bioRxiv @biorxivpreprint@qoto.org

During mammalian insemination, most of the deposited sperm are lost by retrograde flow or the immune response of the female reproductive tract. Once semen enters the uterus, seminal fluid and sperm elicit leukocyte infiltration that contributes to the elimination of sperm in the uterus. However, unlike the uterus, invading sperm do not trigger a phagocytic response in the oviduct in the absence of dysfunction or disease states. Thus, the oviduct possesses a distinct immunological microenvironment that tolerates sperm while maintaining the capacity to respond to pathogens. It has been suggested that sperm glycocalyx contributes to innate oviductal tolerance, but the cell and molecular mechanisms are not understood. The current investigation focused on the role of sialic acid-containing glycoconjugates on sperm and their potential to elicit innate tolerance via cognate sialic acid-binding immunoglobulin-type lectins (Siglecs) expressed in the oviduct. In this manuscript, we report our discovery of eight Siglecs (Siglecs-1, -2, -3, -5, -10, -11, -14, -15) expressed in the lower pig oviduct, five of which are known for immune inhibitory functions (Siglecs-2, -3, -5, -10, and -11) and how these may play a role in achieving sperm-induced immune suppression in the oviduct microenvironment. Mass spectrometry profiling of porcine sperm revealed the presence of a mixture of α2,3 and α2,6 linked sialic acids with α2,3-linked sialic acids as the dominant linkage. Of the detected glycans, several sialic acid-containing glycoconjugates were identified as potential ligands for Siglecs (among O-linked glycans: NeuAc1GalNAc1, NeuGc1GalNAc1, NeuAc2Gal1GalNAc1; attached to glycolipids: NeuAc2Gal1GalNAc1Gal1Glc1, Fuc1Gal1GalNAc1NeuAc1Gal1Glc1). This is the first report of Siglec expression in the mammalian oviduct and total glycan analysis of porcine sperm. The results of this study reveal the potential for a sperm-sialoglycan and oviductal-Siglec axis that may contribute to the distinct immunophysiology of the oviduct fundamentally required for undisrupted reproduction in mammals. ### Competing Interest Statement The authors have declared no competing interest.

To facilitate our understanding of the often rapid and nuanced dynamics of extracellularly exposed proteomes during signaling events, it is important to devise robust workflows affording fast time resolution without biases and confounding factors. Here, we present Surface-exposed protein Labeling using PeroxidaSe, H2O2, and Tyramide-derivative (SLAPSHOT), to label extracellularly exposed proteins in a rapid, sensitive, and specific manner, while preserving cellular integrity. This experimentally simple and flexible method utilizes recombinant soluble APEX2 peroxidase that is applied to cells, thus circumventing biological perturbations, tedious engineering of tools and cells, and labeling biases. APEX2 neither requires metal cations for activity nor contains disulfide bonds, conferring versatility for a wide spectrum of experimental setups. We applied SLAPSHOT followed by quantitative mass spectrometry-based proteomics analysis to examine the immediate and extensive cell surface expansion and ensuing restorative membrane shedding upon the activation of Scott syndrome-linked TMEM16F, a ubiquitously expressed calcium-dependent phospholipid scramblase and ion channel. Time-course data ranging from one to thirty minutes of calcium stimulation using wild-type and TMEM16F deficient cells revealed intricate co-regulation of known protein families, including those in the integrin and ICAM families. Crucially, we identified proteins that are known to reside in intracellular organelles, including ER, as occupants of the freshly deposited membrane, and mitovesicles as an abundant component and contributor to the extracellularly exposed proteome. Our study not only provides the first accounts of the immediate consequences of calcium signaling on the extracellularly exposed proteome, but also presents a blueprint for the application of SLAPSHOT as a general approach for monitoring extracellularly exposed protein dynamics. ### Competing Interest Statement The authors have declared no competing interest.

Fungi in the basidiomycete genus Malassezia are the most prevalent eukaryotic microbes resident on the skin of human and other warm-blooded animals and have been implicated in skin diseases and systemic disorders. Analysis of Malassezia genomes revealed that key adaptations to the skin microenvironment have a direct genomic basis, and the identification of mating and meiotic genes suggests a capacity to reproduce sexually, even though no sexual cycle has been as yet observed. In contrast to other bipolar or tetrapolar basidiomycetes that have either two linked mating-type-determining ( MAT ) loci or two MAT loci on separate chromosomes, in Malassezia species studied thus far the two MAT loci are arranged in a pseudobipolar configuration (linked on the same chromosome but capable of recombining). By incorporating newly-generated chromosome-level genome assemblies, and an improved Malassezia phylogeny, we infer that the pseudobipolar arrangement was the ancestral state of this group and revealed six independent transitions to tetrapolarity, seemingly driven by centromere fission or translocations in centromere-flanking regions. Additionally, in an approach to uncover a sexual cycle, Malassezia furfur strains were engineered to express different MAT alleles in the same cell. The resulting strains produce hyphae reminiscent of early steps in sexual development and display upregulation of genes associated with sexual development as well as others encoding lipases and a protease potentially relevant for pathogenesis of the fungus. Our study reveals a previously unseen genomic relocation of mating-type loci in fungi and provides insight towards the discovery of a sexual cycle in Malassezia , with possible implications for pathogenicity. ### Competing Interest Statement The authors have declared no competing interest.

Volvocine green algae are a model for understanding the evolution of mating types and sexes. They are facultatively sexual, with gametic differentiation occurring in response to nitrogen starvation (-N) in most genera, and to sex inducer hormone (SI) in Volvox. The conserved RWP-RK family transcription factor (TF) MID is encoded by the minus mating type (MT) locus or male sex-determining region (SDR) of heterothallic volvocine species and dominantly determines minus or male gametic differentiation. However, the factor(s) responsible for establishing the default plus or female differentiation programs have remained elusive. We performed a phylo-transcriptomic screen for autosomal RWP-RK TFs induced during gametogenesis in unicellular isogamous Chlamydomonas reinhardtii (Chlamydomonas) and in multicellular oogamous Volvox carteri (Volvox), and identified a single conserved ortho-group we named Volvocine Sex Regulator (VSR1). Chlamydomonas vsr1 mutants of either mating type failed to mate and could not induce expression of key mating-type-specific genes. Similarly, Volvox vsr1 mutants in either sex could initiate sexual embryogenesis, but the presumptive eggs or androgonidia (sperm packet precursors) were infertile and unable to express key sex-specific genes. Yeast two-hybrid assays identified a conserved domain in VSR1 that interacts with itself or with the conserved N terminal domain of MID. In vivo co-immunoprecipitation experiments demonstrated association of VSR1 and MID in both Chlamydomonas and Volvox. These data support a new model for volvocine sexual differentiation where VSR1 homodimers activate expression of plus /female gamete-specific-genes, but when MID is present MID-VSR1 heterodimers are preferentially formed and activate minus /male gamete-specific-genes. ### Competing Interest Statement The authors have declared no competing interest.

Many phage engineering technologies, mainly based on the CRISPR-Cas system, have been recently developed. Here we present a method to genetically engineer the Escherichia coli phages T5, T7, and P1 by adapting a technology, called pORTMAGE, developed for engineering bacterial genomes. The technology comprises of E. coli harboring a plasmid encoding a potent recombinase and a gene transiently silencing a repair system. Oligonucleotides with the desired phage mutation are electroporated into E. coli followed by infection of the target bacteriophage. The high efficiency of this technology, ranging from 1-14% of desired recombinants, allows low-throughput screening for the desired mutant. We demonstrated the use of this technology for single-base substitutions, for deletions of 50 bases, for insertions of 20 bases, and for three different phages. The technology may be adjusted for use across many bacterial and phage strains. ### Competing Interest Statement The authors have declared no competing interest.

When replication forks encounter DNA lesions that cause polymerase stalling a checkpoint pathway is activated. The ATR-dependent intra-S checkpoint pathway mediates detection and processing of sites of replication fork stalling to maintain genomic integrity. Several factors involved in the global checkpoint pathway have been identified, but the response to a single replication fork barrier (RFB) is poorly understood. We utilized the E.coli -based Tus- Ter system in human MCF7 cells and showed that the Tus protein binding to TerB sequences creates an efficient site-specific RFB. The single fork RFB was sufficient to activate a local, but not global, ATR-dependent checkpoint response that leads to phosphorylation and accumulation of DNA damage sensor protein γH2AX, confined locally to within a kilobase of the site of stalling. These data support a model of local management of fork stalling, which allows global replication at sites other than the RFB to continue to progress without delay. ### Competing Interest Statement The authors have declared no competing interest.

The formation of RAD51/DMC1 filaments on single-stranded (ss)DNAs, which is essential for homology search and strand exchange in DNA double-strand break (DSB) repair and for the protection of stalled DNA replication forks, is tightly regulated in time and space. FIGNL1 AAA+++ ATPase plays positive and negative roles in the RAD51-mediated recombination in human cells. However, the role of FIGNL1 in gametogenesis remains unsolved. Here, we characterized a germ-line-specific conditional knockout (cKO) mouse of FIGNL1. The Fignl1 cKO male mice showed defective chromosome synapsis and impaired meiotic DSB repair with the accumulation of RAD51/DMC1 on chromosomes in mid-meiotic prophase I, supporting a role of FIGNL1 in a post-assembly stage of RAD51/DMC1 filaments. Fignl1 cKO spermatocytes accumulate RAD51 and DMC1 ensembles on chromosomes not only in early meiotic prophase I but also in meiotic S-phase. These RAD51/DMC1 assemblies are independent of meiotic DSB formation. Finally, we showed that purified FIGNL1 dismantles RAD51 filament on double-stranded (ds)DNA as well as ssDNA. These results suggest a critical role of FIGNL1 to limit the uncontrolled assembly of RAD51 and DMC1 on native dsDNAs during the meiotic S-phase and meiotic prophase I. ### Competing Interest Statement The authors have declared no competing interest.

The miR-15/16 family is a highly expressed group of tumor suppressor miRNAs that target a large network of genes in T cells to restrict their cell cycle, memory formation and survival. Upon T cell activation, miR-15/16 are downregulated, allowing rapid expansion of differentiated effector T cells to mediate a sustained immune response. Here, using conditional deletion of miR-15/16 in immunosuppressive regulatory T cells (Tregs) that express FOXP3, we identify new functions of the miR-15/16 family in T cell immunity. miR-15/16 are indispensable to maintain peripheral tolerance by securing efficient suppression by a limited number of Tregs. miR-15/16-deficiency alters Treg expression of critical functional proteins including FOXP3, IL2Rα/CD25, CTLA4, PD-1 and IL7Rα/CD127, and results in accumulation of functionally impaired FOXP3loCD25loCD127hi Tregs. Excessive proliferation in the absence of miR-15/16 inhibition of cell cycle programs shifts Treg diversity and produces an effector Treg phenotype characterized by low expression of TCF1, CD25 and CD62L, and high expression of CD44. These Tregs fail to control immune activation of CD4+ effector T cells, leading to spontaneous multi-organ inflammation and increased allergic airway inflammation in a mouse model of asthma. Together, our results demonstrate that miR-15/16 expression in Tregs is essential to maintain immune tolerance. ### Competing Interest Statement The authors have declared no competing interest.

Salmonella infection remains a major global health problem and worsened by lack of appropriate diagnostic tools to aid early detection and teatment, particularly in low-income nations . Salmonella typhi is the most common causative agent of typhoid fever and the prevalence of this illness has been on the increase specifically in areas of poor personal hygiene and sanitation. This study was carried out to further improve the diagnosis of salmonella infection, through a mathematical regression model. An analysis was performed using the logistic regression approach and the predictability of the model was done by extracting fifteen (15) typhoid observations from the obtained samples; for the model to predict their status. The model was able to accurately predict 66.7% of the observations. This study showed an increased prevalence in typhoid fever including a significant correlation between typhoid fever and other parameters. The global burden of this illness can be minimized by proper vaccination, and prompt but appropriate diagnosis and treatment.Further studies and test-meaasures also needs to be carried out to improve diagnosis and treatment regimen. ### Competing Interest Statement The authors have declared no competing interest.

Infections caused by metallo-beta-lactamase-producing organisms (MBLs) are a global health threat. Our understanding of transmission dynamics and how MBLs establish endemicity remains limited. We analysed two decades of blaIMP-4 evolution in a hospital using sequence data from 270 clinical and environmental isolates (including 169 completed genomes) and identified extreme gene promiscuity across 7 Gram-negative genera, 68 bacterial strains and 7 distinct plasmid types. An initial multi-species outbreak of conserved IncC plasmids (95 genomes across 37 strains) allowed endemicity to be established through the ability of blaIMP-4 to disseminate in successful strain-genetic setting pairs we termed propagators, in particular Serratia marcescens and Enterobacter hormaechei. From this reservoir, blaIMP-4 persisted through diversification of genetic settings that resulted from transfer of blaIMP-4 plasmids between bacterial hosts and of the integron carrying blaIMP-4 between plasmids. Our findings provide a framework for understanding endemicity and spread of MBLs and may have broader applicability to other carbapenemase-producing organisms. ### Competing Interest Statement The authors have declared no competing interest.

Here we report the first successful cultivation of SAR202 bacteria, a superorder in the phylum Chloroflexota, which have long been at the top of "most wanted" lists of uncultivated microbial life. It has been proposed that ancient expansions of catabolic enzyme paralogs in SAR202 broadened the spectrum of organic compounds they could oxidize, leading to transformations of the Earth's carbon cycle. We cultured the cells from surface seawater using dilution-to-extinction culturing. Their growth was very slow (0.18-0.24 day-1) and was inhibited by exposure to light. The genomes, of ca. 3.08 Mbp, encoded archaella, archaeal motility structures, and multiple sets of paralogs, including 80 genes in enolase superfamily and 44 genes in NAD(P)-dependent dehydrogenase family. We propose that these paralogs participate in multiple parallel pathways of non-phosphorylative sugar and sugar acid catabolism, and demonstrate that, as predicted by this scheme, the sugars L-fucose and L-rhamnose and their lactone and acid forms are utilized by these cells. ### Competing Interest Statement The authors have declared no competing interest.

The growth and division of mycobacteria, which include several clinically relevant pathogens, deviate significantly from that of canonical bacterial models. Despite their Gram-positive ancestry, mycobacteria synthesize and elongate a diderm envelope asymmetrically from the poles, with the old pole elongating more robustly than the new pole. In addition to being structurally distinct, the molecular components of the mycobacterial envelope are also evolutionarily unique, including the phosphatidylinositol-anchored lipoglycans lipomannan (LM) and lipoarabinomannan (LAM). LM and LAM modulate host immunity during infection, but their role outside of intracellular survival remains poorly understood, despite their widespread conservation among non-pathogenic and opportunistically pathogenic mycobacteria. Previously, Mycobacterium smegmatis and Mycobacterium tuberculosis mutants producing structurally altered LM and LAM were shown to grow slowly under certain conditions and to be more sensitive to antibiotics, suggesting that mycobacterial lipoglycans may support cellular integrity or growth. To test this, we constructed multiple biosynthetic lipoglycan mutants of M. smegmatis and determined the effect of each mutation on cell wall biosynthesis, envelope integrity, and division. We found that mutants deficient in LAM, but not LM, fail to maintain cell wall integrity in a medium-dependent manner, with envelope deformations specifically associated with septa and new poles. Conversely, a mutant producing abnormally large LAM formed multiseptated cells in way distinct from that observed in a septal hydrolase mutant. These results show that LAM plays critical and distinct roles at subcellular locations associated with division in mycobacteria, including maintenance of local cell envelope integrity and septal placement. ### Competing Interest Statement The authors have declared no competing interest.

Type VI secretion systems (T6SS) are common bacterial contractile injection systems that inject toxic "effector" proteins into neighboring cells. We bioinformatically investigated T6SS core proteins in 11,832 genomes of Gram negative bacteria. Comparison of T6SS core proteins that are covalently attached to toxic T6SS effector proteins (T6Es) versus those that are not revealed differences in phylogenetic distribution, physical properties, and genomic position. Using the data generated from our bioinformatic analysis, we developed a new genomic- and Alphafold2-based pipeline for discovery of putative T6Es. We experimentally validated the toxic and immunity activities of four putative antibacterial T6SS effector proteins and four cognate immunity genes from diverse species, respectively. We used Foldseek to predict possible mechanisms of action of the putative T6Es, which was much more effective than sequence-based methods. Evidence of the possible mechanisms of action of the putative T6Es was explored through fluorescence microscopy, where we observed cell wall-targeting, DNA degradation, and cell filamentation. This study shows how combining genomic data mining with new structure-based bioinformatic tools can facilitate identification of novel antibacterial toxins. ### Competing Interest Statement The authors have declared no competing interest.

Biogeochemical sulfur cycling in sulfidic karst systems is largely driven by abiotic and biological sulfide oxidation, but the fate of elemental sulfur (S) that accumulates in these systems is not well understood. The Frasassi Cave system (Italy) is intersected by a sulfidic aquifer that mixes with small quantities of oxygen-rich meteoric water, creating Proterozoic-like conditions and supporting a prolific ecosystem driven by sulfur-based chemolithoautotrophy. To better understand the cycling of S in this environment, we examined the geochemistry and microbiology of sediments underlying widespread sulfide-oxidizing mats dominated by Beggiatoa . Sediment populations were dominated by uncultivated relatives of sulfur cycling chemolithoautotrophs related to Sulfurovum , Halothiobacillus , Thiofaba , Thiovirga , Thiobacillus , and Desulfocapsa , as well as diverse uncultivated anaerobic heterotrophs affiliated with Bacteroidota, Anaerolineaceae, Lentimicrobiaceae, and Prolixibacteraceae. Desulfocapsa and Sulfurovum populations accounted for 12-26% of sediment 16S rRNA amplicon sequences and were closely related to isolates which carry out autotrophic S disproportionation in pure culture. Gibbs energy (Δ G r ) calculations revealed that S disproportionation under in situ conditions is energy yielding. Microsensor profiles through the mat-sediment interface showed that Beggiatoa mats consume dissolved sulfide and oxygen, but a net increase in acidity was only observed in the sediments below. Together, these findings suggest that disproportionation is an important sink for S generated by microbial sulfide oxidation in this oxygen-limited system and may contribute to the weathering of carbonate rocks and sediments in sulfur-rich environments. ### Competing Interest Statement The authors have declared no competing interest.

Organoids, which can reproduce the complex tissue structures found in embryos, are revolutionizing basic research and regenerative medicine. In order to use organoids for research and medicine, it is necessary to assess the composition and arrangement of cell types within the organoid, i.e., spatial gene expression. However, current methods are invasive and require gene editing and immunostaining. In this study, we developed a non-invasive estimation method of spatial gene expression patterns using machine learning. A deep learning model was trained with an encoder-decoder architecture on a dataset of retinal organoids derived from mouse embryonic stem cells. This method successfully estimated spatially plausible fluorescent patterns with appropriate intensities, enabling the non-invasive, quantitative estimation of spatial gene expression patterns within each tissue. Thus, this method could lead to new avenues for evaluating spatial gene expression patterns across a wide range of biology and medicine fields. ### Competing Interest Statement The authors have declared no competing interest.

Proper cellular proteostasis, essential for viability, requires a network of chaperones and cochaperones. ATP-dependent chaperonin TRiC/CCT partners with cochaperones prefoldin (PFD) and phosducin-like proteins (PhLPs) to facilitate the folding of essential eukaryotic proteins. Using cryoEM and biochemical analyses, we determine the ATP-driven cycle of TRiC-PFD-PhLP2A interaction. In the open TRiC state, PhLP2A binds to the chamber's equator while its N-terminal H3-domain binds to the apical domains of CCT3/4, thereby displacing PFD from TRiC. ATP-induced TRiC closure rearranges the contacts of PhLP2A domains within the closed chamber. In the presence of substrate, actin and PhLP2A segregate into opposing chambers, each binding to the positively charged inner surfaces formed by CCT1/3/6/8. Notably, actin induces a conformational change in PhLP2A, causing its N-terminal helices to extend across the inter-ring interface to directly contact a hydrophobic groove in actin. Our findings reveal an ATP-driven PhLP2A structural rearrangement cycle within the TRiC chamber to facilitate folding. ### Competing Interest Statement The authors have declared no competing interest.

Cell proliferation is a fundamental process underlying embryogenesis, homeostasis, wound healing, and cancer. The process involves multiple events during each cell cycle, such as cell growth, contractile ring formation, and division to daughter cells, which affect the surrounding cell population geometrically and mechanically. However, existing methods do not comprehensively describe the dynamics of multicellular structures involving cell proliferation at a subcellular resolution. In this study, we present a novel model for proliferative multicellular dynamics at the subcellular level by building upon the nonconservative fluid membrane (NCF) model that we developed in earlier research. The NCF model utilizes a dynamically-rearranging closed triangular mesh to depict the shape of each cell, enabling us to analyze cell dynamics over extended periods beyond each cell cycle, during which cell surface components undergo dynamic turnover. The proposed model represents the process of cell proliferation by incorporating cell volume growth and contractile ring formation through an energy function and topologically dividing each cell at the cleavage furrow formed by the ring. Numerical simulations demonstrated that the model recapitulated the process of cell proliferation at subcellular resolution, including cell volume growth, cleavage furrow formation, and division to daughter cells. Further analyses suggested that the orientation of actomyosin stress in the contractile ring plays a crucial role in the cleavage furrow formation, i.e., circumferential orientation can form a cleavage furrow but isotropic orientation cannot. Furthermore, the model replicated tissue-scale multicellular dynamics, where the successive proliferation of adhesive cells led to the formation of a cell sheet and stratification on the substrate. Overall, the proposed model provides a basis for analyzing proliferative multicellular dynamics at subcellular resolution. ### Competing Interest Statement The authors have declared no competing interest.

Neuronal mitochondria play important roles beyond ATP generation, including Ca2+ uptake, and therefore have instructive roles in synaptic function and neuronal response properties. Mitochondrial morphology differs significantly in the axon and dendrites of a given neuronal subtype, but in CA1 pyramidal neurons (PNs) of the hippocampus, mitochondria within the dendritic arbor also display a remarkable degree of subcellular, layer-specific compartmentalization. In the dendrites of these neurons, mitochondria morphology ranges from highly fused and elongated in the apical tuft, to more fragmented in the apical oblique and basal dendritic compartments, and thus occupy a smaller fraction of dendritic volume than in the apical tuft. However, the molecular mechanisms underlying this striking degree of subcellular compartmentalization of mitochondria morphology are unknown, precluding the assessment of its impact on neuronal function. Here, we demonstrate that this compartment-specific morphology of dendritic mitochondria requires activity-dependent, Camkk2-dependent activation of AMPK and its ability to phosphorylate two direct effectors: the pro-fission Drp1 receptor Mff and the recently identified anti-fusion, Opa1-inhibiting protein, Mtfr1l. Our study uncovers a new activity-dependent molecular mechanism underlying the extreme subcellular compartmentalization of mitochondrial morphology in dendrites of neurons in vivo through spatially precise regulation of mitochondria fission/fusion balance. ### Competing Interest Statement The authors have declared no competing interest.

Cyclin-dependent kinase (CDK) plays an essential role in determining the temporal ordering of the cell cycle phases. However, despite significant progress in studying regulators of CDK, it remains elusive how they coordinately affect CDK activity at the single-cell level and how CDK controls the temporal order of cell cycle events. This could be due to the lack of tools to monitor CDK activity in living cells. Here, we elucidate the dynamics of CDK activity in fission yeast and mammalian cells by using a newly developed CDK activity biosensor, Eevee-spCDK, based on Forster Resonance Energy Transfer (FRET). Taking advantage of this system, we unravel the profile of CDK activity in vegetatively growing S. pombe cells. Thus, we detect a transient increase in S phase followed by a gradual increment during G2 phase. CDK activity then reaches its maximum in early M phase and rapidly decreases at mitotic exit. During G2 phase, CDK activity exhibits a biphasic pattern, i.e., an early slow increase and a late fast rise prior to the G2/M phase transition, as predicted from mathematical studies. Remarkably, although CDK activity does not necessarily correlate with cyclin levels, we find that it converges to the same level around mitotic onset in several mutant backgrounds, including pom1Δ cells and wee1 or cdc25 overexpressing cells. These data provide the first direct evidence that cells enter M phase when CDK activity reaches a high threshold, consistent with the quantitative model of cell cycle progression in fission yeast. ### Competing Interest Statement The authors have declared no competing interest.

Cytokines of the interleukin (IL)-1 family are widely expressed in epithelial surfaces, including the epidermis, where they play a key role in the maintenance of barrier integrity and host defense. A recent report associated the IL-1 family member IL-33 with stress granules (SGs) in epithelial cells. Formation of SGs is promoted by the aggregation of proteins harboring low complexity regions (LCRs). In this study, using computational analyses, we predicted the presence of LCRs in six of the eleven IL-1 family members. Among these, IL-38 contained a long LCR and localized to Ras GTPase-activating protein binding protein 1 (G3BP1) positive SGs, as well as to G3BP1 negative intracellular protein condensates in keratinocytes exposed to oxidative stress (OS). In addition, we identified two highly aggregation-prone amyloid core (AC) motifs in the IL-38 LCR and detected the formation of amyloid IL-38 aggregates in response to OS in cells and in vitro. Disulfide bond mapping, in silico modelling and the analysis of specific cysteine mutants supported a model in which specific oxidation-sensitive cysteines act as redox switches to modify the conformation of IL-38 and thus the surface exposure of its ACs, shuttling it from a soluble state into biomolecular condensates. Finally, the presence of IL-38 granules in human epidermal layers highly exposed to environmental OS suggests that oxidation-induced formation of amyloid aggregates, as a previously unrecognized intrinsic biological property of IL-38, may be physiologically relevant at this epithelial barrier. ### Competing Interest Statement The authors have declared no competing interest.