Global analysis of posttranslational protein arginylation.
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ABSTRACT:
SUBMITTER: Wong CC
PROVIDER: S-EPMC1988855 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Arginyltransferase 1 (Ate1) mediates protein arginylation, a poorly understood protein posttranslational modification (PTM) in eukaryotic cells. Previous evidence suggest a potential involvement of arginylation in stress response and this PTM was traditionally considered anti-apoptotic based on the studies of individual substrates. However, here we found that arginylation promotes cell death and/or growth arrest, depending on the nature and intensity of the stressing factor. Specifically, in yeast, mouse and human cells, deletion or downregulation of the ATE1 gene disrupts typical stress responses by bypassing growth arrest and suppressing cell death events in the presence of disease-related stressing factors, including oxidative, heat, and osmotic stresses, as well as the exposure to heavy metals or radiation. Conversely, in wild-type cells responding to stress, there is an increase of cellular Ate1 protein level and arginylation activity. Furthermore, the increase of Ate1 protein directly promotes cell death in a manner dependent on its arginylation activity. Finally, we found Ate1 to be required to suppress mutation frequency in yeast and mammalian cells during DNA-damaging conditions such as ultraviolet irradiation. Our study clarifies the role of Ate1/arginylation in stress response and provides a new mechanism to explain the link between Ate1 and a variety of diseases including cancer. This is also the first example that the modulation of the global level of a PTM is capable of affecting DNA mutagenesis.
| S-EPMC5059882 | biostudies-other
Project description:The highly conserved enzyme arginyl-tRNA-protein transferase (Ate1) mediates arginylation, a posttranslational modification that is only incompletely understood at its molecular level. To investigate whether arginylation affects actin-dependent processes in a simple model organism, Dictyostelium discoideum, we knocked out the gene encoding Ate1 and characterized the phenotype of ate1-null cells. Visualization of actin cytoskeleton dynamics by live-cell microscopy indicated significant changes in comparison to wild-type cells. Ate1-null cells were almost completely lacking focal actin adhesion sites at the substrate-attached surface and were only weakly adhesive. In two-dimensional chemotaxis assays toward folate or cAMP, the motility of ate1-null cells was increased. However, in three-dimensional chemotaxis involving more confined conditions, the motility of ate1-null cells was significantly reduced. Live-cell imaging showed that GFP-tagged Ate1 rapidly relocates to sites of newly formed actin-rich protrusions. By mass spectrometric analysis, we identified four arginylation sites in the most abundant actin isoform of Dictyostelium, in addition to arginylation sites in other actin isoforms and several actin-binding proteins. In vitro polymerization assays with actin purified from ate1-null cells revealed a diminished polymerization capacity in comparison to wild-type actin. Our data indicate that arginylation plays a crucial role in the regulation of cytoskeletal activities.
| S-EPMC6594445 | biostudies-literature
Project description:Arginyltransferase1 (ATE1) is a conserved enzyme in eukaryotes mediating posttranslational arginylation, the addition of an extra arginine to an existing protein. In mammals, the dysregulations of the ATE1 gene (ate1) is shown to be involved in cardiovascular abnormalities, cancer, and aging-related diseases. Although biochemical evidence suggested that arginylation may be involved in stress response and/or protein degradation, the physiological role of ATE1 in vivo has never been systematically determined. This gap of knowledge leads to difficulties for interpreting the involvements of ATE1 in diseases pathogenesis. Since ate1 is highly conserved between human and the unicellular organism Schizosaccharomyces pombe (S. pombe), we take advantage of the gene-knockout library of S. pombe, to investigate the genetic interactions between ate1 and other genes in a systematic and unbiased manner. By this approach, we found that ate1 has a surprisingly small and focused impact size. Among the 3659 tested genes, which covers nearly 75% of the genome of S. pombe, less than 5% of them displayed significant genetic interactions with ate1. Furthermore, these ate1-interacting partners can be grouped into a few discrete clustered categories based on their functions or their physical interactions. These categories include translation/transcription regulation, biosynthesis/metabolism of biomolecules (including histidine), cell morphology and cellular dynamics, response to oxidative or metabolic stress, ribosomal structure and function, and mitochondrial function. Unexpectedly, inconsistent to popular belief, very few genes in the global ubiquitination or degradation pathways showed interactions with ate1. Our results suggested that ATE1 specifically regulates a handful of cellular processes in vivo, which will provide critical mechanistic leads for studying the involvements of ATE1 in normal physiologies as well as in diseased conditions.
| S-EPMC7218141 | biostudies-literature
Project description:Posttranslational addition of Arg to proteins, mediated by arginyltransferase ATE1 has been first observed in 1963 and remained poorly understood for decades since its original discovery. Recent work demonstrated the global nature of arginylation and its essential role in multiple physiological pathways during embryogenesis and adulthood and identified over a hundred of proteins arginylated in vivo. Among these proteins, the prominent role belongs to the actin cytoskeleton and the muscle, and follow up studies strongly suggests that arginylation constitutes a novel biological regulator of contractility. This review presents an overview of the studies of protein arginylation that led to the discovery of its major role in the muscle.
| S-EPMC4135399 | biostudies-literature
Project description:We describe an integrated workflow for proteogenomic analysis and global profiling of posttranslational modifications (PTMs) in prokaryotes and use the model cyanobacterium Synechococcus sp. PCC 7002 (hereafter Synechococcus 7002) as a test case. We found more than 20 different kinds of PTMs, and a holistic view of PTM events in this organism grown under different conditions was obtained without specific enrichment strategies. Among 3,186 predicted protein-coding genes, 2,938 gene products (>92%) were identified. We also identified 118 previously unidentified proteins and corrected 38 predicted gene-coding regions in the Synechococcus 7002 genome. This systematic analysis not only provides comprehensive information on protein profiles and the diversity of PTMs in Synechococcus 7002 but also provides some insights into photosynthetic pathways in cyanobacteria. The entire proteogenomics pipeline is applicable to any sequenced prokaryotic organism, and we suggest that it should become a standard part of genome annotation projects.
| S-EPMC4284577 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below.
2020-08-12 | GSE146344 | GEO
Project description:Arginyltransferase ATE1 mediates posttranslational arginylation that plays key roles in mammalian embryogenesis, cell migration, and normal brain function. The molecular mechanisms of arginylation remain elusive. ATE1 utilizes arginyl-tRNAArg as the donor of Arg, putting this reaction into a direct competition with the protein synthesis machinery. Here, we addressed these questions of ATE1- arginyl-tRNAArg specificity as a potential mechanism enabling this competition in vivo. Using in vitro arginylation assays and ATE1 knockout models, we find that while arginylation is specific to tRNAArg, it is able to utilize short tRNAArg derivatives that bear structural resemblance to tRNA-derived fragments (tRF), a new class of small regulatory non-coding RNAs with poorly characterized but critical functions in vivo. Arginyl-tRFArg can be generated in vitro directly from pre-charged -tRNAArg, and ATE1 is able to utilize these arginyl-tRFArg fragments with similar efficiency as arginyl-tRNAArg. Lack of arginylation in ATE1 knockout cells leads to a decrease in tRFArg generation and a significant increase in the ratio of tRNAArg to tRFArg compared to wild type, suggesting a functional link between tRFArg and arginylation in vivo. We propose that generation of physiologically important tRFs can play a critical role as a switch between protein translation and arginylation in vivo.
2020-08-12 | GSE146343 | GEO
Project description:Arginyltransferase ATE1 mediates posttranslational arginylation that plays key roles in mammalian embryogenesis, cell migration, and normal brain function. The molecular mechanisms of arginylation remain elusive. ATE1 utilizes arginyl-tRNAArg as the donor of Arg, putting this reaction into a direct competition with the protein synthesis machinery. Here, we addressed these questions of ATE1- arginyl-tRNAArg specificity as a potential mechanism enabling this competition in vivo. Using in vitro arginylation assays and ATE1 knockout models, we find that while arginylation is specific to tRNAArg, it is able to utilize short tRNAArg derivatives that bear structural resemblance to tRNA-derived fragments (tRF), a new class of small regulatory non-coding RNAs with poorly characterized but critical functions in vivo. Arginyl-tRFArg can be generated in vitro directly from pre-charged -tRNAArg, and ATE1 is able to utilize these arginyl-tRFArg fragments with similar efficiency as arginyl-tRNAArg. Lack of arginylation in ATE1 knockout cells leads to a decrease in tRFArg generation and a significant increase in the ratio of tRNAArg to tRFArg compared to wild type, suggesting a functional link between tRFArg and arginylation in vivo. We propose that generation of physiologically important tRFs can play a critical role as a switch between protein translation and arginylation in vivo.
2020-08-12 | GSE146342 | GEO
Project description:Arginylation is a post-translational modification mediated by the arginyltransferase ATE1, which transfers the amino acid arginine to a protein or peptide substrate from a tRNA molecule. Initially, arginylation was thought to occur only on N-terminally exposed acidic residues, and its function was thought to be limited to targeting proteins for degradation. However, more recent data has shown that ATE1 can arginylate sidechains of internal acidic residues in a protein without necessarily affecting metabolic stability. This greatly expands the potential targets and functions of arginylation, but tools for studying this process have remained limited. Here, we report the first global screen specifically for sidechain arginylation. We generate and validate pan-arginylation antibodies, which are designed to detect sidechain arginylation in any amino acid sequence context. We use these antibodies for immunoaffinity enrichment of sidechain arginylated proteins from wildtype and Ate1 knockout cell lysates. In this way, we identify a limited set of proteins that likely undergo ATE1-dependent sidechain arginylation and that are enriched in specific cellular roles, including translation, splicing, and the cytoskeleton.
2023-03-01 | MSV000091388 | MassIVE
Project description:Arginylation is a post-translational modification mediated by the arginyltransferase (Ate1). We recently showed that conditional deletion of Ate1 in the nervous system leads to increased light-evoked response sensitivities of ON-bipolar cells in the retina, indicating that arginylation regulates the G-protein signaling complexes of those neurons and/or photoreceptors. However, none of the key players in the signaling pathway were previously shown to be arginylated. Here we show that Gαt1, Gβ1, RGS6, and RGS7 are arginylated in the retina and RGS6 and RGS7 protein levels are elevated in Ate1 knockout, suggesting that arginylation plays a direct role in regulating their protein level and the G-protein-mediated responses in the retina.
| S-EPMC8815403 | biostudies-literature