Biology subjects

Schluter, H.

Publications and source records attributed to Schluter, H..

6 recordsLinked to original sources

bioRxiv2026

The Legionella phosphocholinase AnkX modulates IMPDH2 regulation and cytoophidia dynamics

Legionella pneumophila, the causative agent of Legionnaires disease, secretes more than 300 different effector proteins into the host cell to modulate processes such as signal transduction, membrane dynamics, and metabolism. A key regulator of cellular metabolism and proliferation is inosine 5-monophosphate dehydrogenase type II (IMPDH2), which catalyzes the rate-limiting step of de novo GTP biosynthesis. IMPDH2 assembles into filaments and larger structures known as cytoophidia, which enable fine-tuned regulation of enzymatic activity in response to changing nucleotide demands. Here, we identify human IMPDH2 as a previously unrecognized host target of the Legionella effector AnkX. We show that AnkX post-translationally modifies IMPDH2 within its regulatory domain by covalently attaching a phosphocholine moiety. While this post-translational modification does not alter the catalytic activity of IMPDH2, it disrupts filament formation, thereby impairing nucleotide-dependent regulation of enzyme activity. Consequently, AnkX-mediated phosphocholination affects IMPDH2 filament assembly, cytoophidia formation, and subcellular localization to the specific Legionella-containing vacuole (LCV). Thus, L. pneumophila subverts host GTP metabolism by posttranslationally modifying a central enzyme of the GTP biosynthetic pathway. Graphical abstract description O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=170 SRC="FIGDIR/small/701698v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@18f7724org.highwire.dtl.DTLVardef@1d70e86org.highwire.dtl.DTLVardef@1fa8890org.highwire.dtl.DTLVardef@11bc5ba_HPS_FORMAT_FIGEXP M_FIG C_FIG The Legionella effector AnkX post-translationally modifies the human host enzyme IMPDH2 by transferring a phosphocholine group using CDP-choline as co-substrate. We identified the modification site and discovered that this post-translational modification disrupts the ability of IMPDH2 to assemble into filaments and cytoophidia. By shifting the equilibrium toward free octamers, AnkX-mediated PCylation perturbs the structural regulation of IMPDH2 and alters GTP metabolism in L. pneumophila-infected host cells. This figure was inspired by (https://www.biorxiv.org/content/10.1101/2024.07.29.605679v2).

microbiology↗

bioRxiv2023

Proteomic adaptations in the kidney reveal orchestration of local and secreted antimicrobial peptides in human pyelonephritis

Pyelonephritis (PN) is a frequent bacterial infection of the kidney and is often associated with severe diseases, organ loss and sepsis. Antibiotics are the cornerstone of therapy, however, increasing antibiotic resistance threatens therapy success and necessitates novel treatment strategies. Various proteins, such as antimicrobial peptides (AMPs), are key molecules of the innate immune response and insights into their regulation may help overcome multi-drug resistance and severe diseases. Using label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS), several cellular, biological, and metabolic processes important for the antimicrobial response were identified, including a significant increase in previously undescribed proteins in human PN with antimicrobial function. Among others, we observed elevation of AMPs, such as calprotectin, azurocidin-1, and cathepsin G in the kidney, which we validated in the urine. Additionally, we observed a negative correlation of azurocidin-1 with plasma levels of C-reactive protein suggesting that the presence in the kidney may protect from severe diseases and systemic inflammation. This study represents the first renal proteomic dataset of human PN, enabling novel insights into the expression of AMPs in the context of PN. Lay SummaryGrowing antimicrobial resistance necessitates a better understanding of the expression of proteins that are critical for the immune response. Using mass spectrometry we identified AMPs in the kidney and urine of PN patients. Elevated levels of the AMP azurocidin-1 was associated with reduced systemic inflammation, indicated by lower C-reactive protein. Overall, this study identified expression of previously undescribed AMPs in the context of human PN. These proteins may play a pivotal role in protection from severe diseases and systemic inflammation.

immunology↗

bioRxiv2023

Tryptophan stress activates EGFR-RAS-signaling to MTORC1 and p38/MAPK to sustain translation and AHR-dependent autophagy

Tumours face tryptophan (Trp) depletion, but the mechanisms sustaining protein biosynthesis under Trp stress remain unclear. We report that Trp stress increases the levels of the translation repressor EIF4EBP1. Yet, at the same time, EIF4EBP1 is selectively phosphorylated by the metabolic master regulator MTORC1 kinase, preventing EIF4EBP1 from inhibiting translation. MTORC1 activity under Trp stress is unexpected because the absence of amino acids is typically linked with MTORC1 inhibition. EIF4EBP1-sensitive translation in Trp starved cells is sustained by EGFR and RAS signalling to MTORC1. Via this mechanism, Trp stress enhances the synthesis and activity of the aryl hydrocarbon receptor (AHR). This is noteworthy as Trp catabolites are known to activate AHR, and therefore Trp stress was previously considered to inhibit AHR. Trp stress-induced AHR enhances the expression of key regulators of autophagy, which sustains intracellular Trp levels and Trp-charged tRNAs for translation. Hence, Trp stress switches MTORC1 from its established inhibitory function into an enhancer of autophagy, acting through AHR. The clinical potential of this fundamental mechanism is highlighted by the activity of the mTORC1-AHR pathway and an autophagy signature in 20% of glioblastoma patients, opening up new avenues for cancer therapy.

biochemistry↗

bioRxiv2022

TAOK2β represses translation via phosphorylation of eEF2 and ameliorates exaggerated protein synthesis in a mouse model of 16p11.2 microdeletion-driven autism

Microdeletions in the 16p11.2 region of the human genome are frequently associated with autism spectrum disorders (ASDs), but how these genomic rearrangements cause ASD remains unclear. Here, we reveal that TAOK2{beta}, a protein isoform encoded by the human TAOK2 gene located in the 16p11.2 locus, regulates mRNA translation. To identify key functional interaction partners of TAOK2{beta}, we performed proteomic screening from Neuro-2a (N2a) cells, mouse cortices, and cultured neurons. This revealed translation factors as a major class of enriched interacting proteins. Consistently, TAOK2{beta} is present in mouse cortical polyribosomes and cortices from Taok2 knockout mice show increased ribosome density on mRNAs and enhanced protein synthesis. Several lines of evidence support an effect of TAOK2{beta} on translation elongation via phosphorylation of eukaryotic elongation factor (eEF2). TAOK2 can directly phosphorylate eEF2 on Threonine 56 and this phosphorylation is reduced in cortices from Taok2 knockout mice. TAOK2{beta} WT overexpression increased eEF2 phosphorylation levels and reduced protein synthesis, whereas a kinase-dead allele of TAOK2{beta} showed opposite effects. Finally, we show that cortices from the mouse model of the human 16p11.2 microdeletion have increased polysome/monosome (P/M) ratios and protein synthesis, phenocopying Taok2 loss of function. Importantly, defective translation phenotypes observed in the mouse 16p11.2 microdeletion model of ASD could be normalized either by reintroducing Taok2 in vivo or by delivering TAOK2{beta} to cortical neurons derived from 16p11.2 microdeletion mice. Our results uncover a critical role of TAOK2{beta} as a regulator of protein synthesis and support the idea that translational control is a common endpoint of ASD-associated signaling pathways.

neuroscience↗

bioRxiv2022

Comparative assessment of quantification methods for tumor tissue phosphoproteomics

With increasing sensitivity and accuracy in mass spectrometry, the tumor phosphoproteome is getting into reach. However, the selection of quantitation techniques best-suited to the biomedical question and diagnostic requirements remains a trial and error decision as no study has directly compared their performance for tumor tissue phosphoproteomics. We compared label-free quantification (LFQ), spike-in-SILAC (stable isotope labeling by amino acids in cell culture) and TMT isobaric tandem mass tags technology for quantitative phosphosite profiling in tumor tissue. TMT offered the lowest accuracy and the highest precision and robustness towards different phosphosite abundances and matrices. Spike-in-SILAC offered the best compromise between these features but suffered from a low phosphosite coverage. LFQ offered the lowest precision but the highest number of identifications. Both spike-in-SILAC and LFQ presented susceptibility to matrix effects. Match between run (MBR)-based analysis enhanced the phosphosite coverage across technical replicates in LFQ and spike-in-SILAC but further reduced the precision and robustness of quantification. The choice of quantitative methodology is critical for both study design such as sample size in sample groups and quantified phosphosites, and comparison of published cancer phosphoproteomes. Using ovarian cancer tissue as an example, our study builds a resource for the design and analysis of quantitative phosphoproteomic studies in cancer research and diagnostics.

biochemistry↗

bioRxiv2020

Centrosome-dependent microtubule organization sets the conditions for axon formation

Microtubule remodeling is critical during axon development when the more stable microtubules populate the axon. It is not completely understood, however, how this local cytoskeleton remodeling is coordinated. The centrosome, the main microtubule-organizing center (MTOC), has been suggested to be crucial for axon specification 1-5. Conversely, it was proposed that axon elongation is independent of centrosomal functions 6. Here we report that microtubule dynamics in early neurons follow a radial organization which establishes the conditions for the axon formation. Using high-resolution microscopy of early developing neurons, we demonstrate that few somatic acetylated microtubules are restricted near the centrosome. At later stages, however, acetylated microtubules spread out in the soma and concentrate in the growing axon. Furthermore, live-imaging of the microtubule plus-end binding protein EB3 in early differentiating neurons shows that growing microtubules have increased length and growth speed near the MTOC, suggesting local differences that might favor axon selection. Importantly, due to the lack of somatic stable/acetylated microtubules in early developing neurons, disruption of the F-actin cytoskeleton does not induce multiple axons, as it does at later stages of differentiation. Finally, we demonstrate that overexpression of the centrosomal protein 120 (Cep120), known for promoting microtubule acetylation and stabilization, induces multiple axons, while its downregulation decreases the content of proteins regulating microtubule dynamics and stability, hence hampering axon formation. Collectively, our data show that early centrosome-dependent microtubule organization contributes to axon formation.

cell biology↗