570 Biowissenschaften; Biologie
Refine
Year of publication
Document Type
- Preprint (201) (remove)
Language
- English (201) (remove)
Has Fulltext
- yes (201)
Is part of the Bibliography
- no (201)
Keywords
- E2 enzyme (6)
- TRACT (6)
- oligomerization (6)
- ubiquitination (6)
- ISGylation (4)
- autophagy (3)
- HCV (2)
- ISGlyation (2)
- NS3 (2)
- NS5A (2)
Institute
- Biowissenschaften (54)
- Medizin (40)
- Buchmann Institut für Molekulare Lebenswissenschaften (BMLS) (33)
- Biochemie, Chemie und Pharmazie (32)
- Biochemie und Chemie (27)
- Physik (27)
- MPI für Biophysik (20)
- Senckenbergische Naturforschende Gesellschaft (17)
- Institut für Ökologie, Evolution und Diversität (14)
- Frankfurt Institute for Advanced Studies (FIAS) (12)
Owing to their morphological complexity and dense network connections, neurons modify their proteomes locally, using mRNAs and ribosomes present in the neuropil (tissue enriched for dendrites and axons). Although ribosome biogenesis largely takes place in the nucleus and perinuclear region, neuronal ribosomal protein (RP) mRNAs have been frequently detected remotely, in dendrites and axons. Here, using imaging and ribosome profiling, we directly detected the RP mRNAs and their translation in the neuropil. Combining brief metabolic labeling with mass spectrometry, we found that a group of RPs quickly associated with translating ribosomes in the cytoplasm and that this incorporation is independent of canonical ribosome biogenesis. Moreover, the incorporation probability of some RPs was regulated by location (neurites vs. cell bodies) and changes in the cellular environment (in response to oxidative stress). Our results suggest new mechanisms for the local activation, repair and/or specialization of the translational machinery within neuronal processes, potentially allowing remote neuronal synapses a rapid solution to the relatively slow and energy-demanding requirement of nuclear ribosome biogenesis.
Mitochondrial matrix peptidase CLPP is crucial during cell stress. Its loss causes Perrault syndrome type 3 (PRLTS3) with infertility, neurodegeneration and growth deficit. Its target proteins are disaggregated by CLPX, which also regulates heme biosynthesis via unfolding ALAS enzyme, providing access of pyridoxal-5’-phosphate (PLP). Despite efforts in diverse organisms with multiple techniques, CLPXP substrates remain controversial. Here, avoiding recombinant overexpression, we employed complexomics in mitochondria from three mouse tissues to identify endogenous targets. CLPP absence caused accumulation and dispersion of CLPX-VWA8 as AAA+ unfoldases, and of PLPBP. Similar changes and CLPX-VWA8 comigration were evident for mitoribosomal central protuberance clusters, translation factors like GFM1-HARS2, RNA granule components LRPPRC-SLIRP, and enzymes OAT-ALDH18A1. Mitochondrially translated proteins in testis showed reductions to <30% for MTCO1-3, misassembly of complex-IV supercomplex, and accumulated metal-binding assembly factors COX15-SFXN4. Indeed, heavy metal levels were increased for iron, molybdenum, cobalt and manganese. RT-qPCR showed compensatory downregulation only for Clpx mRNA, most accumulated proteins appeared transcriptionally upregulated. Immunoblots validated VWA8, MRPL38, MRPL18, GFM1 and OAT accumulation. Coimmunoprecipitation confirmed CLPX binding to MRPL38, GFM1 and OAT, so excess CLPX and PLP may affect their activity. Our data elucidate mechanistically the mitochondrial translation fidelity deficits, which underlie progressive hearing impairment in PRLTS3.
Viruses that carry a positive-sense, single-stranded (+ssRNA) RNA translate their genomes soon after entering the host cell to produce viral proteins, with the exception of retroviruses. A distinguishing feature of retroviruses is reverse transcription, where the +ssRNA genome serves as a template to synthesize a double-stranded DNA copy that subsequently integrates into the host genome. As retroviral RNAs are produced by the host cell transcriptional machinery and are largely indistinguishable from cellular mRNAs, we investigated the potential of incoming retroviral genomes to directly express proteins. Here we show through multiple, complementary methods that retroviral genomes are translated after entry. Our findings challenge the notion that retroviruses require reverse transcription to produce viral proteins. Synthesis of retroviral proteins in the absence of productive infection has significant implications for basic retrovirology, immune responses and gene therapy applications.
Viruses that carry a positive-sense, single-stranded RNA translate their genomes after entering the host cell to produce viral proteins, with the exception of retroviruses. A distinguishing feature of retroviruses is reverse transcription, where the ssRNA genome serves as a template to synthesize a double-stranded DNA copy that subsequently integrates into the host genome. As retroviral RNAs are produced by the host transcriptional machinery and are largely indistinguishable from cellular mRNAs, we investigated the potential of incoming retroviral genomes to express proteins. Here we show through various biochemical methods that HIV-1 genomes are translated after entry, in case of minimal or full-length genomes, envelopes using different cellular entry pathways and in diverse cell types. Our findings challenge the dogma that retroviruses require reverse transcription to produce viral proteins. Synthesis of retroviral proteins in the absence of productive infection has significant implications for basic retrovirology, immune responses and gene therapy applications.
In Arabidopsis thaliana, the stem cell niche (SCN) within the root apical meristem (RAM) is maintained by an intricate regulatory network that ensures optimal growth and high developmental plasticity. Yet, many aspects of this regulatory network of stem cell quiescence and replenishment are still not fully understood. Here, we investigate the interplay of the key transcription factors (TFs) BRASSINOSTEROID AT VASCULAR AND ORGANIZING CENTRE (BRAVO), PLETHORA 3 (PLT3) and WUSCHEL-RELATED HOMEOBOX 5 (WOX5) involved in SCN maintenance. Phenotypical analysis of mutants involving these TFs uncover their combinatorial regulation of cell fates and divisions in the SCN. Moreover, interaction studies employing fluorescence resonance energy transfer fluorescence lifetime imaging microscopy (FRET-FLIM) in combination with novel analysis methods, allowed us to quantify protein-protein interaction (PPI) affinities as well as higher-order complex formation of these TFs. We integrated our experimental results into a computational model, suggesting that cell type specific profiles of protein complexes and characteristic complex formation, that is also dependent on prion-like domains in PLT3, contribute to the intricate regulation of the SCN. We propose that these unique protein complex ‘signatures’ could serve as a read-out for cell specificity thereby adding another layer to the sophisticated regulatory network that balances stem cell maintenance and replenishment in the Arabidopsis root.
Understanding the complexity of transcriptional regulation is a major goal of computational biology. Because experimental linkage of regulatory sites to genes is challenging, computational methods considering epigenomics data have been proposed to create tissue-specific regulatory maps. However, we showed that these approaches are not well suited to account for the variations of the regulatory landscape between cell-types. To overcome these drawbacks, we developed a new method called STITCHIT, that identifies and links putative regulatory sites to genes. Within STITCHIT, we consider the chromatin accessibility signal of all samples jointly to identify regions exhibiting a signal variation related to the expression of a distinct gene. STITCHIT outperforms previous approaches in various validation experiments and was used with a genome-wide CRISPR-Cas9 screen to prioritize novel doxorubicin-resistance genes and their associated non-coding regulatory regions. We believe that our work paves the way for a more refined understanding of transcriptional regulation at the gene-level.
MicroRNAs (miRNAs) are critical post-transcriptional regulators in many biological processes. They act by guiding RNA-induced silencing complexes to miRNA response elements (MREs) in target mRNAs, inducing translational inhibition and/or mRNA degradation. Functional MREs are expected to predominantly occur in the 3' untranslated region and involve perfect base-pairing of the miRNA seed. Here, we generate a high-resolution map of miR-181a/b-1 (miR-181) MREs to define the targeting rules of miR-181 in developing murine T-cells. By combining a multi-omics approach with computational high-resolution analyses, we uncover novel miR-181 targets and demonstrate that miR-181 acts predominantly through RNA destabilization. Importantly, we discover an alternative seed match and identify a distinct set of targets with repeat elements in the coding sequence which are targeted by miR-181 and mediate translational inhibition. In conclusion, deep profiling of MREs in primary cells is critical to expand physiologically relevant targetomes and establish context-dependent miRNA targeting rules.
In this special issue of Matrix Biology centered on proteoglycan biology we have assembled a blend of articles focused on the state-of-the-art of proteoglycanology. The field has greatly expanded in the past three decades and now encompasses all the areas of biology. This special issue is divided into five chapters describing hyaluronan metabolism, biosynthetic and catabolic pathways of proteoglycans and their roles in inflammation, cancer, repair and development. We hope that the new original work and the reviews from recognized leaders will stimulate investigations in this exciting and fertile field of research.
The epitranscriptome embodies many new and largely unexplored functions of RNA. A major roadblock in the epitranscriptomics field is the lack of transcriptome-wide methods to detect more than a single RNA modification type at a time, identify RNA modifications in individual molecules, and estimate modification stoichiometry accurately. We address these issues with CHEUI (CH3 (methylation) Estimation Using Ionic current), a new method that concurrently detects N6-methyladenosine (m6A) and 5-methylcytidine (m5C) in individual RNA molecules from the same sample, as well as differential methylation between any two conditions. CHEUI processes observed and expected nanopore direct RNA sequencing signals with convolutional neural networks to achieve high single-molecule accuracy and outperforms other methods in detecting m6A and m5C sites and quantifying their stoichiometry. CHEUI’s unique capability to identify two modification types in the same sample reveals a non-random co-occurrence of m6A and m5C in mRNA transcripts in cell lines and tissues. CHEUI unlocks an unprecedented potential to study RNA modification configurations and discover new epitranscriptome functions.
The epitranscriptome embodies many new and largely unexplored functions of RNA. A major roadblock in the epitranscriptomics field is the lack of transcriptome-wide methods to detect more than a single RNA modification type at a time, identify RNA modifications in individual molecules, and estimate modification stoichiometry accurately. We address these issues with CHEUI (CH3 (methylation) Estimation Using Ionic current), a new method that concurrently detects N6-methyladenosine (m6A) and 5-methylcytidine (m5C) in individual RNA molecules from the same sample, as well as differential methylation between any two conditions. CHEUI processes observed and expected nanopore direct RNA sequencing signals with convolutional neural networks to achieve high single-molecule accuracy and outperforms other methods in detecting m6A and m5C sites and quantifying their stoichiometry. CHEUI’s unique capability to identify two modification types in the same sample reveals a non-random co-occurrence of m6A and m5C in mRNA transcripts in cell lines and tissues. CHEUI unlocks an unprecedented potential to study RNA modification configurations and discover new epitranscriptome functions.