Biology subjects

Feederle, R.

Publications and source records attributed to Feederle, R..

13 recordsLinked to original sources

bioRxiv2026

RTTN moonlights beyond the centrosome to control ribosome biogenesis and tRNA modification in human brain organoids

RTTN (rotatin) is a centrosomal protein mutated in severe malformations of cortical development, yet how its dysfunction disrupts human corticogenesis has remained unclear. Here, we show that RTTN has an unrecognized function at the core of the translation machinery. Using human telencephalic and hippocampal organoids carrying distinct RTTN alleles, together with single-cell and bulk transcriptomics, polysome profiling, and tRNA pseudouridine sequencing, we find that RTTN is enriched in cycling first-trimester neural progenitors and physically associates with ribosome-biogenesis and RNA-processing factors. RTTN mutations impair rRNA biogenesis and polysome assembly, reduce cytoplasmic ribosome density and nascent protein synthesis, and remodel the tRNA pseudouridylation landscape through both a PUS7L-dependent variable-arm signature and a broader RTTN-specific defect. These translational deficits are accompanied by prolonged mitosis, reduced entry into S-phase, and impaired interkinetic nuclear migration in mutant progenitors. Our findings redefine RTTN as a regulator of ribosome homeostasis and mRNA translation and implicate defective translational capacity as a driver of RTTN-associated microcephaly. Graphical AbstractRTTN sustains ribosome and tRNA homeostasis in human neural progenitors; its mutation disrupts mRNA translation, stalling progenitor proliferation and interkinetic nuclear migration, and driving cortical malformation and growth failure. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/744412v1_ufig1.gif" ALT="Figure 1"> View larger version (57K): org.highwire.dtl.DTLVardef@16124aaorg.highwire.dtl.DTLVardef@ae23f7org.highwire.dtl.DTLVardef@bb588forg.highwire.dtl.DTLVardef@1b35028_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2026

PINK1 stabilizes RMDN3-VAPB interaction without altering the landscape of mito-ER contact sites

Mitochondria-ER contact sites (MERCS) have emerged as central hubs of organellar biology, participating in the exchange of lipids, Ca2+ and even protein precursors. During mitochondrial dysfunction, the contact between the ER-resident protein VAPB and the outer mitochondrial membrane protein RMDN3 (PTPIP51) increases, yet the mechanism stabilizing this connection despite ongoing mitophagy remains to be determined. Here we develop a splitAPEX2 based proximity biotinylation tool to assess the interaction of VAPB and RMDN3. We determine that activation of the mitochondrial depolarization-induced kinase PINK1 leads to phosphorylation of RMDN3 at T160, which is necessary to increase the interaction with VAPB in the presence of mitochondrial depolarization. Mass Spectrometric analysis of the RMDN3-VAPB MERCS proteome reveals that these contacts also include peroxisomal proteins and that the proteomic composition remains stable during mitochondrial depolarization.

cell biology↗

bioRxiv2026

Mitochondrial hitch-hiking of Mapt mRNA maintains Tau levels in axons

Hitch-hiking of transcripts on organelles, including mitochondria, has emerged as a common pathway to transport mRNAs into the axon to enable their local translation. However, the extent of mitochondrial mRNA association in vivo has not been established. Here we report the landscape of mitochondrial mRNA association in two nerve tissues, the retinal ganglion cell derived optic nerve and the motor neuron derived part of the sciatic nerve. This revealed astonishing tissue diversity between mitochondria associated mRNAs that can partly be explained by the lack of expression of the mitochondrially anchored RNA-binding protein SYNJ2a in motor neurons. This diversity led us to discover that also MAPT mRNA, encoding the axon-enriched microtubule associated protein Tau, is transported via the mitochondrial RNA anchoring complex SYNJ2a/SYNJ2BP into axons of glutamatergic neurons. Loss of SYNJ2BP prevents axonal localization of MAPT and miss-sorting of Tau, consequently stunting axonal growth in cultured hippocampal neurons. This connects mitochondrial transport to the maintenance of the local cytoskeleton and axonal growth in glutamatergic neurons.

neuroscience↗

bioRxiv2025

Development of Motif-Specific Monoclonal Antibodies for Global Protein Citrullination Detection with Minimal Cross-Reactivity to Homocitrullination

Protein citrullination, a post-translational modification (PTM) catalyzed by peptidylarginine deiminases (PADs), plays critical roles in biological processes such as immunity, gene regulation, and inflammation. Dysregulated citrullination is implicated in diseases including rheumatoid arthritis, multiple sclerosis, and cancer, making it a potential biomarker and therapeutic target. Immunodetection is the most commonly used technique to study citrullination. However, most commercially available antibodies against citrullination are either protein-specific or lack the sensitivity and specificity of the broad variety of modified proteins. In addition, existing anti-pan citrullination antibodies often fail to distinguish citrullination from homocitrullination, a chemically similar modification on lysine. This cross-reactivity limits their utility in deciphering the distinct biological roles of these PTMs. To address these challenges, we employed a motif-based strategy to generate monoclonal antibodies against citrullination. This approach leverages PAD enzyme sequence preferences from human proteome, enhancing specificity for citrullination while minimizing cross-reactivity with homocitrullination. We immunized rats with a pool of over 490,000 citrullinated peptides, designed to represent common citrullination motifs in human tissue proteomes. Two monoclonal antibody clones were established and validated for sensitivity and specificity using ELISA and western blot against in vitro citrullinated and homocitrullinated proteomes, as well as ionomycin-activated human neutrophils. Both clones demonstrated great sensitivity to diverse citrullinated proteins, robust discrimination against homocitrullination, and quantitative readout in biological samples. These novel antibodies provide powerful tools for studying global citrullination dynamics and hold promise for biomarker discovery and diagnostic applications in diseases involving PAD dysregulation.

molecular biology↗

bioRxiv2024

Full-length Cryptochrome 1 in the outer segments of the retinal blue cone photoreceptors in humans and great apes suggests a role beyond transcriptional repression

Mammalian cryptochrome 1 (CRY1) is a central player in the circadian transcription-translation feedback loop, crucial for maintaining a roughly 24-hour rhythm. CRY1 was suggested to also function as blue-light photoreceptor in humans and has been found to be expressed at the mRNA level in various cell types of the inner retina. However, attempts to detect CRY1 at the protein level in the human retina have remained unsuccessful so far. Using various C-terminal specific antibodies recognizing full-length CRY1 protein, we consistently detected selective labelling in the outer segments of short wavelength-sensitive (SWS1, "blue") cone photoreceptor cells across human, bonobo, and gorilla retinae. No other retinal cell types were stained, which is in contrast to what would be expected of a ubiquitous clock protein. Subcellular fractionation experiments in transfected HEK cells using a C-terminal specific antibody located full-length CRY1 in the cytosol and membrane fractions. Our findings indicate that human CRY1 has several different functions including at least one non-clock function. Our results also raise the likely possibility that several different versions of CRY1 exists in humans. We suggest that truncation of the C-terminal tail, maybe to different degrees, may affect the localization and function of human CRY1.

cell biology↗

bioRxiv2024

The CHCHD2-CHCHD10 protein complex is modulated by mitochondrial dysfunction and alters lipid homeostasis in the mouse brain.

The highly conserved CHCHD2 and CHCHD10 are small mitochondrial proteins residing in the intermembrane space. Recently, mutations in the CHCHD2 and CHCHD10 genes have been linked to severe disorders, including Parkinsons disease and amyotrophic lateral sclerosis. In cultured cells, a small fraction of CHCHD2 and CHCHD10 oligomerize to form a high molecular weight complex of unknown function. Here, we generated a whole-body Chchd2 knockout mouse to investigate the in vivo role of CHCHD2 and its protein complex. We show that CHCHD2 is crucial for sustaining full motor capacity, normal striatal dopamine levels, and lipid homeostasis in the brain of adult male mice. We also demonstrate that in mouse tissues, CHCHD2 and CHCHD10 exist exclusively as a high molecular weight complex, whose levels are finely tuned under physiological conditions. In response to mitochondrial dysfunction, the abundance and size of the CHCHD2-CHCHD10 complex increases, a mechanism conserved across different tissues. Although the loss of CHCHD2 does not abolish CHCHD10 oligomerization, it enhances cell vulnerability to mitochondrial stress, suggesting that CHCHD2 is protective against mitochondrial damage. Our findings uncover the role of CHCHD2 in preserving tissue homeostasis and provide important insights into the involvement of the CHCHD2-CHCHD10 complex in human diseases.

neuroscience↗

bioRxiv2024

The nuclear speckles protein SRRM2 is a new therapeutic target molecule on the surface of cancer cells

The membrane composition of extracellular vesicles (EVs) largely reflects that of the plasma membrane of the cell of origin. We therefore hypothesized that EVs are a source for the detection of hitherto unknown tumor-associated druggable target molecules. For this, we used EVs derived from cancer cell lines for an immunization of a rat. From this immunization, we obtained a monoclonal antibody specific for SRRM2, a protein involved in splicing a major component of nuclear speckles. Here, we used this antibody to demonstrate that SRRM2 is exposed at the surface of most cancer cell lines from various entities and, even more important, on cancer cells in vivo. Moreover, we demonstrate that SRRM2-specific CAR-T cells are killing SRRM2-positive cancer cells electively. Collectively, we identified SRRM2 as a promising new target molecule exposed on the cancer cell surface and show that our SRRM2-specific antibody can be used as a basis for the development of new targeted cancer therapies.

cancer biology↗

bioRxiv2023

Structures of Arabidopsis thaliana MDL Proteins and Synergistic Effects with the Cytokine MIF on Human Receptors

Vertebrates have developed effective immune mechanisms to fight microbial attacks, relying on a sophisticated network of innate and adaptive responses, a circulatory system, and numerous orchestrating soluble mediators such as cytokines. Mammalian macrophage migration inhibitory factor (MIF) and its paralog D-dopachrome tautomerase (D-DT/MIF-2) are multifunctional inflammatory cytokines with chemokine-like properties that modulate immunity. Plants possess orthologous MIF/D-DT-like (MDL) proteins, whose function is largely unexplored. Driven by the previous discovery of cross-kingdom mimicry of plant (Arabidopsis thaliana) MDL proteins and human MIF receptor signaling, we here characterized the structures of the three A. thaliana MDLs by X-ray crystallography and explored the mechanism underlying the interplay between plant MDLs, human MIF, and its receptors. We obtained high-resolution structures at 1.56 [A], 1.40 [A], and 2.00 [A] resolution for MDL1, MDL2, and MDL3, respectively, revealing a typical trimeric assembly and a high three-dimensional similarity to human MIF. Although residues at the catalytic site of the three MDLs show high identity to human MIF, the proteins showed low tautomerase activity for the substrate 4-hydroxyphenylpyruvate (HPP). Structural differences likely explain the enzymatic inactivity of plant MDLs for HPP. Strikingly, employing in vitro, in vivo, and in planta test systems, we found that MIF and MDL proteins interact with each other and have the capacity to form hetero-oligomeric complexes. The functional consequences of this interaction were demonstrated applying a yeast-based reporter system specific for the MIF chemokine receptors CXCR2 and CXCR4. MDLs not only triggered receptor signaling on their own, but exhibited pronounced synergism regarding the activation of the CXCR2- and CXCR4-dependent signaling pathways, when co-applied with MIF. These findings were substantiated by the co-administration of pharmacological inhibitors that either disrupt MIF receptor binding or block the catalytic cavity. Moreover, biochemical and biophysical experiments using an allosteric oligomer-specific MIF inhibitor established hexa-oligomer formation between MIF and MDLs as the putative basis for the synergistic effect. Our results are the starting point for a mechanistic understanding of the immunomodulatory activity of a family of highly conserved plant proteins. One Sentence SummaryA. thaliana MDLs and human macrophage inhibitory factor interact with each other and induce synergistic effects in activating CXCR2 and CXCR4.

immunology↗

bioRxiv2022

Depletion of the RNA-binding protein PURA triggers changes in posttranscriptional gene regulation and loss of P-bodies

The RNA-binding protein PURA has been implicated in the rare, monogenetic, neurodevelopmental disorder PURA Syndrome. PURA binds both DNA and RNA and has been associated with various cellular functions. Only little is known about its main cellular roles and the molecular pathways affected upon PURA depletion. Here, we show that PURA is predominantly located in the cytoplasm, where it binds to thousands of mRNAs. Many of these transcripts change abundance in response to PURA depletion. The encoded proteins suggest a role for PURA in immune responses, mitochondrial function, autophagy and processing (P)-body activity. Intriguingly, reduced PURA levels decrease the expression of the integral P-body components LSM14A and DDX6 and strongly affect P-body formation in human cells. Furthermore, PURA knockdown results in stabilization of P-body-enriched transcripts, whereas other mRNAs decrease. Hence, reduced PURA levels, as reported in patients with PURA Syndrome, influence the formation and composition of this phase-separated RNA processing machinery. Our study proposes PURA Syndrome as a new model to study the tight connection between P-body-associated RNA regulation and neurodevelopmental disorders. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=150 SRC="FIGDIR/small/479353v2_ufig1.gif" ALT="Figure 1"> View larger version (49K): org.highwire.dtl.DTLVardef@5712f9org.highwire.dtl.DTLVardef@9eb133org.highwire.dtl.DTLVardef@83b7b2org.highwire.dtl.DTLVardef@1912e5c_HPS_FORMAT_FIGEXP M_FIG C_FIG

molecular biology↗

bioRxiv2022

Active site geometry stabilization of a presenilin homolog by the lipid bilayer promotes intramembrane proteolysis

Cleavage of membrane proteins in the lipid bilayer by intramembrane proteases is crucial for health and disease. Although different lipid environments can potently modulate their activity, how this is linked to their structural dynamics is unclear. Here we show that the carboxy-peptidase-like activity of the archaeal intramembrane protease PSH, a homolog of the Alzheime[r]s disease-associated presenilin/{gamma}-secretase is impaired in micelles and promoted in a lipid bilayer. Comparative molecular dynamics simulations revealed that important elements for substrate binding such as transmembrane domain 6a of PSH are more labile in micelles and stabilized in the lipid bilayer. Moreover, consistent with an enhanced interaction of PSH with a transition- state analog inhibitor, the bilayer promoted the formation of the enzymes catalytic active site geometry. Our data indicate that the lipid environment of an intramembrane protease plays a critical role in structural stabilization and active site arrangement of the enzyme-substrate complex thereby promoting intramembrane proteolysis.

biochemistry↗

bioRxiv2021

Structural Basis for Mis18 Complex Assembly: Implications for Centromere Maintenance

The centromere, defined by the enrichment of CENP-A (a Histone H3 variant) containing nucleosomes, is a specialised chromosomal locus that acts as a microtubule attachment site. To preserve centromere identity, CENP-A levels must be maintained through active CENP-A loading during the cell cycle. A central player mediating this process is the Mis18 complex (Mis18, Mis18y and Mis18BP1), which recruits the CENP-A specific chaperone HJURP to centromeres for CENP-A deposition. Here, using a multi-pronged approach, we characterise the structure of the Mis18 complex and show that multiple hetero- and homo-oligomeric interfaces facilitate the hetero-octameric Mis18 complex assembly composed of 4 Mis18, 2 Mis18y and 2 Mis18BP1. Evaluation of structure-guided/separation-of-function mutants reveals structural determinants essential for Mis18 complex assembly and centromere maintenance. Our results provide new mechanistic insights on centromere maintenance, highlighting that while Mis18 can associate with centromeres and deposit CENP-A independently of Mis18y, the latter is indispensable for the optimal level of CENP-A loading required for preserving the centromere identity.

cell biology↗

bioRxiv2020

Defining the RBPome of T helper cells to study higher order post-transcriptional gene regulation

Post-transcriptional gene regulation is complex, dynamic and ensures proper T cell function. The targeted transcripts can simultaneously respond to various factors as evident for Icos, an mRNA regulated by several RNA binding proteins (RBPs), including Roquin. However, fundamental information about the entire RBPome involved in post-transcriptional gene regulation in T cells is lacking. Here, we applied global RNA interactome capture (RNA-IC) and orthogonal organic phase separation (OOPS) to human and mouse primary T cells and identified the core T cell RBPome. This defined 798 mouse and 801 human proteins as RBPs, unexpectedly containing signaling proteins like Stat1, Stat4 and Vav1. Based on the vicinity to Roquin-1 in proximity labeling experiments, we selected [~]50 RBPs for testing coregulation of Roquin targets. Induced expression of these candidate RBPs in wildtype and Roquin-deficient T cells unraveled several Roquin-independent contributions, but also revealed Celf1 as a new Roquin-1-dependent and target-specific coregulator of Icos. One sentence statementWe provide an atlas of RNA-binding proteins in human and mouse T helper cells as a resource for studying higher order post-transcriptional gene regulation.

immunology↗

bioRxiv2020

Lysine acetylation of TDP-43 drives phase separation and pathological aggregation

The trans-activation response DNA-binding protein TDP-43 regulates RNA processing and forms neuropathological aggregates in patients with amyotrophic lateral sclerosis and frontotemporal lobar degeneration. Investigating TDP-43 post-translational modifications, we discovered that K84 acetylation reduced nuclear import whereas K136 acetylation impaired RNA binding and splicing capabilities of TDP-43. Such failure of RNA interaction triggered TDP-43 phase separation mediated by the C-terminal low complexity domain, leading to the formation of insoluble aggregates with pathologically phosphorylated and ubiquitinated TDP-43. Confirming the results from site-directed mutagenesis, we succeeded to introduce authentic acetyl-lysine at the identified sites via amber suppression. [AcK84]TDP-43 showed cytoplasmic mislocalization and the aggregation propensity of [acK136]TDP-43 was confirmed. With newly developed antibodies, we found that the nuclear sirtuin SIRT1 can potently deacetylate [acK136]TDP-43. Moreover, SIRT1 reduced the aggregation propensity of [acK136]TDP-43. Thus, distinct lysine acetylations modulate nuclear import, RNA binding and phase separation of TDP-43, suggesting novel regulatory mechanisms for TDP-43 pathogenesis.

cell biology↗