scholarly journals Identification of a motif in Trm732 required for 2′-O-methylation of the tRNA anticodon loop by Trm7

2021 ◽  
Author(s):  
Holly M. Funk ◽  
Daisy J. DiVita ◽  
Hannah E. Sizemore ◽  
Kendal Wehrle ◽  
Catherine L. Weiner ◽  
...  
Keyword(s):  
Yeast Cells ◽  
Trna Modification ◽  
Anticodon Loop ◽  
Secondary Effects ◽  
Cellular Functions ◽  
Trna Modifications ◽  
Modification Activity ◽  
Human Disorders ◽  
Methylation Activity

Posttranscriptional tRNA modifications are essential for proper gene expression, and defects in the enzymes that perform tRNA modifications are associated with numerous human disorders. Throughout eukaryotes, 2′-O-methylation of residues 32 and 34 of the anticodon loop of tRNA is important for proper translation, and in humans, lack of these modifications results in non-syndromic X-linked intellectual disability. In yeast, the methyltransferase Trm7 forms a complex with Trm732 to 2′-O-methylate tRNA residue 32 and with Trm734 to 2′-O-methylate tRNA residue 34. Trm732 and Trm734 are required for the methylation activity of Trm7, but the role of these auxiliary proteins is not clear. Additionally, Trm732 and Trm734 homologs are implicated in biological processes not directly related to translation, suggesting that these proteins may have additional cellular functions. To identify critical amino acids in Trm732, we generated variants and tested their ability to function in yeast cells. We identified a conserved RRSAGLP motif in the conserved DUF2428 domain of Trm732 that is required for tRNA modification activity by both yeast Trm732 and its human homolog THADA. The identification of Trm732 variants that lack tRNA modification activity will help to determine if other biological functions ascribed to Trm732 and THADA are directly due to tRNA modification, or to secondary effects due to other functions of these proteins.

scholarly journals Roles of Elongator Dependent tRNA Modification Pathways in Neurodegeneration and Cancer

Genes ◽  
2018 ◽  
Vol 10 (1) ◽  
pp. 19 ◽  
Author(s):  
Harmen Hawer ◽  
Alexander Hammermeister ◽  
Keerthiraju Ravichandran ◽  
Sebastian Glatt ◽  
Raffael Schaffrath ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Disease ◽  
Messenger Rna ◽  
Environmental Changes ◽  
Mrna Translation ◽  
Regulatory Control ◽  
Trna Modification ◽  
Positive Role ◽  
Trna Modifications

Transfer RNA (tRNA) is subject to a multitude of posttranscriptional modifications which can profoundly impact its functionality as the essential adaptor molecule in messenger RNA (mRNA) translation. Therefore, dynamic regulation of tRNA modification in response to environmental changes can tune the efficiency of gene expression in concert with the emerging epitranscriptomic mRNA regulators. Several of the tRNA modifications are required to prevent human diseases and are particularly important for proper development and generation of neurons. In addition to the positive role of different tRNA modifications in prevention of neurodegeneration, certain cancer types upregulate tRNA modification genes to sustain cancer cell gene expression and metastasis. Multiple associations of defects in genes encoding subunits of the tRNA modifier complex Elongator with human disease highlight the importance of proper anticodon wobble uridine modifications (xm5U34) for health. Elongator functionality requires communication with accessory proteins and dynamic phosphorylation, providing regulatory control of its function. Here, we summarized recent insights into molecular functions of the complex and the role of Elongator dependent tRNA modification in human disease.

scholarly journals Reciprocal regulation of TORC signaling and tRNA modifications by Elongator enforces nutrient-dependent cell fate

2019 ◽  
Vol 5 (6) ◽  
pp. eaav0184 ◽  
Author(s):  
Julie Candiracci ◽  
Valerie Migeot ◽  
Yok-Hian Chionh ◽  
Fanelie Bauer ◽  
Thomas Brochier ◽  
...  
Keyword(s):  
Cell Differentiation ◽  
Cell Growth ◽  
Cell Fate ◽  
Glycogen Synthase ◽  
Yeast Cells ◽  
Glycogen Synthase Kinase 3 ◽  
Trna Modification ◽  
Reciprocal Regulation ◽  
Trna Modifications ◽  
Dependent Cell

Nutrient availability has a profound impact on cell fate. Upon nitrogen starvation, wild-type fission yeast cells uncouple cell growth from cell division to generate small, round-shaped cells that are competent for sexual differentiation. The TORC1 (TOR complex 1) and TORC2 complexes exert opposite controls on cell growth and cell differentiation, but little is known about how their activity is coordinated. We show that transfer RNA (tRNA) modifications by Elongator are critical for this regulation by promoting the translation of both key components of TORC2 and repressors of TORC1. We further identified the TORC2 pathway as an activator of Elongator by down-regulating a Gsk3 (glycogen synthase kinase 3)–dependent inhibitory phosphorylation of Elongator. Therefore, a feedback control is operating between TOR complex (TORC) signaling and tRNA modification by Elongator to enforce the advancement of mitosis that precedes cell differentiation.

scholarly journals Misactivation of multiple starvation responses in yeast by loss of tRNA modifications

2020 ◽  
Author(s):  
Alexander Bruch ◽  
Teresa Laguna ◽  
Falk Butter ◽  
Raffael Schaffrath ◽  
Roland Klassen
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Mrna Translation ◽  
Exponential Growth Phase ◽  
Trna Modification ◽  
Anticodon Loop ◽  
Transcriptional Responses ◽  
Trna Modifications ◽  
Trna Species ◽  
Transcriptional Stress ◽  
Cellular Phenotypes

Abstract Previously, combined loss of different anticodon loop modifications was shown to impair the function of distinct tRNAs in Saccharomyces cerevisiae. Surprisingly, each scenario resulted in shared cellular phenotypes, the basis of which is unclear. Since loss of tRNA modification may evoke transcriptional responses, we characterized global transcription patterns of modification mutants with defects in either tRNAGlnUUG or tRNALysUUU function. We observe that the mutants share inappropriate induction of multiple starvation responses in exponential growth phase, including derepression of glucose and nitrogen catabolite-repressed genes. In addition, autophagy is prematurely and inadequately activated in the mutants. We further demonstrate that improper induction of individual starvation genes as well as the propensity of the tRNA modification mutants to form protein aggregates are diminished upon overexpression of tRNAGlnUUG or tRNALysUUU, the tRNA species that lack the modifications of interest. Hence, our data suggest that global alterations in mRNA translation and proteostasis account for the transcriptional stress signatures that are commonly triggered by loss of anticodon modifications in different tRNAs.

scholarly journals Identification of the enzymes responsible for m2,2G and acp3U formation on cytosolic tRNA from insects and plants

PLoS ONE ◽  
2020 ◽  
Vol 15 (11) ◽  
pp. e0242737
Author(s):  
Holly M. Funk ◽  
Ruoxia Zhao ◽  
Maggie Thomas ◽  
Sarah M. Spigelmyer ◽  
Nichlas J. Sebree ◽  
...  
Keyword(s):  
Arabidopsis Thaliana ◽  
Drosophila Melanogaster ◽  
Protein Translation ◽  
Yeast Cells ◽  
Trna Modification ◽  
Trna Modifications ◽  
Tryptophan Residues ◽  
Exogenous Genes ◽  
Multicellular Organisms ◽  
Aspartate Residue

Posttranscriptional modification of tRNA is critical for efficient protein translation and proper cell growth, and defects in tRNA modifications are often associated with human disease. Although most of the enzymes required for eukaryotic tRNA modifications are known, many of these enzymes have not been identified and characterized in several model multicellular eukaryotes. Here, we present two related approaches to identify the genes required for tRNA modifications in multicellular organisms using primer extension assays with fluorescent oligonucleotides. To demonstrate the utility of these approaches we first use expression of exogenous genes in yeast to experimentally identify two TRM1 orthologs capable of forming N2,N2-dimethylguanosine (m2,2G) on residue 26 of cytosolic tRNA in the model plant Arabidopsis thaliana. We also show that a predicted catalytic aspartate residue is required for function in each of the proteins. We next use RNA interference in cultured Drosophila melanogaster cells to identify the gene required for m2,2G26 formation on cytosolic tRNA. Additionally, using these approaches we experimentally identify D. melanogaster gene CG10050 as the corresponding ortholog of human DTWD2, which encodes the protein required for formation of 3-amino-3-propylcarboxyuridine (acp3U) on residue 20a of cytosolic tRNA. We further show that A. thaliana gene AT2G41750 can form acp3U20b on an A. thaliana tRNA expressed in yeast cells, and that the aspartate and tryptophan residues in the DXTW motif of this protein are required for modification activity. These results demonstrate that these approaches can be used to study tRNA modification enzymes.

The Role of microRNAs in the Viral Infections

2019 ◽  
Vol 24 (39) ◽  
pp. 4659-4667 ◽  
Author(s):  
Mona Fani ◽  
Milad Zandi ◽  
Majid Rezayi ◽  
Nastaran Khodadad ◽  
Hadis Langari ◽  
...  
Keyword(s):  
Viral Infections ◽  
Rna Viruses ◽  
Regulation Of Gene Expression ◽  
Molecular Structures ◽  
Main Function ◽  
Cellular Functions ◽  
Viral Genes ◽  
Non Coding Rnas ◽  
Post Transcriptional Regulation

MicroRNAs (miRNAs) are non-coding RNAs with 19 to 24 nucleotides which are evolutionally conserved. MicroRNAs play a regulatory role in many cellular functions such as immune mechanisms, apoptosis, and tumorigenesis. The main function of miRNAs is the post-transcriptional regulation of gene expression via mRNA degradation or inhibition of translation. In fact, many of them act as an oncogene or tumor suppressor. These molecular structures participate in many physiological and pathological processes of the cell. The virus can also produce them for developing its pathogenic processes. It was initially thought that viruses without nuclear replication cycle such as Poxviridae and RNA viruses can not code miRNA, but recently, it has been proven that RNA viruses can also produce miRNA. The aim of this articles is to describe viral miRNAs biogenesis and their effects on cellular and viral genes.

Establishing a model to demonstrate physical and mathematical properties of chromatin fibres in fission yeast cells - Research in the Molecular Biophysics Lab at Hiroshima University

Impact ◽  
2018 ◽  
Vol 2018 (3) ◽  
pp. 89-91
Author(s):  
Shin-ichi Tate
Keyword(s):  
Molecular Biology ◽  
Yeast Cells ◽  
Molecular Architecture ◽  
Ministry Of Education ◽  
Cellular Functions ◽  
Sports Science ◽  
Cellular Events ◽  
And Function ◽  
Education Culture ◽  
Open The Doors

The field of molecular biology has provided great insights into the structure and function of key molecules. Thanks to this area of research, we can now grasp the biological details of DNA and have characterised an enormous number of molecules in massive data bases. These 'biological periodic tables' have allowed scientists to connect molecules to particular cellular events, furthering scientific understanding of biological processes. However, molecular biology has yet to answer questions regarding 'higher-order' molecular architecture, such as that of chromatin. Chromatin is the molecular material that serves as the building block for chromosomes, the structures that carry an organism's genetic information inside of the cell's nucleus. Understanding the physical properties of chromatin is crucial in developing a more thorough picture of how chromatin's structure relate to its key cellular functions. Moreover, by establishing a physical model of chromatin, scientists will be able to open the doors into the true inner workings of the cell nucleus. Professor Shin-ichi Tate and his team of researchers at Hiroshima University's Research Center for the Mathematics on Chromatin Live Dynamics (RcMcD), are attempting to do just that. Through a five-year grant funded by the Platform for Dynamic Approaches to Living Systems from the Ministry of Education, Culture, Sports, Science and Technology, Tate is aiming to gain a clearer understanding of the structure and dynamics of chromatin.

scholarly journals Carcinoma-associated fibroblasts derived exosomes modulate breast cancer cell stemness through exonic circHIF1A by miR-580-5p in hypoxic stress

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Yanxia Zhan ◽  
Junxian Du ◽  
Zhihui Min ◽  
Li Ma ◽  
Wei Zhang ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Stem Cell ◽  
Breast Cancer Cells ◽  
Hypoxic Stress ◽  
Cellular Functions ◽  
Breast Cancer Therapy ◽  
Array Analysis ◽  
Cancer Stroma ◽  
Carcinoma Associated Fibroblasts

AbstractHypoxia is a common phenomenon in solid tumors. The roles of exosomes from hypoxic breast cancer stroma are less studied. So, the study was aimed to investigate the role of exosomes from hypoxic cancer-associated fibroblasts (CAFs) cells in breast cancer. The circRNA array analysis was performed to screen differential expressed circRNAs between hypoxic and normoxic CAFs exosomes. Candidate circHIF1A (circ_0032138) was screened out and it was confirmed that circHIF1A was up-regulated in the exosomes from hypoxic CAFs and their exosomes. Through investigating cellular functions including cell proliferation and stem cell features, it was demonstrated that hypoxic CAFs exosomes transferred circHIF1A into breast cancer cells, which played an important role in cancer stem cell properties sponging miR-580-5p by regulating CD44 expression. In a summary, circHIF1A from hypoxic CAFs exosomes played an important role in stem cell properties of breast cancer. CircHIF1A may act as a target molecule of breast cancer therapy.

RNA secondary structure dependence in METTL3–METTL14 mRNA methylation is modulated by the N-terminal domain of METTL3

2020 ◽  
Vol 402 (1) ◽  
pp. 89-98
Author(s):  
Nathalie Meiser ◽  
Nicole Mench ◽  
Martin Hengesbach
Keyword(s):  
Secondary Structure ◽  
Rna Binding ◽  
Specific Protein ◽  
Structure Dependence ◽  
Terminal Domain ◽  
Methylation Assay ◽  
A Site ◽  
Methylation Activity

AbstractN6-methyladenosine (m6A) is the most abundant modification in mRNA. The core of the human N6-methyltransferase complex (MTC) is formed by a heterodimer consisting of METTL3 and METTL14, which specifically catalyzes m6A formation within an RRACH sequence context. Using recombinant proteins in a site-specific methylation assay that allows determination of quantitative methylation yields, our results show that this complex methylates its target RNAs not only sequence but also secondary structure dependent. Furthermore, we demonstrate the role of specific protein domains on both RNA binding and substrate turnover, focusing on postulated RNA binding elements. Our results show that one zinc finger motif within the complex is sufficient to bind RNA, however, both zinc fingers are required for methylation activity. We show that the N-terminal domain of METTL3 alters the secondary structure dependence of methylation yields. Our results demonstrate that a cooperative effect of all RNA-binding elements in the METTL3–METTL14 complex is required for efficient catalysis, and that binding of further proteins affecting the NTD of METTL3 may regulate substrate specificity.

scholarly journals Loss of Ftsj1 perturbs codon-specific translation efficiency in the brain and is associated with X-linked intellectual disability

2021 ◽  
Vol 7 (13) ◽  
pp. eabf3072
Author(s):  
Y. Nagayoshi ◽  
T. Chujo ◽  
S. Hirata ◽  
H. Nakatsuka ◽  
C.-W. Chen ◽  
...  
Keyword(s):  
Intellectual Disability ◽  
State Level ◽  
Memory Deficits ◽  
Ribosome Profiling ◽  
Translation Efficiency ◽  
Trna Modification ◽  
Synaptic Organization ◽  
Transfer Rnas ◽  
The Brain

FtsJ RNA 2′-O-methyltransferase 1 (FTSJ1) gene has been implicated in X-linked intellectual disability (XLID), but the molecular pathogenesis is unknown. We show that Ftsj1 is responsible for 2′-O-methylation of 11 species of cytosolic transfer RNAs (tRNAs) at the anticodon region, and these modifications are abolished in Ftsj1 knockout (KO) mice and XLID patient–derived cells. Loss of 2′-O-methylation in Ftsj1 KO mouse selectively reduced the steady-state level of tRNAPhe in the brain, resulting in a slow decoding at Phe codons. Ribosome profiling showed that translation efficiency is significantly reduced in a subset of genes that need to be efficiently translated to support synaptic organization and functions. Ftsj1 KO mice display immature synaptic morphology and aberrant synaptic plasticity, which are associated with anxiety-like and memory deficits. The data illuminate a fundamental role of tRNA modification in the brain through regulation of translation efficiency and provide mechanistic insights into FTSJ1-related XLID.

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