Biology subjects

Franz-Wachtel, M.

Publications and source records attributed to Franz-Wachtel, M..

16 recordsLinked to original sources

bioRxiv2026

A host-directed virulence factor of Clostridium perfringens is modulated by gut commensal strains

In a healthy host, the residential microbes help regulate the growth of pathobionts, which are common members of the human gut microbiome, preventing them from causing diseases, including infections, under certain conditions. In cases of dysbiosis, this protection may be compromised. Targeted microbiome modulation offers a promising approach to restore healthy conditions in a disrupted community and consequently prevent infections using the natural colonization resistance of the microbiome. Elucidating the interaction mechanisms between microbial species within a microbiome is crucial for understanding how a microbiome can be modulated precisely and effectively to benefit the hosts well-being. Here, we investigated the interactions between the pathobiont C. perfringens and human gut commensals on physiological and molecular levels. We found that commensal strains affect C. perfringens growth by competing for substrates such as amino acids or a carbon source other than glucose. We further observed that Bacteroidaceae strains altered the levels of C. perfringens proteins, among others, the host-directed {theta}-toxin. Our findings reinforce the notion that modulating the composition of the gut microbiome is an effective strategy to prevent infections.

microbiology↗

bioRxiv2025

The ribosome-associated complex regulates cytosolic translation upon mitoprotein-induced stress

The biogenesis of mitochondria relies on the import of newly synthesized precursor proteins from the cytosol. Tom70 is a mitochondrial surface receptor which recognizes precursors and serves as an interface between mitochondrial protein import and the cytosolic proteostasis network. Mitochondrial import defects trigger a complex stress response, in which compromised protein synthesis rates are a characteristic element. The molecular interplay that connects mitochondrial (dys)function to cytosolic translation rates in yeast cells is however poorly understood. Here, we show that the deletion of the two Tom70 paralogs of yeast (TOM70 and TOM71) leads to defects in mitochondrial biogenesis and slow cell growth. Surprisingly, upon heat stress, the deletion of ZUO1, a chaperone of the ribosome-associated complex (RAC), largely prevented the slow growth and the reduced translation rates in the tom70{Delta}/tom71{Delta} double deletion mutant. In contrast, the mitochondrial defects were not cured but even enhanced by ZUO1 deletion. Our study shows that Zuo1 is a critical component in the signaling pathway that mutes protein synthesis upon mitochondrial dysfunction. We propose a novel paradigm according to which RAC serves as a stress-controlled regulatory element of the cytosolic translation machinery.

biochemistry↗

bioRxiv2025

Identification of the negamycin split biosynthetic gene cluster in Kitasatospora purpeofusca ATCC21470

Negamycin is a ribosome-targeting antibiotic with activity against Gram-positive and Gram-negative bacteria including ESKAPE pathogens. Furthermore, it promotes premature stop codon readthrough. Its therapeutic potential is limited by low natural production and synthetic complexity. To enable scalable biosynthesis, we identified and characterized its genetic basis in Kitasatospora purpeofusca ATCC 21470. Two distant chromosomal regions, neg1 and neg2, were found to be essential. Deletion of neg1, involved in nitrite provision for N-N bond formation, reduced production to [~]10%, while deletion of neg2, which directs {beta}-lysine generation and scaffold assembly, abolished it completely. Isotope-labeling experiments confirmed nitrite incorporation. Transcriptomic and proteomic analyses further supported the involvement of both regions. The heterologous expression of neg1 along with the neg2 region in Streptomyces albidoflavus reconstituted negamycin biosynthesis, confirming the unusual involvement of two distant gene clusters in the biosynthesis, and provides a foundation for biotechnological production and further development of this promising antibiotic. SIGNIFICANCEThe rapid rise of antimicrobial resistance (AMR), particularly among Gram-negative ESKAPE pathogens, represents one of the most urgent global health threats. Despite this, the discovery and development of new antibiotics have stagnated. Addressing this challenge requires the exploration of natural products with novel mechanisms of action, alongside the development of scalable production strategies. Negamycin has emerged as a compelling candidate in this regard, characterized by an unusual mechanism of action and therapeutic potential extending beyond traditional antibacterial use. However, its development has been constrained by low production yields in the native producer. In this study, we identify and characterize the biosynthetic genes responsible for negamycin production, providing a foundation for pathway engineering, yield optimization, and the rational design of new analogs.

microbiology↗

bioRxiv2025

Arabidopsis HYPERSENSITIVE INDUCED REACTION 2 affects plasma membrane receptor pathways and organization

The distribution of proteins across the plasma membrane is not uniform; however, the principles governing their organization remain not fully understood. Hypersensitive-induced reaction (HIR) proteins are plant-specific members of the stomatin/prohibitin/flotillin/HflK/C (SPFH) family that have been shown to influence membrane organization. Arabidopsis thaliana HIR2 interacts with multiple plasma membrane proteins, including receptor kinases such as BAK1-INTERACTING RECEPTORS 2 and 3 (BIR2 and 3), BRI1-ASSOCIATED KINASE 1 (BAK1), FLAGELLIN SENSING 2 (FLS2), and BRASSINOSTEROID INSENSITIVE 1 (BRI1). These interactions connect HIR2 to BAK1-mediated signaling pathways, as evidenced by impaired growth and immunity phenotypes in hir2 mutants. HIR2 is anchored to the inner leaflet of the plasma membrane through a hydrophobic interaction domain and S-acylation. Using single-particle tracking photoactivated localization microscopy (sptPALM), we showed that HIR2 affects receptor kinase dynamics and clustering, suggesting a role in spatially coordinating receptor complex activities. Structural modeling with AlphaFold 3 predicts a multimeric circular cup-like assembly for HIR2, consistent with high molecular weight complexes identified through blue native polyacrylamide gel electrophoresis. These findings indicate that HIR2 forms a discrete membrane compartment, providing a novel structural framework for spatial membrane organization and thereby modulating the function of membrane-resident receptors.

plant biology↗

bioRxiv2025

Effector-triggered processing body formation attenuates host translation via ER stress responses and autophagy upon bacterial infection

Pathogens employ sophisticated strategies to modulate host protein homeostasis by targeting proteolytic pathways, but their impact on protein synthesis remains elusive. We report that pathogenic bacteria Pseudomonas syringae (Pst) targets ribonucleoprotein condensates, known as processing bodies (P-bodies), to attenuate host translation through two effectors with liquid-like properties. We uncovered a previously unknown link that Pst-mediated repression of the ER stress response is required for P-body assembly. Furthermore, we identify a novel intersection between P-bodies and autophagy, demonstrating that autophagic clearance of P-bodies is crucial for maintaining the balance between translationally active and inactive mRNAs. Altogether, our discoveries provide novel insights on how host translation is attenuated by bacteria to dampen plant immunity and uncover unknown connections between ER stress responses and autophagy with P-body dynamics.

plant biology↗

bioRxiv2024

The T4bSS of Legionella features a two-step secretion pathway with an inner membrane intermediate for secretion of transmembrane effectors

To promote intracellular survival and infection, Legionella spp. translocate hundreds of effector proteins into eukaryotic host cells using a type IV b protein secretion system (T4bSS). T4bSS are well known to translocate soluble as well as transmembrane domain-containing effector proteins (TMD-effectors) but the mechanisms of secretion are still poorly understood. Herein we investigated the secretion of hydrophobic TMD-effectors, of which about 80 were previously reported to be encoded by L. pneumophila. A proteomic analysis of fractionated membranes revealed that TMD-effectors are targeted to and inserted into the bacterial inner membranes of L. pneumophila independent of the presence of a functional T4bSS. While the T4bSS chaperones IcmS and IcmW were critical for secretion of all tested TMD-effectors, they did not influence inner membrane targeting of these proteins. As for soluble effector proteins, translocation of TMD-effectors into host cells depended on a C-terminal secretion signal and this signal needed to be presented towards the cytoplasmic side of the inner membrane. A different secretion behavior of TMD- and soluble effectors and the need for small periplasmic loops within TMD-effectors provided strong evidence that TMD-effectors are secreted in a two-step secretion process: Initially, an inner membrane intermediate is formed, that is extracted towards the cytoplasmic side, possibly by the help of the type IV coupling protein complex and subsequently secreted into eukaryotic host cells by the T4bSS core complex. Overall, our study highlights the amazing versatility of T4bSS to secrete soluble and TMD-effectors from different subcellular locations of the bacterial cell.

microbiology↗

bioRxiv2023

ER-anchored protein sorting controls the fate of two proteasome activators for intracellular organelle communication during proteotoxic stress

Proteotoxic stress, characterized by the accumulation of damaged proteins, poses a significant challenge to cellular homeostasis. To mitigate proteotoxicity eukaryotes employ the proteasome that is regulated by proteasome activators, e.g. transcription factors that promote gene expression of proteasome subunits. As proteotoxicity originates in different compartments, cells need to perceive signals from various locations. Understanding which components integrate signals to address proteotoxicity is essential to develop strategies to cope with proteotoxicity but remain elusive. Here, we identify that the proteasome autoregulatory feedback loop acts as a gatekeeper to facilitate the communication between nucleus and chloroplast. We reveal that the ER-anchored protein sorting system (ERAPS) controls the proteasomal degradation or nuclear translocation of proteasome activators NAC53 and NAC78. While both transcription factors activate the proteasome gene expression, they repress photosynthesis-associated nuclear genes during proteotoxicity through association with a conserved cis-element. Our data implicate a general trade-off between proteasome function and energy metabolism unravelling an unprecedented mechanism of how eukaryotic cells cope with proteotoxicity. Collectively, our discoveries provide a novel conceptual framework in which the proteasome autoregulatory feedback loop coordinates subcellular proteostasis and the trade-off between growth and defence.

plant biology↗

bioRxiv2023

Characterization of the Pristionchus pacificus epigenetic toolkit reveals the evolutionary loss of the histone methyltransferase complex PRC2

Comparative approaches have revealed both divergent and convergent paths to achieving shared developmental outcomes. Thus, only through assembling multiple case studies can we understand biological principles. Yet, despite appreciating the conservation - or lack thereof - of developmental networks, the conservation of epigenetic mechanisms regulating these networks is poorly understood. The nematode Pristionchus pacificus has emerged as a model system of plasticity and epigenetic regulation as it exhibits a bacterivorous or omnivorous morph depending on its environment. Here, we determined the "epigenetic toolkit" available to P. pacificus as a resource for future functional work on plasticity, and as a comparison with C. elegans to investigate the conservation of epigenetic mechanisms. Broadly, we observed a similar cast of genes with putative epigenetic function between C. elegans and P. pacificus. However, we also found striking differences. Most notably, the histone methyltransferase complex PRC2 appears to be missing in P. pacificus. We described the deletion/pseudogenization of the PRC2 genes mes-2 and mes-6 and concluded that both were lost in the last common ancestor of P. pacificus and a related species P. arcanus. Interestingly, we observed the enzymatic product of PRC2 (H3K27me3) by mass spectrometry and immunofluorescence, suggesting that a currently unknown methyltransferase has been co-opted for heterochromatin silencing. Altogether, we have provided an inventory of epigenetic genes in P. pacificus to enable reverse-genetic experiments related to plasticity, and in doing so have described the first loss of PRC2 in a multicellular organism.

genetics↗

bioRxiv2022

Horizontal transfer of bacteriocin biosynthesis genes requires metabolic adaptation to improve compound production and cellular fitness

Biosynthetic gene clusters (BGCs) encoding the production of bacteriocins are widespread amongst bacterial isolates and are important genetic determinants of competitive fitness within a given habitat. Staphylococci produce a tremendous diversity of compounds and the corresponding BGCs are frequently associated with mobile genetic elements, suggesting gain and loss of biosynthetic capacity. Pharmaceutical biology has shown that compound production in heterologous hosts is often challenging and many BGC recipients produce initially low compound amounts or show reduced growth rates. To assess whether transfer of BGCs between closely related S. aureus strains can be instantly effective or requires elaborate metabolic adaptation, we investigated the intra species transfer of a BGC encoding the ribosomally synthesized and post-translationally modified peptide (RiPP) micrococcin P1 (MP1). We found that acquisition of the BGC by S. aureus RN4220 enabled immediate MP1 production but also imposed a metabolic burden, which was relieved after prolonged cultivation by adaptive mutation. We used a multiomics approach to study this phenomenon and found adaptive evolution to select for strains with increased activity of the tricarboxylic acid cycle (TCA), which enhanced metabolic fitness and levels of compound production. Metabolome analysis revealed increases of central metabolites including citrate and -ketoglutarate in the adapted strain, suggesting metabolic adaptation to overcome the BGC-associated growth defects. Our results indicate that BCG acquisition requires genetic and metabolic predispositions allowing the integration of bacteriocin production into the cellular metabolism. Inappropriate metabolic characteristics of recipients can entail physiological burdens, negatively impacting the competitive fitness of recipients within natural bacterial communities. ImportanceHuman microbiomes are critically associated with human health and disease. Importantly, pathogenic bacteria can hide in human associated communities and can cause disease when the composition of the community becomes dysbalanced. Bacteriocin producing commensals are able to displace pathogens from microbial communities, suggesting that their targeted introduction in human microbiomes might prevent pathogen colonisation and infection. However, in view of future probiotic approaches, strains are needed that produce high levels of bioactive compounds and retain cellular fitness within mixed bacterial communities. Our work offers insights into the metabolic burdens associated with the production of the bacteriocin micrococcin P1 and highlights evolutionary strategies that increase cellular fitness in the context of production. Most likely metabolic adaptations are broadly relevant for bacteriocin producers and need to be considered for the future development of effective microbiome editing strategies.

microbiology↗

bioRxiv2022

Histone 4 lysine 5/12 acetylation provides a plasticity code with epigenetic memory of environmental exposure

Development can be altered to match phenotypes with the environment, and the genetic mechanisms that direct such alternative phenotypes are beginning to be elucidated1,2. Yet, the rules that govern environmental sensitivity vs. invariant development (canalization), and potential epigenetic memory, remain unknown. Here, we show that plasticity of nematode mouth forms is determined by histone 4 lysine 5 and 12 acetylation (H4K5/12ac). Acetylation in early larval stages provides a permissive chromatin state at specific switch genes, which is susceptible to induction during the critical window of environmental sensitivity. As development proceeds deacetylation shuts off switch gene expression to end the critical period. We show that inhibiting deacetylase enzymes leads to long-term epigenetic memory, demonstrating that histone modifications in juveniles can carry environmental information to affect organismal traits in adults. This epigenetic regulation of plasticity appears to be derived from an ancient mechanism of licensing developmental speed that is conserved between flies and nematodes. Thus, H4K5/12ac provides a histone plasticity code with epigenetic potential that can be stored and erased by acetylation and deacetylation, respectively. HighlightsO_LIReciprocal transplant experiments reveal a critical time window of mouth-form plasticity. C_LIO_LIEntry and exit of the critical window is determined by H4K5/12ac at the switch gene eud-1. C_LIO_LIH4K12ac maintains transcriptional competence by supporting elongation. C_LIO_LIInhibition of deacetylation freezes an initial developmental trajectory, resulting in long-term epigenetic memory. C_LIO_LIH4K5/12 acetylation control of plasticity was co-opted from an ancestral role in controlling developmental speed. C_LI

developmental biology↗

bioRxiv2022

A network of cytosolic (co)chaperones promotes the biogenesis of mitochondrial signal-anchored outer membrane proteins

Signal-anchored (SA) proteins are anchored into the mitochondrial outer membrane (OM) via a single transmembrane segment at their N-terminus while the bulk of the proteins is facing the cytosol. These proteins are encoded by nuclear DNA, translated on cytosolic ribosomes, and are then targeted to the organelle and inserted into its OM by import factors. Recently, research on the insertion mechanisms of these proteins into the mitochondrial OM have gained a lot of attention. In contrast, the early cytosolic steps of their biogenesis are unresolved. Using various proteins from this category and a broad set of in vivo, in organello, and in vitro assays, we reconstituted the early steps of their biogenesis. We identified a subset of molecular (co)chaperones that interact with newly synthesized SA proteins, namely, Hsp70 and Hsp90 chaperones and co-chaperones from the Hsp40 family like Ydj1 and Sis1. These interactions were mediated by the hydrophobic transmembrane segments of the SA proteins. We further demonstrate that interfering with these interactions inhibits the biogenesis of SA proteins to varying extents. Finally, we could demonstrate direct interaction of peptides corresponding to the transmembrane segments of SA proteins with the (co)chaperones and reconstitute in vitro the transfer of such peptides from the Hsp70 chaperone to the mitochondrial Tom70 receptor. Collectively, this study unravels an array of cytosolic chaperones and mitochondrial import factors that facilitates the targeting and membrane integration of mitochondrial SA proteins.

biochemistry↗

bioRxiv2021

The TIR-NBS-LRR protein CSA1 is required for autoimmune cell death in Arabidopsis pattern recognition co-receptor bak1 and bir3 mutants

The BRI1-associated kinase BAK1/SERK3 is a positive regulator of multiple leucine rich receptor kinase-mediated signaling pathways including pattern triggered immunity (PTI). Absence or overexpression of BAK1 leads to spontaneous cell death formation. BAK1-interacting receptors (BIR) constitutively interact with BAK1, and plants lacking or overexpressing BIR proteins phenocopy the cell death symptoms observed in bak1 knock outs or overexpressors. In the interactome of BIR3, the TIR-NBS-LRR protein CONSTITUTIVE SHADE-AVOIDANCE 1 (CSA1) was identified by mass spectrometry. CSA1 physically interacts with BIR proteins and can be detected in complexes with BAK1. Direct interaction was shown only for CSA1 with BIR proteins but not BAK1. Double mutant bak1 bir3 genotypes develop strong dwarfism and cell death symptoms that are dependent on EDS1 and salicylic acid. Loss of CSA1 blocks bak1 and bak1 bir3-mediated cell death formation thus demonstrating that CSA1 is causal for this type of cell death. We propose that CSA1 guards BIR proteins and initiates autoimmune cell death that is observed when BAK1 BIR complexes are impaired. Our findings reveal how cell death in the absence of BAK1 and BIR3 is executed and links BAK1, a common co-receptor of many pattern recognition receptors, to NLR proteins typically implicated in effector-triggered immunity.

plant biology↗

bioRxiv2021

Self-ubiquitination of a pathogen type-III effector traps and blocks the autophagy machinery to promote disease

Beyond its role in cellular homeostasis, autophagy plays anti- and pro-microbial roles in host-microbe interactions, both in animals and plants. One prominent role of anti-microbial autophagy is to degrade intracellular pathogens or microbial molecules, in a process termed xenophagy. Consequently, microbes evolved mechanisms to hijack or modulate autophagy to escape elimination. Although well-described in animals, the extent to which xenophagy contributes to plant-bacteria interactions remains unknown. Here, we provide evidence that Xanthomonas campestris pv. vesicatoria (Xcv) suppresses host autophagy by utilizing type-III effector XopL. XopL interacts with and degrades the autophagy component SH3P2 via its E3 ligase activity to promote infection. Intriguingly, XopL is targeted for degradation by defense-related selective autophagy mediated by NBR1/Joka2, revealing a complex antagonistic interplay between XopL and the host autophagy machinery. Our results implicate plant antimicrobial autophagy in depletion of a bacterial virulence factor and unravels an unprecedented pathogen strategy to counteract defense-related autophagy.

plant biology↗

bioRxiv2021

Nemertean, brachiopod and phoronid neuropeptidomics reveals ancestral spiralian signalling systems

Neuropeptides are diverse signalling molecules in animals commonly acting through G-protein coupled receptors (GPCRs). Neuropeptides and their receptors underwent extensive diversification in bilaterians and the relationships of many peptide-receptor systems have been clarified. However, we lack a detailed picture of neuropeptide evolution in lophotrochozoans as in-depth studies only exist for molluscs and annelids. Here we analyse peptidergic systems in Nemertea, Brachiopoda and Phoronida. We screened transcriptomes from thirteen nemertean, six brachiopod and four phoronid species for proneuropeptides and neuropeptide GPCRs. With mass spectrometry from the nemertean Lineus longissimus, we validated several predicted peptides and identified novel ones. Molecular phylogeny combined with peptide-sequence and gene-structure comparisons allowed us to comprehensively map spiralian neuropeptide evolution. We found most mollusc and annelid peptidergic systems also in nemerteans, brachiopods and phoronids. We uncovered previously hidden relationships including the orthologies of spiralian CCWamides to arthropod agatoxin-like peptides and of mollusc APGWamides to RGWamides from annelids, with orthologues systems in nemerteans, brachiopods and phoronids. We found that pleurin neuropeptides previously only found in molluscs are also present in nemerteans and brachiopods. We also identified cases of gene family duplications and losses. These include a protostome-specific expansion of RFamide/Wamide signalling, a spiralian expansion of GnRH-related peptides, and duplications of vasopressin/oxytocin before the divergence of brachiopods, phoronids and nemerteans. This analysis expands our knowledge of peptidergic signalling in spiralians and protostomes. Our annotated dataset of nearly 1,300 proneuropeptide sequences and 600 GPCRs presents a useful resource for further studies of neuropeptide signalling in protostomes.

evolutionary biology↗

bioRxiv2018

β-actin mRNA interactome mapping by proximity biotinylation

The molecular function and fate of mRNAs are controlled by RNA binding proteins (RBPs). However, identification of the interacting proteome of a specific mRNA in vivo is still very challenging. Based on the widely-used RNA tagging with MS2 aptamers for RNA visualization, we developed a novel RNA proximity biotinylation (RNA-BioID) method by tethering biotin ligase (BirA*) via MS2 coat protein (MCP) at the 3UTR of endogenously MS2 tagged {beta}-actin mRNA in mouse embryonic fibroblasts (MEFs). We demonstrate the dynamics of the {beta}-actin mRNA interactome by characterizing its changes upon serum-induced localization of the mRNA. Apart from the previously known interactors, we identified over 60 additional {beta}-actin associated RBPs by RNA-BioID, among them the KH-domain containing protein FUBP3/MARTA2 has shown to be required for {beta}-actin mRNA localization. This protein binds to the 3-untranslated region of {beta}-actin mRNA, is essential for {beta}-actin mRNA localization but does not interact with the characterized {beta}-actin zipcode element. RNA-BioID provides a tool to identify new mRNA interactors and to study the dynamic view of the interacting proteome of endogenous mRNAs in space and time.

cell biology↗

bioRxiv2018

Changes in the neuropeptide complement correlate with nervous system architectures in xenacoelomorphs

Neuropeptides are essential neurosecretory signaling molecules common in protostomes and deuterostomes (together Nephrozoa). Not much, however, is known about the neuropeptide complement of the sister group Xenacoelomorpha. This group is comprised of the three clades Xenoturbella, Nemertodermatida, and Acoela, which differ strongly in their nervous system anatomy. In order to reconstruct the ancestral bilaterian neuropeptide complement and gain insights into neuropeptide evolution within Xenacoelomorpha, we analyzed transcriptomes of 13 acoels, nemertodermatids, and Xenoturbella species. Together with motif searches, similarity searches, mass spectrometry and phylogenetic analyses of neuropeptide precursors and neuropeptide receptors, we reconstruct the xenacoelomorph neuropeptide complement. Our comparison of xenacoelomorph GPCRs with cnidarian and nephrozoan neuropeptide receptors shows that the neuropeptide signaling diversified into at least 20 ancestral peptidergic systems in the lineage to Bilateria. We find that Xenoturbella species possess many of the ancestral bilaterian peptidergic systems and only a few clade-specific neuropeptides. Nemertodermatids seem to have nearly the complete complement of ancestral bilaterian systems and several novel neuropeptides. Acoels show an extensive loss of conserved bilaterian systems, but gained the highest number of novel and group-specific neuropeptides. While it is difficult to correlate the emergence of the bilaterian neuropeptide complement with the evolution of centralized nervous systems, we find a correlation between nervous system novelties and the expansion of taxon-specific neuropeptides in Xenacoelomorpha.

evolutionary biology↗