Resistance of fungi to predation is thought to be mediated by toxic metabolites and proteins. Many of these fungal defense effectors are highly abundant in the fruiting body and not produced in the vegetative mycelium. The defense function of fruiting body-specific proteins, however, including cytoplasmically localized lectins and antinutritional proteins such as biotin-binding proteins, is mainly based on toxicity assays using bacteria as a heterologous expression system, with bacterivorous/omnivorous model organisms as predators. Here, we present an ecologically more relevant experimental setup to assess the toxicity of potential fungal defense proteins towards the fungivorous, stylet-feeding nematodes Aphelenchus avenae and Bursaphelenchus okinawaensis As a heterologous expression host, we exploited the filamentous fungus Ashbya gossypii Using this new system, we assessed the toxicity of six previously characterized, cytoplasmically localized, potential defense proteins from fruiting bodies of different fungal phyla against the two fungivorous nematodes. We found that all of the tested proteins were toxic against both nematodes, albeit to various degrees. The toxicity of these proteins against both fungivorous and bacterivorous nematodes suggests that their targets have been conserved between the different feeding groups of nematodes and that bacterivorous nematodes are valid model organisms to assess the nematotoxicity of potential fungal defense proteins.IMPORTANCE Our results support the hypothesis that cytoplasmic proteins abundant in fungal fruiting bodies are involved in fungal resistance against predation. The toxicity of these proteins toward stylet-feeding nematodes, which are also capable of feeding on plants, and the abundance of these proteins in edible mushrooms, may open possible avenues for biological crop protection against parasitic nematodes, e.g., by expression of these proteins in crops.

Receptors on the cell surfaces of bacterial hosts are essential during the infection cycle of bacteriophages. To date, the phage receptors of the industrial relevant dairy starter bacterium Streptococcus thermophilus remain elusive. Thus, we set out to identify cell surface structures that are involved in host recognition by dairy streptococcal phages. Five industrial S. thermophilus strains sensitive to different phages (pac type, cos type, and the new type 987), were selected to generate spontaneous bacteriophage-insensitive mutants (BIMs). Of these, approximately 50% were deselected as clustered regularly interspaced short palindromic repeat (CRISPR) mutants, while the other pool was further characterized to identify receptor mutants. On the basis of genome sequencing data, phage resistance in putative receptor mutants was attributed to nucleotide changes in genes encoding glycan biosynthetic pathways. Superresolution structured illumination microscopy was used to visualize the interactions between S. thermophilus and its phages. The phages were either regularly distributed along the cells or located at division sites of the cells. The cell wall structures mediating the latter type of phage adherence were further analyzed via phenotypic and biochemical assays. Altogether, our data suggested that phage adsorption to S. thermophilus is mediated by glycans associated with the bacterial cell surface. Specifically, the pac-type phage CHPC951 adsorbed to polysaccharides anchored to peptidoglycan, while the 987-type phage CHPC926 recognized exocellular polysaccharides associated with the cell surface.IMPORTANCEStreptococcus thermophilus is widely used in starter cultures for cheese and yoghurt production. During dairy fermentations, infections of bacteria with bacteriophages result in acidification failures and a lower quality of the final products. An understanding of the molecular factors involved in phage-host interactions, in particular, the phage receptors in dairy bacteria, is a crucial step for developing better strategies to prevent phage infections in dairy plants.

FRET-based biosensor experiments in adult cardiomyocytes are a powerful way of dissecting the spatiotemporal dynamics of the complicated signaling networks that regulate cardiac health and disease. However, although much information has been gleaned from FRET studies on cardiomyocytes from larger species, experiments on adult cardiomyocytes from mice have been difficult at best. Thus the large variety of genetic mouse models cannot be easily used for this type of study. Here we develop cell culture conditions for adult mouse cardiomyocytes that permit robust expression of adenoviral FRET biosensors and reproducible FRET experimentation. We find that addition of 6.25 µM blebbistatin or 20 µM (S)-nitro-blebbistatin to a minimal essential medium containing 10 mM HEPES and 0.2% BSA maintains morphology of cardiomyocytes from physiological, pathological, and transgenic mouse models for up to 50 h after adenoviral infection. This provides a 10-15-h time window to perform reproducible FRET readings using a variety of CFP/YFP sensors between 30 and 50 h postinfection. The culture is applicable to cardiomyocytes isolated from transgenic mouse models as well as models with cardiac diseases. Therefore, this study helps scientists to disentangle complicated signaling networks important in health and disease of cardiomyocytes.

Hebbian synaptic plasticity at hippocampal Schaffer collateral synapses is tightly regulated by postsynaptic small conductance (SK) channels that restrict NMDA receptor activity. SK channels are themselves modulated by G-protein-coupled signaling pathways, but it is not clear under what conditions these are activated to enable synaptic plasticity. Here, we show that muscarinic M1 receptor (M1R) and type 1 metabotropic glutamate receptor (mGluR1) signaling pathways, which are known to inhibit SK channels and thereby disinhibit NMDA receptors, converge to facilitate spine calcium transients during the induction of long-term potentiation (LTP) at hippocampal Schaffer collateral synapses onto CA1 pyramidal neurons of male rats. Furthermore, mGluR1 activation is required for LTP induced by reactivated place-cell firing patterns that occur in sharp-wave ripple events during rest or sleep. In contrast, M1R activation is required for LTP induced by place-cell firing patterns during exploration. Thus, we describe a common mechanism that enables synaptic plasticity during both encoding and consolidation of memories within hippocampal circuits.SIGNIFICANCE STATEMENT Memory ensembles in the hippocampus are formed during active exploration and consolidated during rest or sleep. These two distinct phases each require strengthening of synaptic connections by long-term potentiation (LTP). The neuronal activity patterns in each phase are very different, which makes it hard to map generalized rules for LTP induction onto both formation and consolidation phases. In this study, we show that inhibition of postsynaptic SK channels is a common necessary feature of LTP induction and that SK channel inhibition is achieved by separate but convergent metabotropic signaling pathways. Thus, we reveal a common mechanism for enabling LTP under distinct behavioral conditions.

Metazoan Hedgehog (Hh) morphogens are essential regulators of growth and patterning at significant distances from their source, despite being produced as N-terminally palmitoylated and C-terminally cholesteroylated proteins, which firmly tethers them to the outer plasma membrane leaflet of producing cells and limits their spread. One mechanism to overcome this limitation is proteolytic processing of both lipidated terminal peptides, called shedding, but molecular target site requirements for effective Hh shedding remained undefined. In this work, by using Drosophila melanogaster as a model, we show that mutagenesis of the N-terminal Cardin-Weintraub (CW) motif inactivates recombinant Hh proteins to variable degrees and, if overexpressed in the same compartment, converts them into suppressors of endogenous Hh function. In vivo, additional removal of N-palmitate membrane anchors largely restored endogenous Hh function, supporting the hypothesis that proteolytic CW processing controls Hh solubilization. Importantly, we also observed that CW repositioning impairs anterior/posterior compartmental boundary maintenance in the third instar wing disc. This demonstrates that Hh shedding not only controls the differentiation of anterior cells, but also maintains the sharp physical segregation between these receiving cells and posterior Hh-producing cells.

The ability to understand intentions based on another's movements is crucial for human interaction. This ability has been ascribed to the so-called motor chaining mechanism: anytime a motor chain is activated (e.g., grasp-to-drink), the observer attributes to the agent the corresponding intention (i.e., to drink) from the first motor act (i.e., the grasp). However, the mechanisms by which a specific chain is selected in the observer remain poorly understood. In the current study, we investigate the possibility that in the absence of discriminative contextual cues, slight kinematic variations in the observed grasp inform mapping to the most probable chain. Chaining of motor acts predicts that, in a sequential grasping task (e.g., grasp-to-drink), electromyographic (EMG) components that are required for the final act [e.g., the mouth-opening mylohyoid (MH) muscle] show anticipatory activation. To test this prediction, we used MH EMG, transcranial magnetic stimulation (TMS; MH motor-evoked potentials), and predictive models of movement kinematics to measure the level and timing of MH activation during the execution (Experiment 1) and the observation (Experiment 2) of reach-to-grasp actions. We found that MH-related corticobulbar excitability during grasping observation varied as a function of the goal (to drink or to pour) and the kinematics of the observed grasp. These results show that subtle changes in movement kinematics drive the selection of the most probable motor chain, allowing the observer to link an observed act to the agent's intention.

Surface protein dynamics dictate synaptic connectivity and function in neuronal circuits. ASTN2, a gene disrupted by copy number variations (CNVs) in neurodevelopmental disorders, including autism spectrum, was previously shown to regulate the surface expression of ASTN1 in glial-guided neuronal migration. Here, we demonstrate that ASTN2 binds to and regulates the surface expression of multiple synaptic proteins in postmigratory neurons by endocytosis, resulting in modulation of synaptic activity. In cerebellar Purkinje cells (PCs), by immunogold electron microscopy, ASTN2 localizes primarily to endocytic and autophagocytic vesicles in the cell soma and in subsets of dendritic spines. Overexpression of ASTN2 in PCs, but not of ASTN2 lacking the FNIII domain, recurrently disrupted by CNVs in patients, including in a family presented here, increases inhibitory and excitatory postsynaptic activity and reduces levels of ASTN2 binding partners. Our data suggest a fundamental role for ASTN2 in dynamic regulation of surface proteins by endocytic trafficking and protein degradation.

Epidermal growth factor receptor (EGFR) activation by growth factors (GFs) relies on dimerization and allosteric activation of its intrinsic kinase activity, resulting in trans-phosphorylation of tyrosines on its C-terminal tail. While structural and biochemical studies identified this EGF-induced allosteric activation, imaging collective EGFR activation in cells and molecular dynamics simulations pointed at additional catalytic EGFR activation mechanisms. To gain more insight into EGFR activation mechanisms in living cells, we develop a Förster resonance energy transfer (FRET)-based conformational EGFR indicator (CONEGI) using genetic code expansion that reports on conformational transitions in the EGFR activation loop. Comparing conformational transitions, self-association and auto-phosphorylation of CONEGI and its Y845F mutant reveals that Y845 phosphorylation induces a catalytically active conformation in EGFR monomers. This conformational transition depends on EGFR kinase activity and auto-phosphorylation on its C-terminal tail, generating a looped causality that leads to autocatalytic amplification of EGFR phosphorylation at low EGF dose.

To explore new protective measure against visceral leishmaniasis, reverse vaccinology approach was employed to identify key immunogenic regions which can mediate long-term immunity. In-depth computational analysis revealed nine promiscuous epitopes which can possibly be presented by 46 human leukocyte antigen, thereby broadening the worldwide population up to 94.16%. This is of reasonable significance that most of the epitopes shared 100% sequence homology with other Leishmania species and could evoke a common pattern of protective immune response. Transporter associated with antigen processing binding affinity, molecular docking approach followed by dynamics simulation and human leukocyte antigen stabilization assay suggested that the best five optimal set of epitopes bind in between α1 and α2 binding groove with sufficient affinity and stability which allows the translocation of intact epitope to the cell surface. Fascinatingly, the human leukocyte antigen stabilization assay exhibited a modest correlation with the positive immunogenicity score predicted by class I pMHC immunogenicity predictor. A support for this notion came from ELISA and FACS analysis where the epitopes as a cocktail induced CD8+ IFN-γ and Granzyme B levels significantly in treated visceral leishmaniasis subject which suggests the immunogenic ability of the selected epitopes.

The purpose of this study is to explore Dvl3 variants and their interaction with negative life events on MDD susceptibility in a Chinese Han population. Additionally, we also attempted to identify whether there is an association between Dvl3 variants and pro-inflammatory cytokines. A total of 1102 participants, consisting of 550 patients with MDD and 552 healthy subjects, were recruited for genotyping by TaqMan allelic discrimination assay. Pro-inflammatory cytokine mRNA levels in peripheral blood were measured by QPCR. After the assessment of negative life events by the Life Events Scale, the Dvl3 gene-environment interaction (G × E) and risk factors were evaluated using generalized multifactor dimensionality reduction method (GMDR) and logistic regression analysis, respectively. This study is the first to reveal the interaction between Dvl3 allelic variations and negative life events as well as pro-inflammatory cytokines on MDD susceptibility in a Chinese Han population.

Tuberous Sclerosis Complex (TSC), a rare genetic disorder with mechanistic target of rapamycin complex 1 (mTORC1) hyperactivation, is characterized by multi-organ hamartomatous benign tumors including brain, skin, kidney, and lung (Lymphangioleiomyomatosis). mTORC1 hyperactivation drives metabolic reprogramming including glucose and glutamine utilization, protein, nucleic acid and lipid synthesis. To investigate the mechanisms of exogenous nutrients uptake in Tsc2-deficient cells, we measured dextran uptake, a polysaccharide internalized via macropinocytosis. Tsc2-deficient cells showed a striking increase in dextran uptake (3-fold, p < 0.0001) relative to Tsc2-expressing cells, which was decreased (3-fold, p < 0.0001) with mTOR inhibitor, Torin1. Pharmacologic and genetic inhibition of the lipid kinase Vps34 markedly abrogated uptake of Dextran in Tsc2-deficient cells. Macropinocytosis was further increased in Tsc2-deficient cells that lack autophagic mechanisms, suggesting that autophagy inhibition leads to dependence on exogenous nutrient uptake in Tsc2-deficient cells. Treatment with a macropinocytosis inhibitor, ethylisopropylamiloride (EIPA), resulted in selective growth inhibition of Atg5-deficient, Tsc2-deficient cells (50%, p < 0.0001). Genetic inhibition of autophagy (Atg5-/- MEFs) sensitized cells with Tsc2 downregulation to the Vps34 inhibitor, SAR405, resulting in growth inhibition (75%, p < 0.0001). Finally, genetic downregulation of Vps34 inhibited tumor growth and increased tumor latency in an in vivo xenograft model of TSC. Our findings show that macropinocytosis is upregulated with Tsc2-deficiency via a Vps34-dependent mechanism to support their anabolic state. The dependence of Tsc2-deficient cells on exogenous nutrients may provide novel approaches for the treatment of TSC.

Cochlear implantation, a surgical method to bypass cochlear hair cells and directly stimulate the spiral ganglion, is the standard treatment for severe-to-profound hearing loss. Changes in cochlear implant electrode array design and surgical approach now allow for preservation of acoustic hearing in the implanted ear. Electrocochleography (ECochG) was performed in eight hearing preservation subjects to assess hair cell and neural function and elucidate underlying genetic hearing loss. Three subjects had pathogenic variants in TMPRSS3 and five had pathogenic variants in genes known to affect the cochlear sensory partition. The mechanism by which variants in TMPRSS3 cause genetic hearing loss is unknown. We used a 500-Hz tone burst to record ECochG responses from an intracochlear electrode. Responses consist of a cochlear microphonic (hair cell) and an auditory nerve neurophonic. Cochlear microphonics did not differ between groups. Auditory nerve neurophonics were smaller, on average, in subjects with TMPRSS3 deafness. Results of this proof-of-concept study provide evidence that pathogenic variants in TMPRSS3 may impact function of the spiral ganglion. While ECochG as a clinical and research tool has been around for decades, this study illustrates a new application of ECochG in the study of genetic hearing and deafness in vivo.

Holometaboly is a key evolutionary innovation that has facilitated the spectacular radiation of insects. Despite the undeniable advantage of complete metamorphosis, the female of some holometabolous species have lost the typical holometabolous development through neoteny. In Xenos vesparum Rossi (Strepsiptera: Stylopidae), a derived species of the holometabolous endoparasitic order Strepsiptera, neotenic females reach sexual maturity without the pupal and the imaginal stages, thus retaining their larval morphology (with the exception of the anterior part of the body or cephalothorax), while males undergo normal pupal-based metamorphosis. Expression of the "adult-specifier" E93 factor has been shown to be required for proper metamorphosis in holometabolous insects. Here, we investigated the involvement of E93 in female neoteny by cloning XvE93. Interestingly, while we detected a clear up-regulation of XvE93 expression in pupal and adult stages of males, persistent low levels of XvE93 were detected in X. vesparum females. However, a specific up-regulation of XvE93 was observed in the cephalothorax of late 4th female instar larva, which correlates with the occurrence of neotenic-specific features in the anterior part of the female body. Moreover, the same expression dynamic in the cephalothorax and abdomen was also observed for other two critical metamorphic regulators, the anti-metamorphic XvKr-h1 and the pupal specifier XvBr-C. The specific up-regulation of XvE93 and XvBr-C in the female cephalothorax seems to be the result of an increase in 20-hydroxyecdysone (20E) signaling in this region for we detected higher expression levels of the 20E-dependent nuclear receptors XvHR3 and XvE75 in the cephalothorax. Overall, our results detect a sex-specific expression pattern of critical metamorphic genes in X. vesparum, suggesting that neoteny in Strepsiptera results from the modification of the normal expression of E93, Br-C and Kr-h1 genes.

We present the first large-scale methylome-wide association studies (MWAS) for major depressive disorder (MDD) to identify sites of potential importance for MDD etiology. Using a sequencing-based approach that provides near-complete coverage of all 28 million common CpGs in the human genome, we assay methylation in MDD cases and controls from both blood (N = 1132) and postmortem brain tissues (N = 61 samples from Brodmann Area 10, BA10). The MWAS for blood identified several loci with P ranging from 1.91 × 10-8 to 4.39 × 10-8 and a resampling approach showed that the cumulative association was significant (P = 4.03 × 10-10) with the signal coming from the top 25,000 MWAS markers. Furthermore, a permutation-based analysis showed significant overlap (P = 5.4 × 10-3) between the MWAS findings in blood and brain (BA10). This overlap was significantly enriched for a number of features including being in eQTLs in blood and the frontal cortex, CpG islands and shores, and exons. The overlapping sites were also enriched for active chromatin states in brain including genic enhancers and active transcription start sites. Furthermore, three loci located in GABBR2, RUFY3, and in an intergenic region on chromosome 2 replicated with the same direction of effect in the second brain tissue (BA25, N = 60) from the same individuals and in two independent brain collections (BA10, N = 81 and 64). GABBR2 inhibits neuronal activity through G protein-coupled second-messenger systems and RUFY3 is implicated in the establishment of neuronal polarity and axon elongation. In conclusion, we identified and replicated methylated loci associated with MDD that are involved in biological functions of likely importance to MDD etiology.

The mitochondrial Ca2+ uniporter machinery is a multiprotein complex composed by the Ca2+ selective pore-forming subunit, the mitochondrial uniporter (MCU), and accessory proteins, including MICU1, MICU2 and EMRE. Their concerted action is required to fine-tune the uptake of Ca2+ into the mitochondrial matrix which both sustains cell bioenergetics and regulates the apoptotic response. To adequately fulfil such requirements and avoid impairment in mitochondrial Ca2+ handling, the intracellular turnover of all the MCU components must be tightly regulated. Here we show that the MCU complex regulator MICU1, but not MCU and MICU2, is rapidly and selectively degraded by the Ubiquitin Proteasome System (UPS). Moreover, we show that the multifunctional E3 ubiquitin ligase Parkin (PARK2), whose mutations cause autosomal recessive early-onset Parkinson's disease (PD), is a potential candidate involved in this process since its upregulation strongly decreases the basal level of MICU1. Parkin was found to interact with MICU1 and, interestingly, Parkin Ubl-domain, but not its E3-ubquitin ligase activity, is required for the degradation of MICU1, suggesting that in addition to the well documented role in the control of Parkin basal auto-inhibition, the Ubl-domain might exert important regulatory functions by acting as scaffold for the proteasome-mediated degradation of selected substrates under basal conditions, i.e. to guarantee their turnover. We have found that also MICU2 stability was affected upon Parkin overexpression, probably as a consequence of increased MICU1 degradation. Our findings support a model in which the PD-related E3 ubiquitin ligase Parkin directly participates in the selective regulation of the MCU complex regulator MICU1 and, indirectly, also of the MICU2 gatekeeper, thus indicating that Parkin loss of function could contribute to the impairment of the ability of mitochondria to handle Ca2+ and consequently to the pathogenesis of PD.

It is widely accepted that the periodic cycle of hair follicles is controlled by the biological clock, but the molecular regulatory mechanisms of the hair follicle cycle have not been thoroughly studied. The secondary hair follicle of the cashmere goat is characterized by seasonal periodic changes throughout life. In the hair follicle cycle, the initiation of hair follicles is of great significance for hair follicle regeneration. To provide a reference for hair follicle research, our study compared differences in mRNA expression and microRNA expression during the growth and repose stages of cashmere goat skin samples. Through microRNA and mRNA association analysis, we found microRNAs and target genes that play major regulatory roles in hair follicle initiation. We further constructed an mRNA-microRNA interaction network and found that hair follicle initiation and development were related to MiR-195 and the genes CHP1, SMAD2, FZD6 and SIAH1.

Caries is associated with shifts of microbiota in dental biofilms and primarily driven by frequent sucrose consumption. Data on environmentally induced in vivo microbiota shifts are scarce therefore we investigated the influence of frequent sucrose consumption on the oral biofilm. Splint systems containing enamel slabs were worn for 3 × 7 days with 7-day intervals to obtain oral biofilm samples. After a three-month dietary change of sucking 10 g of sucrose per day in addition to the regular diet, biofilm was obtained again at the end of the second phase. The microbiota was analysed using Illumina MiSeq amplicon sequencing (v1-v2 region). In addition, roughness of the enamel surface was measured with laser scanning microscopy. The sucrose phase resulted in significant differences in beta-diversity and significantly decreased species richness. It was marked by a significant increase in abundance of streptococci, specifically Streptococcus gordonii, Streptococcus parasanguinis and Streptococcus sanguinis. Enamel surface roughness began to increase, reflecting initial impairment of dental enamel surface. The results showed that frequent sucrose consumption provoked compositional changes in the microbiota, leading to an increase of non-mutans streptococci, hence supporting the extended ecological plaque hypothesis and emphasizing the synergy of multiple bacterial species in the development of caries.

A rare population of salivary gland cells isolated based on c-Kit immunoreactivity are thought to represent tissue stem cells since they exhibit the most robust proliferative and differentiation capacity ex vivo. Despite their high promise for cell-based therapies aimed at restoring salivary function, the precise location and in vivo function of c-Kit+ stem cells remain unclear. Here, by combining immunostaining with c-KitCreERT2-based genetic labeling and lineage tracing in the adult mouse salivary glands, we show that c-Kit is expressed in a relatively large and heterogeneous cell population that consists mostly of differentiated cells. Moreover, c-Kit does not mark ductal stem cells that are known to express cytokeratin K14. Tracking the fate of in vivo-labeled c-Kit+ or that of K14+ cells in spheroid cultures reveals a limited proliferative potential for c-Kit+ cells and identifies K14+ cells as the major source of salispheres in these cultures. Long-term in vivo lineage tracing studies indicate that although c-Kit marks at least two discrete ductal cell lineages, c-Kit+ cells do not contribute to the normal maintenance of any other cell lineages. Our results indicate that c-Kit is not a reliable marker for salivary gland stem cells, which has important implications for salivary gland regenerative therapies.

Self-organization is the generation of order out of local interactions. It is deeply connected to many fields of science from physics, chemistry to biology, all based on physical interactions. The emergence of collective animal behavior is the result of self-organization processes as well, though they involve abstract interactions arising from sensory inputs, information processing, storage, and feedback. Resulting collective behaviors are found, for example, in crowds of people, flocks of birds, and swarms of bacteria. Here we introduce interactions between active microparticles which are based on the information about other particle positions. A real-time feedback of multiple active particle positions is the information source for the propulsion direction of these particles. The emerging structures require continuous information flows. They reveal frustrated geometries due to confinement to two dimensions and internal dynamical degrees of freedom that are reminiscent of physically bound systems, though they exist only as nonequilibrium structures.

The intestinal barrier encompasses structural, permeability and immune aspects of the gut mucosa that, when disrupted, may contribute to chronic inflammation. Although gnotobiotic studies have demonstrated the effects of microbiota on mucosal and systemic immunity, as well as intestinal barrier architecture and innate immune characteristics, its impact on barrier function remains unclear. We compared germ-free and conventional mice, as well as mice colonized with human fecal microbiota that were followed for 21 days post-colonization. Colonic barrier structure was investigated by immunohistochemistry, molecular and electron microscopy techniques. Permeability was assessed in colon tissue by Ussing chambers, and by serum LPS and MDP detection using TLR4- and NOD2-NFκB reporter assays. Microbiota profile was determined by Illumina 16S rRNA gene sequencing. Low dose dextran sodium sulfate was administered to assess microbiota-induced barrier changes on resistance to colonic injury. Permeability to paracellular probes and mucus layer structure resembled that of conventional mice by day 7 post-colonization, coinciding with reduced claudin-1 expression and transient IL-18 production by intestinal epithelial cells. These post-colonization adaptations were associated with decreased systemic bacterial antigen exposure and reduced susceptibility to intestinal injury. In conclusion, commensal colonization promotes physiological barrier structural and functional adaptations that contribute to intestinal homeostasis.