High-entropy engineering with regulated defect structure and electron interaction tuning active sites for trifunctional electrocatalysis.

High-entropy alloy nanoparticles (HEANs) possessing regulated defect structure and electron interaction exhibit a guideline for constructing multifunctional catalysts. However, the microstructure-activity relationship between active sites of HEANs for multifunctional electrocatalysts is rarely reported. In this work, HEANs distributed on multi-walled carbon nanotubes (HEAN/CNT) are prepared by Joule heating as an example to explain the mechanism of trifunctional electrocatalysis for oxygen reduction, oxygen evolution, and hydrogen evolution reaction. HEAN/CNT excels with unmatched stability, maintaining a 0.8V voltage window for 220 h in zinc-air batteries. Even after 20 h of water electrolysis, its performance remains undiminished, highlighting exceptional endurance and reliability. Moreover, the intrinsic characteristics of the defect structure and electron interaction for HEAN/CNT are investigated in detail. The electrocatalytic mechanism of trifunctional electrocatalysis of HEAN/CNT under different conditions is identified by in situ monitoring and theoretical calculation. Meanwhile, the electron interaction and adaptive regulation of active sites in the trifunctional electrocatalysis of HEANs were further verified by density functional theory. These findings could provide unique ideas for designing inexpensive multifunctional high-entropy electrocatalysts.

Revealing the Orbital Interactions between Dissimilar Metal Sites during Oxygen Reduction Process.

A FeCo/DA@NC catalyst with the well-defined FeCoN6 moiety is customized through a novel and ultrafast Joule heating technique. This catalyst demonstrates superior oxygen reduction reaction activity and stability in an alkaline environment. The power density and charge-discharge cycling of znic-air batteries driven by FeCo/DA@NC also surpass those of Pt/C catalyst. The source of the excellent oxygen reduction reaction activity of FeCo/DA@NC originates from the significantly changed charge environment and 3d orbital spin state. These not only improve the bonding strength between active sites and oxygen-containing intermediates, but also provide spare reaction sites for oxygen-containing intermediates. Moreover, various in situ detection techniques reveal that the rate-determining step in the four-electron oxygen reduction reaction is *O2 protonation. This work provides strong support for the precise design and rapid preparation of bimetallic catalysts and opens up new ideas for understanding orbital interactions during oxygen reduction reactions.

Regulated High-Spin State and Constrained Charge Behavior of Active Cobalt Sites in Covalent Organic Frameworks for Promoting Electrocatalytic Oxygen Reduction.

A novel type of covalent organic frameworks has been developed by assembling definite cobalt-nitrogen-carbon configurations onto carbon nanotubes using linkers that have varying electronic effects. This innovative approach has resulted in an efficient electrocatalyst for oxygen reduction, which is understood by a combination of in situ spectroelectrochemistry and the bond order theorem. The strong interaction between the electron-donating carbon nanotubes and the electron-accepting linker mitigates the trend of charge loss at cobalt sites, while inducing the generation of high spin state. This enhances the adsorption strength and electron transfer between the cobalt center and reactants/intermediates, leading to an improved oxygen reduction capability. This work not only presents an effective strategy for developing efficient non-noble metal electrocatalysts through reticular chemistry, but also provides valuable insights into regulating the electronic configuration and charge behavior of active sites in designing high-performance electrocatalysts.

Lectin receptor-like kinase LecRK-VIII.2 is a missing link in MAPK signaling-mediated yield control.

The energy allocation for vegetative and reproductive growth is regulated by developmental signals and environmental cues, which subsequently affects seed output. However, the molecular mechanism underlying how plants coordinate yield-related traits to control yield in changing source-sink relationships remains largely unknown. Here, we discovered the lectin receptor-like kinase LecRK-VIII.2 as a specific receptor-like kinase that coordinates silique number, seed size, and seed number to determine seed yield in Arabidopsis (Arabidopsis thaliana). The lecrk-VIII.2 mutants develop smaller seeds, but more siliques and seeds, leading to increased yield. In contrast, the plants overexpressing LecRK-VIII.2 form bigger seeds, but less siliques and seeds, which results in similar yield to that of wild-type plants. Interestingly, LecRK-VIII.2 promotes the growth of the rosette, root, and stem by coordinating the source-sink relationship. Additionally, LecRK-VIII.2 positively regulates cell expansion and proliferation in the seed coat, and maternally controls seed size. The genetic and biochemical analyses demonstrated that LecRK-VIII.2 acts upstream of the mitogen-activated protein kinase (MAPK) gene MPK6 to regulate silique number, seed size, and seed number. Collectively, these findings uncover LecRK-VIII.2 as an upstream component of the MAPK signaling pathway to control yield-related traits and suggest its potential for crop improvement aimed at developing plants with stable yield, a robust root system, and improved lodging resistance.

Genome Analysis of a Novel Polysaccharide-Degrading Bacterium Paenibacillus algicola and Determination of Alginate Lyases.

Carbohydrate-active enzymes (CAZymes) are an important characteristic of bacteria in marine systems. We herein describe the CAZymes of Paenibacillus algicola HB172198T, a novel type species isolated from brown algae in Qishui Bay, Hainan, China. The genome of strain HB172198T is a 4,475,055 bp circular chromosome with an average GC content of 51.2%. Analysis of the nucleotide sequences of the predicted genes shows that strain HB172198T encodes 191 CAZymes. Abundant putative enzymes involved in the degradation of polysaccharides were identified, such as alginate lyase, agarase, carrageenase, xanthanase, xylanase, amylases, cellulase, chitinase, fucosidase and glucanase. Four of the putative polysaccharide lyases from families 7, 15 and 38 were involved in alginate degradation. The alginate lyases of strain HB172198T exhibited the maximum activity 152 U/mL at 50 °C and pH 8.0, and were relatively stable at pH 7.0 and temperatures lower than 40 °C. The average degree of polymerization (DP) of the sodium alginate oligosaccharide (AOS) degraded by the partially purified alginate lyases remained around 14.2, and the thin layer chromatography (TCL) analysis indicated that it contained DP2-DP8 oligosaccharides. The complete genome sequence of P. algicola HB172198T will enrich our knowledge of the mechanism of polysaccharide lyase production and provide insights into its potential applications in the degradation of polysaccharides such as alginate.

Identification of polycistronic transcriptional units and non-canonical introns in green algal chloroplasts based on long-read RNA sequencing data.

BACKGROUND: Chloroplasts are important semi-autonomous organelles in plants and algae. Unlike higher plants, the chloroplast genomes of green algal linage have distinct features both in organization and expression. Despite the architecture of chloroplast genome having been extensively studied in higher plants and several model species of algae, little is known about the transcriptional features of green algal chloroplast-encoded genes. RESULTS: Based on full-length cDNA (Iso-Seq) sequencing, we identified widely co-transcribed polycistronic transcriptional units (PTUs) in the green alga Caulerpa lentillifera. In addition to clusters of genes from the same pathway, we identified a series of PTUs of up to nine genes whose function in the plastid is not understood. The RNA data further allowed us to confirm widespread expression of fragmented genes and conserved open reading frames, which are both important features in green algal chloroplast genomes. In addition, a newly fragmented gene specific to C. lentillifera was discovered, which may represent a recent gene fragmentation event in the chloroplast genome. With the newly annotated exon-intron boundary information, gene structural annotation was greatly improved across the siphonous green algae lineages. Our data also revealed a type of non-canonical Group II introns, with a deviant secondary structure and intronic ORFs lacking known splicing or mobility domains. These widespread introns have conserved positions in their genes and are excised precisely despite lacking clear consensus intron boundaries. CONCLUSION: Our study fills important knowledge gaps in chloroplast genome organization and transcription in green algae, and provides new insights into expression of polycistronic transcripts, freestanding ORFs and fragmented genes in algal chloroplast genomes. Moreover, we revealed an unusual type of Group II intron with distinct features and conserved positions in Bryopsidales. Our data represents interesting additions to knowledge of chloroplast intron structure and highlights clusters of uncharacterized genes that probably play important roles in plastids.

Comprehensive genomic survey, structural classification and expression analysis of C2H2-type zinc finger factor in wheat (Triticum aestivum L.).

BACKGROUND: The C2H2-type zinc finger proteins (C2H2-ZFPs) are one of major classes of transcription factors that play important roles in plant growth, development and stress responses. Limit information about the C2H2-ZF genes hinders the molecular breeding in bread wheat (Triticum aestivum). RESULTS: In this study, 457 C2H2-ZFP proteins (including 253 splice variants), which contain four types of conserved domain (named Q, M, Z, and D), could be further classified into ten subsets. They were identified to be distributed in 21 chromosomes in T. aestivum. Subset-specific motifs, like NPL-, SFP1-, DL- (EAR-like-motif), R-, PL-, L- and EK-, might make C2H2-ZFP diverse multifunction. Interestingly, NPL- and SFP1-box were firstly found to be located in C2H2-ZFP proteins. Synteny analyses showed that only 4 pairs of C2H2 family genes in T. aestivum, 65 genes in B. distachyon, 66 genes in A. tauschii, 68 genes in rice, 9 genes in Arabidopsis, were syntenic relationships respectively. It indicated that TaZFPs were closely related to genes in Poaceae. From the published transcriptome data, totally 198 of 204 TaC2H2-ZF genes have expression data. Among them, 25 TaC2H2-ZF genes were certificated to be significantly differentially expressed in 5 different organs and 15 different development stages by quantitative RT-PCR. The 18 TaC2H2-ZF genes were verified in response to heat, drought, and heat & drought stresses. According to expression pattern analysis, several TaZFPs, like Traes_5BL_D53A846BE.1, were not only highly expressed in L2DAAs, RTLS, RMS, but also endowed tolerance to drought and heat stresses, making them good candidates for molecular breeding. CONCLUSIONS: This study systematically characterized the TaC2H2-ZFPs and their potential roles in T. aestivum. Our findings provide new insights into the C2H2-ZF genes in T. aestivum as well as a foundation for further studies on the roles of TaC2H2-ZF genes in T. aestivum molecular breeding.

Microbulbifer harenosus sp. nov., an alginate-degrading bacterium isolated from coastal sand.

A Gram-stain-negative, aerobic, rod-shaped bacterium with peritrichous flagella, designated strain HB161719T, was isolated from coastal sand collected from Tanmen Port in Hainan, PR China. The isolate was found to grow with 2-11 % (w/v) NaCl, at 15-45 °C and pH 6.0-10.0, with an optima of 2-3 % NaCl, 37 °C and pH 7.0, respectively. Chemotaxonomic analysis showed that Q-8 was detected as the sole respiratory quinone and that iso-C15 : 0 and summed features 3, 8 and 9 were the major cellular fatty acids. The G+C content of the genomic DNA was 58.2 mol%. Analysis of the 16S rRNA gene sequence of the strain showed an affiliation with the genus Microbulbifer, sharing 98.7, 98.4, 97.8 and 97.8 % sequence similarities to the closest relatives of Microbulbifer okinawensis ABABA23T, Microbulbifer pacificus SPO729T, Microbulbifer taiwanensis CC-LN1-12T and Microbulbifer gwangyangensis GY2T, respectively. Low DNA-DNA hybridization values showed that it formed a distinct genomic species. The combined phenotypic and molecular features supported that strain HB161719T represents a novel species of the genus Microbulbifer, for which the name Microbulbifer harenosus sp. nov. is proposed. The type strain is HB161719T (=CGMCC 1.13584T=JCM 32688T).

Alteromonas portus sp. nov., an alginate lyase-excreting marine bacterium.

An alginate lyase-excreting bacterium, designated strain HB161718T, was isolated from coastal sand collected from Tanmen Port in Hainan, PR China. Cells were Gram-stain-negative rods and motile with a single polar flagellum. Its major isoprenoid quinone was ubiquinone 8 (Q-8), and its cellular fatty acid profile mainly consisted of C16 : 1 ω7c and/or C16 : 1 ω6c, C18 : 1 ω6c and/or C18 : 1 ω7c, C16 : 0, C17 : 0 10-methyl and C16 : 0 N alcohol. The G+C content of the genomic DNA was 44.1 mol%. 16S rRNA gene sequence analysis suggested that strain HB161718T belonged to the genus Alteromonas, sharing 99.5, 99.4, 99.2, 98.9 and 98.5 % sequence similarities to its closest relatives, Alteromonas macleodii JCM 20772T, Alteromonas gracilis 9a2T, Alteromonas australica H17T, Alteromonas marina SW-47T and Alteromonas mediterranea DET, respectively. The low values of DNA-DNA hybridization and average nucleotide identity showed that it formed a distinct genomic species. The combined phenotypic and molecular features supported the conclusion that strain HB161718T represents a novel species of the genus Alteromonas, for which the name Alteromonas portus sp. nov. is proposed. The type strain is HB161718T (=CGMCC 1.13585T=JCM 32687T).

Paenibacillus algicola sp. nov., a novel alginate lyase-producing marine bacterium.

A Gram-stain-variable, facultatively anaerobic, endospore-forming, rod-shaped bacterium, designated HB172198T, was isolated from brown alga collected at Qishui Bay, Hainan, PR China. Phylogenetic analysis of 16S rRNA gene sequences indicated that strain HB172198T belonged to the genus Paenibacillus, and the closest phylogenetically related species was Paenibacillus lemnae NBRC 109972T (97.6% similarity). The other 16S rRNA gene sequence similarities were under 97.0%. The whole genome average nucleotide identity value between strain HB172198T and the closest type strain was 75.3% and the in silico DNA-DNA hybridization value was 20.2%. The predominant isoprenoid quinone was menaquinone 7 and the major fatty acids were anteiso-C15:0, C16:0, anteiso-C17:0, iso C16:0 and C16:1 ω11c. The combined phylogenetic relatedness, phenotypic and genotypic features supported the conclusion that strain HB172198T represents a novel species of the genus Paenibacillus, for which the name Paenibacillus algicola sp. nov. is proposed. The type strain is HB172198T (=CGMCC 1.13583T=JCM 32683T).

Nonomuraea mangrovi sp. nov., an actinomycete isolated from mangrove soil.

A novel aerobic actinomycete, designated HA15826T, was isolated from a mangrove soil sample collected in Sanya, China. Scanning electron microscopy revealed that the isolate produced straight to slightly flexural spore chains with rough cylindrical spores. Chemotaxonomic tests showed that the cell wall contained meso-diaminopimelic acid and the major fatty acids were iso-C16 : 0, 10-methyl-C17 : 0, C17 : 1ω8c and C16 : 0. 16S rRNA gene sequence similarity analysis showed that strain HA15826T belonged to the genus Nonomuraea, being most closely related to Nonomuraea dietziae DSM 44320T (98.7 %), Nonomuraea candida HMC10T (98.4 %), Nonomuraea africana IFO 14745T (98.4 %), Nonomuraea roseola IFO 14685T (98.2 %) and Nonomuraea recticatena IFO 14525T (98.1 %). The DNA G+C content of the type strain is 73.2 %. DNA-DNA relatedness and comparative analyses of physiological, biochemical and chemotaxonomic data allowed genotypic and phenotypic differentiation of strain HA15826T from the closely related species. Thus, strain HA15826T should be classified as a novel species of the genus Nonomuraea, for which the name Nonomuraeamangrovi sp. nov. is proposed. The type strain is HA15826T (=CGMCC 4.7425T=DSM 105694T).

Streptomyces caeni sp. nov., isolated from mangrove mud.

A novel aerobic actinomycete, designated as HA15955T, was isolated from a mangrove mud sample collected in Sanya, China. Scanning electron microscopy revealed that HA15955T produced straight to spiral spore chains with smooth cylindrical spores. 16S rRNA gene sequence similarity showed that strain HA15955T belonged to the genus Streptomyces, was most closely related to Streptomyces speibonae NRRL B-24240T (98.7 % similarity) and formed a distinct subclade. The low relatedness value of DNA-DNA hybridization showed that it formed a distinct genomic species. Based on phenotypic, genotypic and phylogenetic data, strain HA15955T should be classified as a novel species of the genus Streptomyces, for which the name Streptomycescaeni sp. nov. is proposed. The type strain is HA15955T (=CGMCC 4.7426T=DSM 105693T).

Paenibacillus silvae sp. nov., isolated from a tropical rainforest soil.

Two Gram-stain-positive, facultatively aerobic, endospore-forming and rod-shaped bacteria, designated DB13031T and DB13311, were isolated from the soil of the Jiaxi Nature Reserve in Hainan, PR China. 16S rRNA gene analysis of strains DB13031T and DB13311 showed that they fell within the Paenibacillus cluster, with highest similarities to Paenibacillus cucumis AP-115T (98.4 and 98.3 %, respectively), Paenibacillus barcinonensis BP-23T (98.3 and 98.2 %, respectively) and Paenibacillus oceanisediminis L10T (97.7 and 97.7 %, respectively). The DNA-DNA hybridization values between strain DB13031T and the type strains of its closest related species were 48.2, 38.1 and 43.5 %. Strain DB13031T contained menaquinone-7 (MK-7) as the predominant isoprenoid quinone and anteiso-C15 : 0, iso-C16 : 0 and C16 : 0 as the major cellular fatty acids. The cell-wall peptidoglycan was of the A1γ type and the major polar lipid profiles were diphosphatidylglycerol, phosphatidylethanolamine, four unknown aminophospholipids and four unknown phospholipids. Based on the phenotypic and genotypic data, it is proposed that the two isolates represent a novel species of the genus Paenibacillus, for which the name Paenibacillus silvae sp. nov. is proposed. The type strain is DB13031T (=CGMCC 1.12770T=DSM 28013T).

Widespread and evolutionary analysis of a MITE family Monkey King in Brassicaceae.

BACKGROUND: Miniature inverted repeat transposable elements (MITEs) are important components of eukaryotic genomes, with hundreds of families and many copies, which may play important roles in gene regulation and genome evolution. However, few studies have investigated the molecular mechanisms involved. In our previous study, a Tourist-like MITE, Monkey King, was identified from the promoter region of a flowering time gene, BnFLC.A10, in Brassica napus. Based on this MITE, the characteristics and potential roles on gene regulation of the MITE family were analyzed in Brassicaceae. RESULTS: The characteristics of the Tourist-like MITE family Monkey King in Brassicaceae, including its distribution, copies and insertion sites in the genomes of major Brassicaceae species were analyzed in this study. Monkey King was actively amplified in Brassica after divergence from Arabidopsis, which was indicated by the prompt increase in copy number and by phylogenetic analysis. The genomic variations caused by Monkey King insertions, both intra- and inter-species in Brassica, were traced by PCR amplification. Genomic sequence analysis showed that most complete Monkey King elements are located in gene-rich regions, less than 3kb from genes, in both the B. rapa and A. thaliana genomes. Sixty-seven Brassica expressed sequence tags carrying Monkey King fragments were also identified from the NCBI database. Bisulfite sequencing identified specific DNA methylation of cytosine residues in the Monkey King sequence. A fragment containing putative TATA-box motifs in the MITE sequence could bind with nuclear protein(s) extracted from leaves of B. napus plants. A Monkey King-related microRNA, bna-miR6031, was identified in the microRNA database. In transgenic A. thaliana, when the Monkey King element was inserted upstream of 35S promoter, the promoter activity was weakened. CONCLUSION: Monkey King, a Brassicaceae Tourist-like MITE family, has amplified relatively recently and has induced intra- and inter-species genomic variations in Brassica. Monkey King elements are most abundant in the vicinity of genes and may have a substantial effect on genome-wide gene regulation in Brassicaceae. Monkey King insertions potentially regulate gene expression and genome evolution through epigenetic modification and new regulatory motif production.

Streptomyces mangrovi sp. nov., an actinomycete from mangrove soil.

A novel aerobic actinomycete, designated HA11110(T), was isolated from a mangrove soil sample collected in Haikou, China. It formed white aerial mycelium and pale yellow substrate mycelium on Gause's synthetic agar no. 1. Scanning electron microscopy revealed that cells of HA11110(T) produced straight to spiral spore chains with spiny spores. Chemotaxonomic tests showed that the cell wall contained LL-diaminopimelic acid and the major fatty acids were iso-C16 : 0, anteiso-C15 : 0 and iso-C14 : 0.16S rRNA gene sequence similarity showed that strain HA11110(T) belonged to the genus Streptomyces, most closely related to Streptomyces fenghuangensis GIMN4.003(T) (99.1%), Streptomyces nanhaiensis SCSIO 01248(T) (98.8%) and Streptomyces radiopugnans R97(T) (98.8%). However, DNA-DNA hybridization studies of strain HA11110T with these three closest relatives showed relatedness values of 58.4, 49.7 and 47.2%, respectively. On the basis of phenotypic and genotypic data, strain HA11110(T) represents a novel species of the genus Streptomyces, for which the name Streptomyces mangrovi sp. nov. is proposed. The type strain is HA11110(T) ( = CGMCC 4.7117(T)= DSM 42113(T)).

RNA barcode segments for SARS-CoV-2 identification from HCoVs and SARSr-CoV-2 lineages.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the pathogen responsible for coronavirus disease 2019 (COVID-19), continues to evolve, giving rise to more variants and global reinfections. Previous research has demonstrated that barcode segments can effectively and cost-efficiently identify specific species within closely related populations. In this study, we designed and tested RNA barcode segments based on genetic evolutionary relationships to facilitate the efficient and accurate identification of SARS-CoV-2 from extensive virus samples, including human coronaviruses (HCoVs) and SARSr-CoV-2 lineages. Nucleotide sequences sourced from NCBI and GISAID were meticulously selected and curated to construct training sets, encompassing 1733 complete genome sequences of HCoVs and SARSr-CoV-2 lineages. Through genetic-level species testing, we validated the accuracy and reliability of the barcode segments for identifying SARS-CoV-2. Subsequently, 75 main and subordinate species-specific barcode segments for SARS-CoV-2, located in ORF1ab, S, E, ORF7a, and N coding sequences, were intercepted and screened based on single-nucleotide polymorphism sites and weighted scores. Post-testing, these segments exhibited high recall rates (nearly 100%), specificity (almost 30% at the nucleotide level), and precision (100%) performance on identification. They were eventually visualized using one and two-dimensional combined barcodes and deposited in an online database (http://virusbarcodedatabase.top/). The successful integration of barcoding technology in SARS-CoV-2 identification provides valuable insights for future studies involving complete genome sequence polymorphism analysis. Moreover, this cost-effective and efficient identification approach also provides valuable reference for future research endeavors related to virus surveillance.

Bioinformatic Identification and Expression Analyses of the MAPK-MAP4K Gene Family Reveal a Putative Functional MAP4K10-MAP3K7/8-MAP2K1/11-MAPK3/6 Cascade in Wheat (Triticum aestivum L.).

The mitogen-activated protein kinase (MAPK) cascades act as crucial signaling modules that regulate plant growth and development, response to biotic/abiotic stresses, and plant immunity. MAP3Ks can be activated through MAP4K phosphorylation in non-plant systems, but this has not been reported in plants to date. Here, we identified a total of 234 putative TaMAPK family members in wheat (Triticum aestivum L.). They included 48 MAPKs, 17 MAP2Ks, 144 MAP3Ks, and 25 MAP4Ks. We conducted systematic analyses of the evolution, domain conservation, interaction networks, and expression profiles of these TaMAPK-TaMAP4K (representing TaMAPK, TaMAP2K, TaMAP3K, and TaMAP4K) kinase family members. The 234 TaMAPK-TaMAP4Ks are distributed on 21 chromosomes and one unknown linkage group (Un). Notably, 25 of these TaMAP4K family members possessed the conserved motifs of MAP4K genes, including glycine-rich motif, invariant lysine (K) motif, HRD motif, DFG motif, and signature motif. TaMAPK3 and 6, and TaMAP4K10/24 were shown to be strongly expressed not only throughout the growth and development stages but also in response to drought or heat stress. The bioinformatics analyses and qRT-PCR results suggested that wheat may activate the MAP4K10-MEKK7-MAP2K11-MAPK6 pathway to increase drought resistance in wheat, and the MAP4K10-MAP3K8-MAP2K1/11-MAPK3 pathway may be involved in plant growth. In general, our work identified members of the MAPK-MAP4K cascade in wheat and profiled their potential roles during their response to abiotic stresses and plant growth based on their expression pattern. The characterized cascades might be good candidates for future crop improvement and molecular breeding.

Single-cell profiling of the pig cecum at various developmental stages.

The gastrointestinal tract is essential for food digestion, nutrient absorption, waste elimination, and microbial defense. Single-cell transcriptome profiling of the intestinal tract has greatly enriched our understanding of cellular diversity, functional heterogeneity, and their importance in intestinal tract development and disease. Although such profiling has been extensively conducted in humans and mice, the single-cell gene expression landscape of the pig cecum remains unexplored. Here, single-cell RNA sequencing was performed on 45 572 cells obtained from seven cecal samples in pigs at four different developmental stages (days (D) 30, 42, 150, and 730). Analysis revealed 12 major cell types and 38 subtypes, as well as their distinctive genes, transcription factors, and regulons, many of which were conserved in humans. An increase in the relative proportions of CD8 + T and Granzyme A (low expression) natural killer T cells (GZMA low NKT) cells and a decrease in the relative proportions of epithelial stem cells, Tregs, RHEX + T cells, and plasmacytoid dendritic cells (pDCs) were noted across the developmental stages. Moreover, the post-weaning period exhibited an up-regulation in mitochondrial genes, COX2 and ND2, as well as genes involved in immune activation in multiple cell types. Cell-cell crosstalk analysis indicated that IBP6 + fibroblasts were the main signal senders at D30, whereas IBP6 - fibroblasts assumed this role at the other stages. NKT cells established interactions with epithelial cells and IBP6 + fibroblasts in the D730 cecum through mediation of GZMA-F2RL1/F2RL2 pairs. This study provides valuable insights into cellular heterogeneity and function in the pig cecum at different development stages.

The subcommissural organ regulates brain development via secreted peptides.

The subcommissural organ (SCO) is a gland located at the entrance of the aqueduct of Sylvius in the brain. It exists in species as distantly related as amphioxus and humans, but its function is largely unknown. To explore its function, we compared transcriptomes of SCO and non-SCO brain regions and found three genes, Sspo, Car3, and Spdef, that are highly expressed in the SCO. Mouse strains expressing Cre recombinase from endogenous promoter/enhancer elements of these genes were used to genetically ablate SCO cells during embryonic development, resulting in severe hydrocephalus and defects in neuronal migration and development of neuronal axons and dendrites. Unbiased peptidomic analysis revealed enrichment of three SCO-derived peptides, namely thymosin beta 4, thymosin beta 10, and NP24, and their reintroduction into SCO-ablated brain ventricles substantially rescued developmental defects. Together, these data identify a critical role for the SCO in brain development.