author(s):  
Tiago R. Ferreira ◽  
Adam A. Dowle ◽  
Ewan Parry ◽  
Eliza V. C. Alves-Ferreira ◽  
Foteini Kolokousi ◽  
...  
Keyword(s):  
Protein Modification ◽  
Transcriptional Control ◽  
Leishmania Major ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Capacity ◽  
Arginine Methylation ◽  
Protein Arginine Methyltransferases

ABSTRACTRNA binding proteins (RBPs) are the primary gene regulators in kinetoplastids as transcriptional control is nearly absent, makingLeishmaniaan exceptional model for investigating methylation of non-histone substrates. Arginine methylation is an evolutionarily conserved protein modification catalyzed by Protein aRginine MethylTransferases (PRMTs). The chromatin modifier PRMT7 is the only Type III PRMT found in higher eukaryotes and a restricted number of unicellular eukaryotes. InLeishmania major, PRMT7 is a cytoplasmic protein implicit in pathogenesis with unknown substrates. Using comparative methyl-SILAC proteomics for the first time in protozoa, we identified 40 putative targets, including 17 RBPs hypomethylated upon PRMT7 knockout. PRMT7 can modify Alba3 and RBP16trans-regulators (mammalian RPP25 and YBX2 homologs, respectively) as direct substratesin vitro. The absence of PRMT7 levelsin vivoselectively reduces Alba3 mRNA-binding capacity to specific target transcripts and can impact the relative stability of RBP16 in the cytoplasm. RNA immunoprecipitation analyses demonstrate PRMT7-dependent methylation promotes Alba3 association with select target transcripts and stability ofδ-amastinsurface antigen. These results highlight a novel role for PRMT7-mediated arginine methylation upon RBP substrates, suggesting a regulatory pathway controlling gene expression and virulence inLeishmania. This work introducesLeishmaniaPRMTs as epigenetic regulators of mRNA metabolism with mechanistic insight into the functional manipulation of RBPs by methylation.

scholarly journals PRMT7 regulates RNA-binding capacity and protein stability in Leishmania parasites

2020 ◽  
Vol 48 (10) ◽  
pp. 5511-5526
Author(s):  
Tiago R Ferreira ◽  
Adam A Dowle ◽  
Ewan Parry ◽  
Eliza V C Alves-Ferreira ◽  
Karen Hogg ◽  
...  
Keyword(s):  
Protein Modification ◽  
Transcriptional Control ◽  
Leishmania Major ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Capacity ◽  
Arginine Methylation ◽  
Mrna Metabolism

Abstract RNA binding proteins (RBPs) are the primary gene regulators in kinetoplastids as transcriptional control is nearly absent, making Leishmania an exceptional model for investigating methylation of non-histone substrates. Arginine methylation is an evolutionarily conserved protein modification catalyzed by Protein aRginine Methyl Transferases (PRMTs). The chromatin modifier PRMT7 is the only Type III PRMT found in higher eukaryotes and a restricted number of unicellular eukaryotes. In Leishmania major, PRMT7 is a cytoplasmic protein implicit in pathogenesis with unknown substrates. Using comparative methyl-SILAC proteomics for the first time in protozoa, we identified 40 putative targets, including 17 RBPs hypomethylated upon PRMT7 knockout. PRMT7 can modify Alba3 and RBP16 trans-regulators (mammalian RPP25 and YBX2 homologs, respectively) as direct substrates in vitro. The absence of PRMT7 levels in vivo selectively reduces Alba3 mRNA-binding capacity to specific target transcripts and can impact the relative stability of RBP16 in the cytoplasm. RNA immunoprecipitation analyses demonstrate PRMT7-dependent methylation promotes Alba3 association with select target transcripts and thus indirectly stabilizes mRNA of a known virulence factor, δ-amastin surface antigen. These results highlight a novel role for PRMT7-mediated arginine methylation of RBP substrates, suggesting a regulatory pathway controlling gene expression and virulence in Leishmania. This work introduces Leishmania PRMTs as epigenetic regulators of mRNA metabolism with mechanistic insight into the functional manipulation of RBPs by methylation.

scholarly journals Protein methyltransferase 7 deficiency in Leishmania major increases neutrophil associated pathology in murine model

2021 ◽  
Vol 15 (3) ◽  
pp. e0009230
Author(s):  
Juliana Alcoforado Diniz ◽  
Mariana M. Chaves ◽  
Slavica Vaselek ◽  
Rubens D. Miserani Magalhães ◽  
Rafael Ricci-Azevedo ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Leishmania Major ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Capacity ◽  
Sand Fly ◽  
Parasite Development ◽  
Post Translational Modification ◽  
Phlebotomus Duboscqi ◽  
The Impact

Leishmania major is the main causative agent of cutaneous leishmaniasis in the Old World. In Leishmania parasites, the lack of transcriptional control is mostly compensated by post-transcriptional mechanisms. Methylation of arginine is a conserved post-translational modification executed by Protein Arginine Methyltransferase (PRMTs). The genome from L. major encodes five PRMT homologs, including the cytosolic protein associated with several RNA-binding proteins, LmjPRMT7. It has been previously reported that LmjPRMT7 could impact parasite infectivity. In addition, a more recent work has clearly shown the importance of LmjPRMT7 in RNA-binding capacity and protein stability of methylation targets, demonstrating the role of this enzyme as an important epigenetic regulator of mRNA metabolism. In this study, we unveil the impact of PRMT7-mediated methylation on parasite development and virulence. Our data reveals that higher levels of LmjPRMT7 can impair parasite pathogenicity, and that deletion of this enzyme rescues the pathogenic phenotype of an attenuated strain of L. major. Interestingly, lesion formation caused by LmjPRMT7 knockout parasites is associated with an exacerbated inflammatory reaction in the tissue correlated with an excessive neutrophil recruitment. Moreover, the absence of LmjPRMT7 also impairs parasite development within the sand fly vector Phlebotomus duboscqi. Finally, a transcriptome analysis shed light onto possible genes affected by depletion of this enzyme. Taken together, this study highlights how post-transcriptional regulation can affect different aspects of the parasite biology.

scholarly journals In vivo and in vitro arginine methylation of RNA-binding proteins.

1995 ◽  
Vol 15 (5) ◽  
pp. 2800-2808 ◽  
Author(s):  
Q Liu ◽  
G Dreyfuss
Keyword(s):  
Hela Cells ◽  
Posttranslational Modifications ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Arginine Methylation ◽  
Hnrnp A1 ◽  
Arginine Residues

Heterogenous nuclear ribonucleoproteins (hnRNPs) bind pre-mRNAs and facilitate their processing into mRNAs. Many of the hnRNPs undergo extensive posttranslational modifications including methylation on arginine residues. hnRNPs contain about 65% of the total NG,NG-dimethylarginine found in the cell nucleus. The role of this modification is not known. Here we identify the hnRNPs that are methylated in HeLa cells and demonstrate that most of the pre-mRNA-binding proteins receive this modification. Using recombinant human hnRNP A1 as a substrate, we have partially purified and characterized a protein-arginine N-methyltransferase specific for hnRNPs from HeLa cells. This methyltransferase can methylate the same subset of hnRNPs in vitro as are methylated in vivo. Furthermore, it can also methylate other RNA-binding proteins that contain the RGG motif RNA-binding domain. This activity is evolutionarily conserved from lower eukaryotes to mammals, suggesting that methylation has a significant role in the function of RNA-binding proteins.

scholarly journals Characterization of the Drosophila protein arginine methyltransferases DART1 and DART4

2004 ◽  
Vol 379 (2) ◽  
pp. 283-289 ◽  
Author(s):  
Marie-Chloé BOULANGER ◽  
Tina Branscombe MIRANDA ◽  
Steven CLARKE ◽  
Marco di FRUSCIO ◽  
Beat SUTER ◽  
...  
Keyword(s):  
Rna Binding ◽  
Developmental Stages ◽  
Rna Binding Proteins ◽  
Genetic System ◽  
Arginine Methylation ◽  
Type I ◽  
Protein Arginine Methyltransferases ◽  
Arginine Residues ◽  
Drosophila Protein

The role of arginine methylation in Drosophila melanogaster is unknown. We identified a family of nine PRMTs (protein arginine methyltransferases) by sequence homology with mammalian arginine methyltransferases, which we have named DART1 to DART9 (Drosophilaarginine methyltransferases 1–9). In keeping with the mammalian PRMT nomenclature, DART1, DART4, DART5 and DART7 are the putative homologues of PRMT1, PRMT4, PRMT5 and PRMT7. Other DART family members have a closer resemblance to PRMT1, but do not have identifiable homologues. All nine genes are expressed in Drosophila at various developmental stages. DART1 and DART4 have arginine methyltransferase activity towards substrates, including histones and RNA-binding proteins. Amino acid analysis of the methylated arginine residues confirmed that both DART1 and DART4 catalyse the formation of asymmetrical dimethylated arginine residues and they are type I arginine methyltransferases. The presence of PRMTs in D. melanogaster suggest that flies are a suitable genetic system to study arginine methylation.

scholarly journals Epigenetic control via allosteric regulation of mammalian protein arginine methyltransferases

2017 ◽  
Vol 114 (38) ◽  
pp. 10101-10106 ◽  
Author(s):  
Kanishk Jain ◽  
Cyrus Y. Jin ◽  
Steven G. Clarke
Keyword(s):  
Cancer Metastasis ◽  
Gene Activation ◽  
Kinetic Studies ◽  
Arginine Methylation ◽  
Histone H4 ◽  
Protein Arginine Methyltransferases ◽  
Wide Range ◽  
Substrate Concentrations

Arginine methylation on histones is a central player in epigenetics and in gene activation and repression. Protein arginine methyltransferase (PRMT) activity has been implicated in stem cell pluripotency, cancer metastasis, and tumorigenesis. The expression of one of the nine mammalian PRMTs, PRMT5, affects the levels of symmetric dimethylarginine (SDMA) at Arg-3 on histone H4, leading to the repression of genes which are related to disease progression in lymphoma and leukemia. Another PRMT, PRMT7, also affects SDMA levels at the same site despite its unique monomethylating activity and the lack of any evidence for PRMT7-catalyzed histone H4 Arg-3 methylation. We present evidence that PRMT7-mediated monomethylation of histone H4 Arg-17 regulates PRMT5 activity at Arg-3 in the same protein. We analyzed the kinetics of PRMT5 over a wide range of substrate concentrations. Significantly, we discovered that PRMT5 displays positive cooperativity in vitro, suggesting that this enzyme may be allosterically regulated in vivo as well. Most interestingly, monomethylation at Arg-17 in histone H4 not only raised the general activity of PRMT5 with this substrate, but also ameliorated the low activity of PRMT5 at low substrate concentrations. These kinetic studies suggest a biochemical explanation for the interplay between PRMT5- and PRMT7-mediated methylation of the same substrate at different residues and also suggest a general model for regulation of PRMTs. Elucidating the exact relationship between these two enzymes when they methylate two distinct sites of the same substrate may aid in developing therapeutics aimed at reducing PRMT5/7 activity in cancer and other diseases.

scholarly journals Identification of mRNAs Associated with αCP2-Containing RNP Complexes

2003 ◽  
Vol 23 (19) ◽  
pp. 7055-7067 ◽  
Author(s):  
Shelly A. Waggoner ◽  
Stephen A. Liebhaber
Keyword(s):  
Binding Proteins ◽  
Rna Binding ◽  
Cell Function ◽  
Rna Binding Proteins ◽  
Translation Control ◽  
Viral Mrnas ◽  
Posttranscriptional Gene Regulation ◽  
Direct Binding

ABSTRACT Posttranscriptional controls in higher eukaryotes are central to cell differentiation and developmental programs. These controls reflect sequence-specific interactions of mRNAs with one or more RNA binding proteins. The α-globin poly(C) binding proteins (αCPs) comprise a highly abundant subset of K homology (KH) domain RNA binding proteins and have a characteristic preference for binding single-stranded C-rich motifs. αCPs have been implicated in translation control and stabilization of multiple cellular and viral mRNAs. To explore the full contribution of αCPs to cell function, we have identified a set of mRNAs that associate in vivo with the major αCP2 isoforms. One hundred sixty mRNA species were consistently identified in three independent analyses of αCP2-RNP complexes immunopurified from a human hematopoietic cell line (K562). These mRNAs could be grouped into subsets encoding cytoskeletal components, transcription factors, proto-oncogenes, and cell signaling factors. Two mRNAs were linked to ceroid lipofuscinosis, indicating a potential role for αCP2 in this infantile neurodegenerative disease. Surprisingly, αCP2 mRNA itself was represented in αCP2-RNP complexes, suggesting autoregulatory control of αCP2 expression. In vitro analyses of representative target mRNAs confirmed direct binding of αCP2 within their 3′ untranslated regions. These data expand the list of mRNAs that associate with αCP2 in vivo and establish a foundation for modeling its role in coordinating pathways of posttranscriptional gene regulation.

scholarly journals SSMART: Sequence-structure motif identification for RNA-binding proteins

2018 ◽  
Author(s):  
Alina Munteanu ◽  
Neelanjan Mukherjee ◽  
Uwe Ohler
Keyword(s):  
Binding Sites ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Sequence Motif ◽  
Primary Sequence ◽  
Sequence Structure ◽  
Rna Targets

AbstractMotivationRNA-binding proteins (RBPs) regulate every aspect of RNA metabolism and function. There are hundreds of RBPs encoded in the eukaryotic genomes, and each recognize its RNA targets through a specific mixture of RNA sequence and structure properties. For most RBPs, however, only a primary sequence motif has been determined, while the structure of the binding sites is uncharacterized.ResultsWe developed SSMART, an RNA motif finder that simultaneously models the primary sequence and the structural properties of the RNA targets sites. The sequence-structure motifs are represented as consensus strings over a degenerate alphabet, extending the IUPAC codes for nucleotides to account for secondary structure preferences. Evaluation on synthetic data showed that SSMART is able to recover both sequence and structure motifs implanted into 3‘UTR-like sequences, for various degrees of structured/unstructured binding sites. In addition, we successfully used SSMART on high-throughput in vivo and in vitro data, showing that we not only recover the known sequence motif, but also gain insight into the structural preferences of the RBP.AvailabilitySSMART is freely available at https://ohlerlab.mdc-berlin.de/software/SSMART_137/[email protected]

scholarly journals A single domain of yeast poly(A)-binding protein is necessary and sufficient for RNA binding and cell viability.

1987 ◽  
Vol 7 (9) ◽  
pp. 3268-3276 ◽  
Author(s):  
A B Sachs ◽  
R W Davis ◽  
R D Kornberg
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Association Constant ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Binding Protein ◽  
High Affinity ◽  
Terminal Domain ◽  
Necessary And Sufficient

The poly(A)-binding protein (PAB) gene of Saccharomyces cerevisiae is essential for cell growth. A 66-amino acid polypeptide containing half of a repeated N-terminal domain can replace the entire protein in vivo. Neither an octapeptide sequence conserved among eucaryotic RNA-binding proteins nor the C-terminal domain of PAB is required for function in vivo. A single N-terminal domain is nearly identical to the entire protein in the number of high-affinity sites for poly(A) binding in vitro (one site with an association constant of approximately 2 X 10(7) M-1) and in the size of the binding site (12 A residues). Multiple N-terminal domains afford a mechanism of PAB transfer between poly(A) strands.

scholarly journals A novel methyltransferase (Hmt1p) modifies poly(A)+-RNA-binding proteins.

1996 ◽  
Vol 16 (7) ◽  
pp. 3668-3678 ◽  
Author(s):  
M F Henry ◽  
P A Silver
Keyword(s):  
Normal Cell ◽  
Binding Proteins ◽  
Rna Binding ◽  
Rna Binding Proteins ◽  
Temperature Sensitive ◽  
Arginine Residues ◽  
Cognate Gene ◽  
Heterogeneous Nuclear Ribonucleoproteins

RNA-binding proteins play many essential roles in the metabolism of nuclear pre-mRNA. As such, they demonstrate a myriad of dynamic behaviors and modifications. In particular, heterogeneous nuclear ribonucleoproteins (hnRNPs) contain the bulk of methylated arginine residues in eukaryotic cells. We have identified the first eukaryotic hnRNP-specific methyltransferase via a genetic screen for proteins that interact with an abundant poly(A)+-RNA-binding protein termed Npl3p. We have previously shown that npl3-1 mutants are temperature sensitive for growth and defective for export of mRNA from the nucleus. New mutants in interacting genes were isolated by their failure to survive in the presence of the npl3-1 allele. Four alleles of the same gene were identified in this manner. Cloning of the cognate gene revealed an encoded protein with similarity to methyltransferases that was termed HMT1 for hnRNP methyltransferase. HMT1 is not required for normal cell viability except when NPL3 is also defective. The Hmt1 protein is located in the nucleus. We demonstrate that Npl3p is methylated by Hmt1p both in vivo and in vitro. These findings now allow further exploration of the function of this previously uncharacterized class of enzymes.

scholarly journals The Emerging Role and Promise of Circular RNAs in Obesity and Related Metabolic Disorders

Cells ◽  
2020 ◽  
Vol 9 (6) ◽  
pp. 1473
Author(s):  
Mohamed Zaiou
Keyword(s):  
Rna Binding ◽  
Metabolic Diseases ◽  
Rna Binding Proteins ◽  
In Vivo Studies ◽  
Future Research ◽  
Circular Rnas ◽  
Exciting Field ◽  
Rna Molecules

Circular RNAs (circRNAs) are genome transcripts that are produced from back-splicing of specific regions of pre-mRNA. These single-stranded RNA molecules are widely expressed across diverse phyla and many of them are stable and evolutionary conserved between species. Growing evidence suggests that many circRNAs function as master regulators of gene expression by influencing both transcription and translation processes. Mechanistically, circRNAs are predicted to act as endogenous microRNA (miRNA) sponges, interact with functional RNA-binding proteins (RBPs), and associate with elements of the transcriptional machinery in the nucleus. Evidence is mounting that dysregulation of circRNAs is closely related to the occurrence of a range of diseases including cancer and metabolic diseases. Indeed, there are several reports implicating circRNAs in cardiovascular diseases (CVD), diabetes, hypertension, and atherosclerosis. However, there is very little research addressing the potential role of these RNA transcripts in the occurrence and development of obesity. Emerging data from in vitro and in vivo studies suggest that circRNAs are novel players in adipogenesis, white adipose browning, obesity, obesity-induced inflammation, and insulin resistance. This study explores the current state of knowledge on circRNAs regulating molecular processes associated with adipogenesis and obesity, highlights some of the challenges encountered while studying circRNAs and suggests some perspectives for future research directions in this exciting field of study.

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