scholarly journals Stop codon readthrough alters the activity of a POU/Oct transcription factor during Drosophila development

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Yunpo Zhao ◽  
Bo Gustav Lindberg ◽  
Shiva Seyedoleslami Esfahani ◽  
Xiongzhuo Tang ◽  
Stefano Piazza ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Larval Development ◽  
Stop Codon ◽  
Steroid Hormone ◽  
Protein Isoforms ◽  
Prothoracic Gland ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Translational Readthrough ◽  
Stop Codon Readthrough

Abstract Background A number of cellular processes have evolved in metazoans that increase the proteome repertoire in relation to the genome, such as alternative splicing and translation recoding. Another such process, translational stop codon readthrough (SCR), generates C-terminally extended protein isoforms in many eukaryotes, including yeast, plants, insects, and humans. While comparative genome analyses have predicted the existence of programmed SCR in many species including humans, experimental proof of its functional consequences are scarce. Results We show that SCR of the Drosophila POU/Oct transcription factor Ventral veins lacking/Drifter (Vvl/Dfr) mRNA is prevalent in certain tissues in vivo, reaching a rate of 50% in the larval prothoracic gland. Phylogenetically, the C-terminal extension is conserved and harbors intrinsically disordered regions and amino acid stretches implied in transcriptional activation. Elimination of Vvl/Dfr translational readthrough by CRISPR/Cas9 mutagenesis changed the expression of a large number of downstream genes involved in processes such as chromatin regulation, neurogenesis, development, and immune response. As a proof-of-principle, we demonstrate that the C-terminal extension of Vvl/Dfr is necessary for correct timing of pupariation, by increasing the capacity to regulate its target genes. The extended Vvl/Dfr isoform acts in synergy with the transcription factor Molting defective (Mld) to increase the expression and biosynthesis of the steroid hormone ecdysone, thereby advancing pupariation. Consequently, late-stage larval development was prolonged and metamorphosis delayed in vvl/dfr readthrough mutants. Conclusions We demonstrate that translational recoding of a POU/Oct transcription factor takes place in a highly tissue-specific and temporally controlled manner. This dynamic and regulated recoding is necessary for normal expression of a large number of genes involved in many cellular and developmental processes. Loss of Vvl/Dfr translational readthrough negatively affects steroid hormone biosynthesis and delays larval development and progression into metamorphosis. Thus, this study demonstrates how SCR of a transcription factor can act as a developmental switch in a spatiotemporal manner, feeding into the timing of developmental transitions between different life-cycle stages. Graphical abstract

scholarly journals Stop codon readthrough of a POU transcription factor regulates steroidogenesis and developmental transitions

2020 ◽  
Author(s):  
Yunpo Zhao ◽  
Bo Gustav Lindberg ◽  
Shiva Seyedoleslami Esfahani ◽  
Xiongzhuo Tang ◽  
Stefano Piazza ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Transcriptional Activation ◽  
Stop Codon ◽  
Phylogenetic Analyses ◽  
Protein Isoforms ◽  
High Rate ◽  
Prothoracic Gland ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Stop Codon Readthrough

AbstractTranslational stop codon readthrough generates C-terminally extended protein isoforms. While evidence mounts of readthrough as a global phenomenon, proofs of its functional consequences are scarce. We show that readthrough of the mRNA for the Drosophila POU/Oct transcription factor Drifter occurs at a high rate and in a spatiotemporal manner in vivo, reaching above 50% in the prothoracic gland. Phylogenetic analyses suggested that readthrough of drifter is conserved among Dipterans, with C-terminal extensions harboring intrinsically disordered regions, and amino acids streches implied in transcriptional activation. The C-terminally extended Drifter isoform is required for maintaining normal levels of the growth hormone ecdysone through regulation of its biosynthetic genes, acting in synergy with the transcription factor Molting defective. A 14-bp deletion that abolished readthrough, caused prolonged larval development and delayed metamorphosis. This study provides a striking example of alternative genetic decoding that feeds into the progression from one life cycle stage to another.

scholarly journals Identification of mRNAs that undergo stop codon readthrough in Arabidopsis thaliana

2021 ◽  
Author(s):  
Sarthak Sahoo ◽  
Divyoj Singh ◽  
Anumeha Singh ◽  
Sandeep M. Eswarappa
Keyword(s):  
Arabidopsis Thaliana ◽  
Stop Codon ◽  
Protein Localization ◽  
Abiotic Stress Tolerance ◽  
Ribosome Profiling ◽  
Protein Isoforms ◽  
Functional Enrichment ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Stop Codon Readthrough

A stop codon ensures termination of translation at a specific position on an mRNA. Sometimes, termination fails as translation machinery recognizes a stop codon as a sense codon. This leads to stop codon readthrough (SCR) resulting in the continuation of translation beyond the stop codon, generating protein isoforms with C-terminal extension. SCR has been observed in viruses, fungi, and multicellular organisms including mammals. However, SCR is largely unexplored in plants. In this study, we have analyzed ribosome profiling datasets to identify mRNAs that undergo SCR in Arabidopsis thaliana. Analyses of the ribosome density, ribosome coverage and three-nucleotide periodicity of the ribosome profiling reads, in the mRNA region downstream of the stop codon, provided strong evidence for SCR in mRNAs of 144 genes. This process generates putative peroxisomal targeting signal, nuclear localization signal, prenylation signal, transmembrane helix and intrinsically disordered regions in the C-terminal extension of several of these proteins. Gene ontology (GO) functional enrichment analysis revealed that these 144 genes belong to three major functional groups - translation, photosynthesis and abiotic stress tolerance. Finally, using a luminescence-based assay, we experimentally demonstrate SCR in representative mRNAs belonging to these functional classes. Based on these observations, we propose that SCR plays an important role in plant physiology by regulating the protein localization and function.

scholarly journals The functional readthrough extension of malate dehydrogenase reveals a modification of the genetic code

Open Biology ◽  
2016 ◽  
Vol 6 (11) ◽  
pp. 160246 ◽  
Author(s):  
Julia Hofhuis ◽  
Fabian Schueren ◽  
Christopher Nötzel ◽  
Thomas Lingner ◽  
Jutta Gärtner ◽  
...  
Keyword(s):  
Malate Dehydrogenase ◽  
Stop Codon ◽  
Protein Isoforms ◽  
Translational Readthrough ◽  
Peroxisomal Targeting ◽  
Codon Context ◽  
Targeting Signal ◽  
Stop Codon Readthrough ◽  
Insight Into ◽  
Actual Ratio

Translational readthrough gives rise to C-terminally extended proteins, thereby providing the cell with new protein isoforms. These may have different properties from the parental proteins if the extensions contain functional domains. While for most genes amino acid incorporation at the stop codon is far lower than 0.1%, about 4% of malate dehydrogenase (MDH1) is physiologically extended by translational readthrough and the actual ratio of MDH1x (e x tended protein) to ‘normal' MDH1 is dependent on the cell type. In human cells, arginine and tryptophan are co-encoded by the MDH1x UGA stop codon. Readthrough is controlled by the 7-nucleotide high-readthrough stop codon context without contribution of the subsequent 50 nucleotides encoding the extension. All vertebrate MDH1x is directed to peroxisomes via a hidden peroxisomal targeting signal (PTS) in the readthrough extension, which is more highly conserved than the extension of lactate dehydrogenase B. The hidden PTS of non-mammalian MDH1x evolved to be more efficient than the PTS of mammalian MDH1x. These results provide insight into the genetic and functional co-evolution of these dually localized dehydrogenases.

scholarly journals Robustness by intrinsically disordered C-termini and translational readthrough

2018 ◽  
Vol 46 (19) ◽  
pp. 10184-10194 ◽  
Author(s):  
April Snofrid Kleppe ◽  
Erich Bornberg-Bauer
Keyword(s):  
High Frequency ◽  
Ribosomal Subunit ◽  
Conformational Flexibility ◽  
Ribosome Profiling ◽  
Protein Isoforms ◽  
Evolutionary Constraints ◽  
Protein Chain ◽  
Intrinsically Disordered ◽  
Translational Readthrough ◽  
Evolutionary Paths

Abstract During protein synthesis genetic instructions are passed from DNA via mRNA to the ribosome to assemble a protein chain. Occasionally, stop codons in the mRNA are bypassed and translation continues into the untranslated region (3′-UTR). This process, called translational readthrough (TR), yields a protein chain that becomes longer than would be predicted from the DNA sequence alone. Protein sequences vary in propensity for translational errors, which may yield evolutionary constraints by limiting evolutionary paths. Here we investigated TR in Saccharomyces cerevisiae by analysing ribosome profiling data. We clustered proteins as either prone or non-prone to TR, and conducted comparative analyses. We find that a relatively high frequency (5%) of genes undergo TR, including ribosomal subunit proteins. Our main finding is that proteins undergoing TR are highly expressed and have a higher proportion of intrinsically disordered C-termini. We suggest that highly expressed proteins may compensate for the deleterious effects of TR by having intrinsically disordered C-termini, which may provide conformational flexibility but without distorting native function. Moreover, we discuss whether minimizing deleterious effects of TR is also enabling exploration of the phenotypic landscape of protein isoforms.

scholarly journals Mapping the Promiscuous Binding Interface of HOX-A11 with KIX by Experimentally Guided in-silico docking

2021 ◽  
Author(s):  
Soumya Ganguly ◽  
Günter P. Wagner ◽  
Jens Meiler
Keyword(s):  
Transcription Factor ◽  
Protein Interactions ◽  
Binding Protein ◽  
Creb Binding Protein ◽  
Computational Docking ◽  
In Silico Docking ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Binding Interface ◽  
Kix Domain

AbstractTranscription factors (TFs) regulate levels of transcription through a complex array of protein-protein interactions, thereby controlling key physiological processes such as development, stress response and cell growth. The transcription factor HOXA11 contains an intrinsically disordered regions (IDR) through which it interacts with CREB binding protein (CBP) and regulates endometrial development and function in eutherian mammals. The interaction between the IDR of HOXA11 and CBP was analyzed using computational docking guided by experimental constraints. HOXA11 IDR interacts with the KIX domain of CBP at two discrete sites – MLL and cMyb, mediated by sticky hydrophobic grooves on the surface of KIX. A five residue motif FDQFF on HOXA11 can interact both at cMyb and MLL site of KIX resulting in a promiscuous binding.Author SummaryWe demonstrate how the intrinsically disordered region (IDR) of transcription factor HOXA11 interacts at two distinct sites of the transcription coactivator CREB binding protein (CBP). By combining computational docking with limited experimental data we construct models of the complex of the KIX domain within CBP and a short helical segment within the IDR of HOXA11. The interaction between HOXA11 and CBP is believed to trigger the downstream expression of genes important in embryonic development.

scholarly journals Oxidative Stress-Induced Misfolding and Inclusion Formation of Nrf2 and Keap1

2021 ◽  
Author(s):  
Vy Ngo ◽  
Nadun C. Karunatilleke ◽  
Anne Brickenden ◽  
Wing-Yiu Choy ◽  
Martin L. Duennwald
Keyword(s):  
Oxidative Stress ◽  
Transcription Factor ◽  
Protein Folding ◽  
Protein Misfolding ◽  
Mammalian Cells ◽  
Inclusion Formation ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Antioxidant Proteins ◽  
Cysteine Content

Cells that experience high levels of oxidative stress respond with the induction of antioxidant proteins through the activation of the transcription factor Nrf2. Nrf2 is negatively regulated by Keap1 which binds to Nrf2 to facilitate its ubiquitination and ensuing proteasomal degradation under basal conditions. Here, we study protein folding and misfolding in Nrf2 and Keap1 in yeast, mammalian cells, and purified proteins under oxidative stress conditions. Both Nrf2 and Keap1 are susceptible to protein misfolding and inclusion formation upon oxidative stress. We propose that the intrinsically disordered regions within Nrf2 and the high cysteine content of Keap1 contribute to their oxidation and the ensuing misfolding. Our work reveals previously unexplored aspects of Nrf2 and Keap1 regulation and dysregulation by oxidation-induced protein misfolding.

scholarly journals Glucocorticoid Receptor Condensates Link DNA-Dependent Receptor Dimerization and Transcriptional Transactivation

2021 ◽  
Vol 5 (Supplement_1) ◽  
pp. A809-A809
Author(s):  
Filipp Frank ◽  
Xu Liu ◽  
Eric Ortlund
Keyword(s):  
Transcription Factor ◽  
Dna Binding ◽  
Glucocorticoid Receptor ◽  
Target Genes ◽  
Dna Recognition ◽  
Receptor Dimerization ◽  
Intrinsically Disordered ◽  
Intrinsically Disordered Regions ◽  
Recognition Elements

Abstract The glucocorticoid receptor (GR) is a ligand-regulated transcription factor (TF) that controls the tissue- and gene-specific transactivation and transrepression of thousands of target genes. Distinct GR DNA binding sequences with activating or repressive activities have been identified, but how they modulate transcription in opposite ways is not known. We show that GR forms phase-separated condensates that specifically concentrate known co-regulators via their intrinsically disordered regions (IDRs) in vitro. A combination of dynamic, multivalent (between IDRs) and specific, stable interactions (between LxxLL motifs and the GR ligand binding domain) control the degree of recruitment. Importantly, GR DNA-binding directs the selective partitioning of co-regulators within GR condensates such that activating DNAs cause enhanced recruitment of co-activators. Our work shows that condensation controls GR function by modulating co-regulator recruitment and provides a mechanism for the up- and down-regulation of GR target genes controlled by distinct DNA recognition elements.

Sign in / Sign up

Export Citation Format

Share Document