Gene Expression Changes and Community Turnover Differentially Shape the Global Ocean Metatranscriptome.

Ocean microbial communities strongly influence the biogeochemistry, food webs, and climate of our planet. Despite recent advances in understanding their taxonomic and genomic compositions, little is known about how their transcriptomes vary globally. Here, we present a dataset of 187 metatranscriptomes and 370 metagenomes from 126 globally distributed sampling stations and establish a resource of 47 million genes to study community-level transcriptomes across depth layers from pole-to-pole. We examine gene expression changes and community turnover as the underlying mechanisms shaping community transcriptomes along these axes of environmental variation and show how their individual contributions differ for multiple biogeochemically relevant processes. Furthermore, we find the relative contribution of gene expression changes to be significantly lower in polar than in non-polar waters and hypothesize that in polar regions, alterations in community activity in response to ocean warming will be driven more strongly by changes in organismal composition than by gene regulatory mechanisms. VIDEO ABSTRACT.

Global Trends in Marine Plankton Diversity across Kingdoms of Life.

The ocean is home to myriad small planktonic organisms that underpin the functioning of marine ecosystems. However, their spatial patterns of diversity and the underlying drivers remain poorly known, precluding projections of their responses to global changes. Here we investigate the latitudinal gradients and global predictors of plankton diversity across archaea, bacteria, eukaryotes, and major virus clades using both molecular and imaging data from Tara Oceans. We show a decline of diversity for most planktonic groups toward the poles, mainly driven by decreasing ocean temperatures. Projections into the future suggest that severe warming of the surface ocean by the end of the 21st century could lead to tropicalization of the diversity of most planktonic groups in temperate and polar regions. These changes may have multiple consequences for marine ecosystem functioning and services and are expected to be particularly significant in key areas for carbon sequestration, fisheries, and marine conservation. VIDEO ABSTRACT.

Mirusviruses link herpesviruses to giant viruses.

DNA viruses have a major influence on the ecology and evolution of cellular organisms1-4, but their overall diversity and evolutionary trajectories remain elusive5. Here we carried out a phylogeny-guided genome-resolved metagenomic survey of the sunlit oceans and discovered plankton-infecting relatives of herpesviruses that form a putative new phylum dubbed Mirusviricota. The virion morphogenesis module of this large monophyletic clade is typical of viruses from the realm Duplodnaviria6, with multiple components strongly indicating a common ancestry with animal-infecting Herpesvirales. Yet, a substantial fraction of mirusvirus genes, including hallmark transcription machinery genes missing in herpesviruses, are closely related homologues of giant eukaryotic DNA viruses from another viral realm, Varidnaviria. These remarkable chimaeric attributes connecting Mirusviricota to herpesviruses and giant eukaryotic viruses are supported by more than 100 environmental mirusvirus genomes, including a near-complete contiguous genome of 432 kilobases. Moreover, mirusviruses are among the most abundant and active eukaryotic viruses characterized in the sunlit oceans, encoding a diverse array of functions used during the infection of microbial eukaryotes from pole to pole. The prevalence, functional activity, diversification and atypical chimaeric attributes of mirusviruses point to a lasting role of Mirusviricota in the ecology of marine ecosystems and in the evolution of eukaryotic DNA viruses.

Complex Genomes of Early Nucleocytoviruses Revealed by Ancient Origins of Viral Aminoacyl-tRNA Synthetases.

Aminoacyl-tRNA synthetases (aaRSs), also known as tRNA ligases, are essential enzymes in translation. Owing to their functional essentiality, these enzymes are conserved in all domains of life and used as informative markers to trace the evolutionary history of cellular organisms. Unlike cellular organisms, viruses generally lack aaRSs because of their obligate parasitic nature, but several large and giant DNA viruses in the phylum Nucleocytoviricota encode aaRSs in their genomes. The discovery of viral aaRSs led to the idea that the phylogenetic analysis of aaRSs can shed light on ancient viral evolution. However, conflicting results have been reported from previous phylogenetic studies: one posited that nucleocytoviruses recently acquired their aaRSs from their host eukaryotes, while another hypothesized that the viral aaRSs have ancient origins. Here, we investigated 4,168 nucleocytovirus genomes, including metagenome-assembled genomes (MAGs) derived from large-scale metagenomic studies. In total, we identified 780 viral aaRS sequences in 273 viral genomes. We generated and examined phylogenetic trees of these aaRSs with a large set of cellular sequences to trace evolutionary relationships between viral and cellular aaRSs. The analyses suggest that the origins of some viral aaRSs predate the last common eukaryotic ancestor. Inside viral aaRS clades, we identify intricate evolutionary trajectories of viral aaRSs with horizontal transfers, losses, and displacements. Overall, these results suggest that ancestral nucleocytoviruses already developed complex genomes with an expanded set of aaRSs in the proto-eukaryotic era.

Gene Transfer Among Viruses Substantially Contributes to Gene Gain of Giant Viruses.

The phylum Nucleocytoviricota comprises a diverse group of double-stranded DNA viruses that display a wide range of gene repertoires. Although these gene repertoires determine the characteristics of individual viruses, the evolutionary processes that have shaped the gene repertoires of extant viruses since their common ancestor are poorly characterized. In this study, we aimed to address this gap in knowledge by using amalgamated likelihood estimation, a probabilistic tree reconciliation method that infers evolutionary scenarios by distinguishing origination, gene duplications, virus-to-virus horizontal gene transfer (vHGT), and gene losses. We analyzed over 4,700 gene families from 195 genomes spanning all known viral orders. The evolutionary reconstruction suggests a history of extensive gene gains and losses during the evolution of these viruses, notably with vHGT contributing to gene gains at a comparable level to duplications and originations. The vHGT frequently occurred between phylogenetically closely related viruses, as well as between distantly related viruses with an overlapping host range. We observed a pattern of massive gene duplications that followed vHGTs for gene families that was potentially related to host range control and virus-host arms race. These results suggest that vHGT represents a previously overlooked, yet important, evolutionary force that integrates the evolutionary paths of multiple viruses and affects shaping of Nucleocytoviricota virus gene repertoires.

Giant Viruses Encode Actin-Related Proteins.

The emergence of the eukaryotic cytoskeleton is a critical yet puzzling step of eukaryogenesis. Actin and actin-related proteins (ARPs) are ubiquitous components of this cytoskeleton. The gene repertoire of the Last Eukaryotic Common Ancestor (LECA) would have therefore harbored both actin and various ARPs. Here, we report the presence and expression of actin-related genes in viral genomes (viractins) of some Imitervirales, a viral order encompassing the giant Mimiviridae. Phylogenetic analyses suggest an early recruitment of an actin-related gene by viruses from ancient protoeukaryotic hosts before the emergence of modern eukaryotes, possibly followed by a back transfer that gave rise to eukaryotic actins. This supports a coevolutionary scenario between pre-LECA lineages and their viruses, which could have contributed to the emergence of the modern eukaryotic cytoskeleton.

A rapid genome-wide analysis of isolated giant viruses using MinION sequencing.

Following the discovery of Acanthamoeba polyphaga mimivirus, diverse giant viruses have been isolated. However, only a small fraction of these isolates have been completely sequenced, limiting our understanding of the genomic diversity of giant viruses. MinION is a portable and low-cost long-read sequencer that can be readily used in a laboratory. Although MinION provides highly error-prone reads that require correction through additional short-read sequencing, recent studies assembled high-quality microbial genomes only using MinION sequencing. Here, we evaluated the accuracy of MinION-only genome assemblies for giant viruses by re-sequencing a prototype marseillevirus. Assembled genomes presented over 99.98% identity to the reference genome with a few gaps, demonstrating a high accuracy of the MinION-only assembly. As a proof of concept, we de novo assembled five newly isolated viruses. Average nucleotide identities to their closest known relatives suggest that the isolates represent new species of marseillevirus, pithovirus and mimivirus. The assembly of subsampled reads demonstrated that their taxonomy and genomic composition could be analysed at the 50× sequencing coverage. We also identified a pithovirus gene whose homologues were detected only in metagenome-derived relatives. Collectively, we propose that MinION-only assembly is an effective approach to rapidly perform a genome-wide analysis of isolated giant viruses.

MSNet-4mC: learning effective multi-scale representations for identifying DNA N4-methylcytosine sites.

MOTIVATION: N4-methylcytosine (4mC) is an essential kind of epigenetic modification that regulates a wide range of biological processes. However, experimental methods for detecting 4mC sites are time-consuming and labor-intensive. As an alternative, computational methods that are capable of automatically identifying 4mC with data analysis techniques become a reasonable option. A major challenge is how to develop effective methods to fully exploit the complex interactions within the DNA sequences to improve the predictive capability. RESULTS: In this work, we propose MSNet-4mC, a lightweight neural network building upon convolutional operations with multi-scale receptive fields to perceive cross-element relationships over both short and long ranges of given DNA sequences. With strong imbalances in the number of candidates in different species in mind, we compute and apply class weights in the cross-entropy loss to balance the training process. Extensive benchmarking experiments show that our method achieves a significant performance improvement and outperforms other state-of-the-art methods. AVAILABILITY AND IMPLEMENTATION: The source code and models are freely available for download at https://github.com/LIU-CT/MSNet-4mC, implemented in Python and supported on Linux and Windows. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

"Mamonoviridae", a proposed new family of the phylum Nucleocytoviricota.

Acanthamoeba castellanii medusavirus J1 is a giant virus that was isolated from a hot spring in Japan in 2019. Recently, a close relative of this virus, named medusavirus stheno T3, was isolated in Japan. Here, we describe their morphological, genomic, and gene content similarities and also propose to create a new family, "Mamonoviridae", a new genus, "Medusavirus", and two species, "Medusavirus medusae" and "Medusavirus sthenus", to classify these two viruses within the phylum Nucleocytoviricota.

Taxonomic update for giant viruses in the order Imitervirales (phylum Nucleocytoviricota).

Large DNA viruses in the phylum Nucleocytoviricota, sometimes referred to as "giant viruses" owing to their large genomes and virions, have been the subject of burgeoning interest over the last decade. Here, we describe recently adopted taxonomic updates for giant viruses within the order Imitervirales. The families Allomimiviridae, Mesomimiviridae, and Schizomimiviridae have been created to accommodate the increasing diversity of mimivirus relatives that have sometimes been referred to in the literature as "extended Mimiviridae". In addition, the subfamilies Aliimimivirinae, Megamimivirinae, and Klosneuvirinae have been established to refer to subgroups of the Mimiviridae. Binomial names have also been adopted for all recognized species in the order. For example, Acanthamoeba polyphaga mimivirus is now classified in the species Mimivirus bradfordmassiliense.

Functional Differentiation among the Arabidopsis Phosphatidylinositol 4-Phosphate 5-Kinase Genes PIP5K1, PIP5K2 and PIP5K3.

Phosphatidylinositol 4-phosphate 5-kinase (PIP5K) is involved in regulating various cellular processes through the signaling function of its product, phosphatidylinositol (4,5)-bisphosphate. Higher plants encode a large number of PIP5Ks forming distinct clades in their molecular phylogenetic tree. Although biological functions of PIP5K genes have been analyzed intensively in Arabidopsis thaliana, it remains unclear how those functions differ across clades of paralogs. We performed comparative functional analysis of the Arabidopsis genes encoding PIP5K1, PIP5K2 and PIP5K3, of which the first two and the last belong to closely related but distinct clades, to clarify their conserved and/or differentiated functions. Genetic analysis with their single and multiple mutants revealed that PIP5K1 and PIP5K3 have non-overlapping functions, with the former in total plant growth and the latter in root hair elongation, whereas PIP5K2 redundantly functions in both phenomena. This pattern of functional redundancy is explainable in terms of the overlapping pattern of their promoter activities. In transformation rescue experiments, PIP5K3 promoter-directed PIP5K1-YFP completely rescued the short-root-hair phenotype of pip5k3. However, PIP5K3-YFP could substitute for PIP5K1-YFP only partially in rescuing the severe dwarfism of pip5k1pip5k2 when directed by the PIP5K1 promoter. Phylogenetic analysis of angiosperm PIP5Ks revealed that PIP5K3 orthologs have a faster rate of diversification in their amino-acid sequences compared with PIP5K1/2 orthologs after they arose through a eudicot-specific duplication event. These findings suggest that PIP5K3 specialized to promote root hair elongation and lost some of the protein-encoded functions retained by PIP5K1 and PIP5K2, whereas PIP5K1 differentiated from PIP5K2 only in its promoter-directed expression pattern.

A persistent giant algal virus, with a unique morphology, encodes an unprecedented number of genes involved in energy metabolism.

Viruses have long been viewed as entities possessing extremely limited metabolic capacities. Over the last decade, however, this view has been challenged, as metabolic genes have been identified in viruses possessing large genomes and virions-the synthesis of which is energetically demanding. Here, we unveil peculiar phenotypic and genomic features of Prymnesium kappa virus RF01 (PkV RF01), a giant virus of the Mimiviridae family. We found that this virus encodes an unprecedented number of proteins involved in energy metabolism, such as all four succinate dehydrogenase (SDH) subunits (A-D) as well as key enzymes in the ß-oxidation pathway. The SDHA gene was transcribed upon infection, indicating that the viral SDH is actively used by the virus- potentially to modulate its host's energy metabolism. We detected orthologous SDHA and SDHB genes in numerous genome fragments from uncultivated marine Mimiviridae viruses, which suggests that the viral SDH is widespread in oceans. PkV RF01 was less virulent compared with other cultured prymnesioviruses, a phenomenon possibly linked to the metabolic capacity of this virus and suggestive of relatively long co-evolution with its hosts. It also has a unique morphology, compared to other characterized viruses in the Mimiviridae family. Finally, we found that PkV RF01 is the only alga-infecting Mimiviridae virus encoding two aminoacyl-tRNA synthetases and enzymes corresponding to an entire base-excision repair pathway, as seen in heterotroph-infecting Mimiviridae These Mimiviridae encoded-enzymes were found to be monophyletic and branching at the root of the eukaryotic tree of life. This placement suggests that the last common ancestor of Mimiviridae was endowed with a large, complex genome prior to the divergence of known extant eukaryotes.IMPORTANCE Viruses on Earth are tremendously diverse in terms of morphology, functionality, and genomic composition. Over the last decade, the conceptual gap separating viruses and cellular life has tightened because of the detection of metabolic genes in viral genomes that express complex virus phenotypes upon infection. Here, we describe Prymnesium kappa virus RF01, a large alga-infecting virus with a unique morphology, an atypical infection profile, and an unprecedented number of genes involved in energy metabolism (such as the tricarboxylic (TCA) cycle and the ß-oxidation pathway). Moreover, we show that the gene corresponding to one of these enzymes (the succinate dehydrogenase subunit A) is transcribed during infection and is widespread among marine viruses. This discovery provides evidence that a virus has the potential to actively regulate energy metabolism with its own gene.

Plankton networks driving carbon export in the oligotrophic ocean.

The biological carbon pump is the process by which CO2 is transformed to organic carbon via photosynthesis, exported through sinking particles, and finally sequestered in the deep ocean. While the intensity of the pump correlates with plankton community composition, the underlying ecosystem structure driving the process remains largely uncharacterized. Here we use environmental and metagenomic data gathered during the Tara Oceans expedition to improve our understanding of carbon export in the oligotrophic ocean. We show that specific plankton communities, from the surface and deep chlorophyll maximum, correlate with carbon export at 150 m and highlight unexpected taxa such as Radiolaria and alveolate parasites, as well as Synechococcus and their phages, as lineages most strongly associated with carbon export in the subtropical, nutrient-depleted, oligotrophic ocean. Additionally, we show that the relative abundance of a few bacterial and viral genes can predict a significant fraction of the variability in carbon export in these regions.

KofamKOALA: KEGG Ortholog assignment based on profile HMM and adaptive score threshold.

SUMMARY: KofamKOALA is a web server to assign KEGG Orthologs (KOs) to protein sequences by homology search against a database of profile hidden Markov models (KOfam) with pre-computed adaptive score thresholds. KofamKOALA is faster than existing KO assignment tools with its accuracy being comparable to the best performing tools. Function annotation by KofamKOALA helps linking genes to KEGG resources such as the KEGG pathway maps and facilitates molecular network reconstruction. AVAILABILITY AND IMPLEMENTATION: KofamKOALA, KofamScan and KOfam are freely available from GenomeNet (https://www.genome.jp/tools/kofamkoala/). SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

The complete genomic sequence of the novel myovirus RP13 infecting Ralstonia solanacearum, the causative agent of bacterial wilt.

A novel lytic bacteriophage, Ralstonia phage RP13, was isolated from tomato fields in Pang Nga, Thailand. Electron microscopic observation showed it to have the features of a myovirus with a novel triangulation number (T = 21, dextro). The RP13 DNA appeared to be heavily modified. By applying RNA sequencing and RNA-sequence-mediated DNA sequencing, the whole genome of RP31 was determined to be 170,942 bp in length with a mean G+C content of 39.2%. A total of 277 ORFs were identified as structural, functional, or hypothetical genes in addition to four tRNA genes. Phylogenetic analysis suggested that RP13 is not closely related to any other known phages. Thus, we concluded that the RP13 is a novel phage infecting R. solanacearum strains and will be a useful biocontrol agent against bacterial wilt disease.

Multiple mutations in RNA polymerase ß-subunit gene (rpoB) in Streptomyces incarnatus NRRL8089 enhance production of antiviral antibiotic sinefungin: modeling rif cluster region by density functional theory.

Streptomyces incarnatus NRRL8089 produces the antiviral, antifungal, antiprotozoal nucleoside antibiotic sinefungin. To enhance sinefungin production, multiple mutations were introduced to the rpoB gene encoding RNA polymerase (RNAP) ß-subunit at the target residues, D447, S453, H457, and R460. Sparse regression analysis using elastic-net lasso-ridge penalties on previously reported H457X mutations identified a numeric parameter set, which suggested that H457R/Y/F may cause production enhancement. H457R/R460C mutation successfully enhanced the sinefungin production by 3-fold, while other groups of mutations, such as D447G/R460C or D447G/H457Y, made moderate or even negative effects. To identify why the rif cluster residues have diverse effects on sinefungin production, an RNAP/DNA/mRNA complex model was constructed by homology modeling and molecular dynamics simulation. The 4 residues were located near the mRNA strand. Density functional theory-based calculation suggested that D447, H457, and R460 are in direct contact with ribonucleotide, and partially positive charges are induced by negatively charged chain of mRNA.

The Evolution of Rag Gene Enhancers and Transcription Factor E and Id Proteins in the Adaptive Immune System.

Adaptive immunity relies on the V(D)J DNA recombination of immunoglobulin (Ig) and T cell receptor (TCR) genes, which enables the recognition of highly diverse antigens and the elicitation of antigen-specific immune responses. This process is mediated by recombination-activating gene (Rag) 1 and Rag2 (Rag1/2), whose expression is strictly controlled in a cell type-specific manner; the expression of Rag1/2 genes represents a hallmark of lymphoid lineage commitment. Although Rag genes are known to be evolutionally conserved among jawed vertebrates, how Rag genes are regulated by lineage-specific transcription factors (TFs) and how their regulatory system evolved among vertebrates have not been fully elucidated. Here, we reviewed the current body of knowledge concerning the cis-regulatory elements (CREs) of Rag genes and the evolution of the basic helix-loop-helix TF E protein regulating Rag gene CREs, as well as the evolution of the antagonist of this protein, the Id protein. This may help to understand how the adaptive immune system develops along with the evolution of responsible TFs and enhancers.

Medusavirus, a Novel Large DNA Virus Discovered from Hot Spring Water.

Recent discoveries of new large DNA viruses reveal high diversity in their morphologies, genetic repertoires, and replication strategies. Here, we report the novel features of medusavirus, a large DNA virus newly isolated from hot spring water in Japan. Medusavirus, with a diameter of 260 nm, shows a T=277 icosahedral capsid with unique spherical-headed spikes on its surface. It has a 381-kb genome encoding 461 putative proteins, 86 of which have their closest homologs in Acanthamoeba, whereas 279 (61%) are orphan genes. The virus lacks the genes encoding DNA topoisomerase II and RNA polymerase, showing that DNA replication takes place in the host nucleus, whereas the progeny virions are assembled in the cytoplasm. Furthermore, the medusavirus genome harbored genes for all five types of histones (H1, H2A, H2B, H3, and H4) and one DNA polymerase, which are phylogenetically placed at the root of the eukaryotic clades. In contrast, the host amoeba encoded many medusavirus homologs, including the major capsid protein. These facts strongly suggested that amoebae are indeed the most promising natural hosts of medusavirus, and that lateral gene transfers have taken place repeatedly and bidirectionally between the virus and its host since the early stage of their coevolution. Medusavirus reflects the traces of direct evolutionary interactions between the virus and eukaryotic hosts, which may be caused by sharing the DNA replication compartment and by evolutionarily long lasting virus-host relationships. Based on its unique morphological characteristics and phylogenomic relationships with other known large DNA viruses, we propose that medusavirus represents a new family, MedusaviridaeIMPORTANCE We have isolated a new nucleocytoplasmic large DNA virus (NCLDV) from hot spring water in Japan, named medusavirus. This new NCLDV is phylogenetically placed at the root of the eukaryotic clades based on the phylogenies of several key genes, including that encoding DNA polymerase, and its genome surprisingly encodes the full set of histone homologs. Furthermore, its laboratory host, Acanthamoeba castellanii, encodes many medusavirus homologs in its genome, including the major capsid protein, suggesting that the amoeba is the genuine natural host from ancient times of this newly described virus and that lateral gene transfers have repeatedly occurred between the virus and amoeba. These results suggest that medusavirus is a unique NCLDV preserving ancient footprints of evolutionary interactions with its hosts, thus providing clues to elucidate the evolution of NCLDVs, eukaryotes, and virus-host interaction. Based on the dissimilarities with other known NCLDVs, we propose that medusavirus represents a new viral family, Medusaviridae.

Effects of growth hormone treatment on thyroid function in pediatric patients with Prader-Willi syndrome.

It is unclear whether hypothyroidism is present in patients with Prader-Willi syndrome (PWS). This study aimed to clarify the state of the hypothalamic-pituitary-thyroid axis and the effects of growth hormone (GH) treatment on thyroid function in pediatric patients with PWS. We retrospectively evaluated thyroid function in 51 patients with PWS before GH treatment using a thyroid-releasing hormone (TRH) stimulation test (29 males and 22 females; median age, 22 months). We also evaluated the effect of GH therapy on thyroid function by comparing serum free triiodothyronine (fT3), free thyroxine (fT4), and thyroid stimulating hormone (TSH) levels at baseline, 1 year, and 2 years after GH therapy. TSH, fT4, and fT3 levels were 2.28 µU/ml (interquartile range [IQR]; 1.19-3.61), 1.18 ng/dl (IQR; 1.02-1.24), and 4.02 pg/dl (IQR; 3.54-4.40) at baseline, respectively. In 49 of 51 patients, the TSH response to TRH administration showed a physiologically normal pattern; in two patients (4.0%), the pattern suggested hypothalamic hypothyroidism (delayed and prolonged TSH peak after TRH administration). TSH, fT4, and fT3 levels did not change significantly during 1 or 2 years after GH treatment. The TSH response to TRH showed a normal pattern in most patients, and thyroid function did not change significantly during the 2 years after initiating GH treatment.

Deficiency of Gankyrin in the small intestine is associated with augmented colitis accompanied by altered bacterial composition of intestinal microbiota.

BACKGROUND: Gankyrin (GK) is an oncoprotein which regulates inflammatory responses and its inhibition is considered as a possible anti-inflammatory therapy for inflammatory bowel disease (IBD). METHODS: In this study, we investigated the role of GK in epithelial cells using mice with intestinal epithelial cell-specific GK deletion in (i) the entire small intestine and colon (Villin-Cre;Gankyrinf/f) and (ii) the distal intestine and colon (Cdx2-Cre;Gankyrinf/f). RESULT: Unexpectedly, GK-deficiency in the upper small bowel augmented inflammatory activity compared with control mice when colitis was induced with dextran sodium sulfate. Biochemical analyses have revealed GK-deficiency to have caused reduction in the expression of antimicrobial peptides, α-Defensin-5 and -6, in the upper small bowel. Examination of human samples have further confirmed that the reduction of GK expression in the small bowel is associated with colonic involvement in human Crohn's disease. Through the sequencing of bacterial 16S rRNA gene amplicons, bacteria potentially deleterious to intestinal homeostasis such as Helicobacter japonicum and Bilophila were found to be over-represented in colitis induced Villin-Cre;Gankyrinf/f mice when compared to Gankyrinf/f control mice under the same condition. CONCLUSION: These results highlight the distinct site dependence of the pro- and anti-inflammatory functions of GK and provide important insights into the pathogenesis of IBD.