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A wide variety of bioactive peptides have been identified in the central nervous system and several peripheral tissues in the ascidian Ciona intestinalis type A (Ciona robusta). However, hemocyte endocrine peptides have yet to be explored. Here, we report a novel 14-amino-acid peptide, CiEMa, that is predominant in the granular hemocytes and unilocular refractile granulocytes of Ciona. RNA-seq and qRT-PCR revealed the high CiEma expression in the adult pharynx and stomach. Immunohistochemistry further revealed the highly concentrated CiEMa in the hemolymph of the pharynx and epithelial cells of the stomach, suggesting biological roles in the immune response. Notably, bacterial lipopolysaccharide stimulation of isolated hemocytes for 1–4 h resulted in 1.9- to 2.4-fold increased CiEMa secretion. Furthermore, CiEMa-stimulated pharynx exhibited mRNA upregulation of the growth factor (Fgf3/7/10/22), vanadium binding proteins (CiVanabin1 and CiVanabin3), and forkhead and homeobox transcription factors (Foxl2, Hox3, and Dbx) but not antimicrobial peptides (CrPap-a and CrMam-a) or immune-related genes (Tgfbtun3, Tnfa, and Il17-2). Collectively, these results suggest that CiEMa plays roles in signal transduction involving tissue development or repair in the immune response, rather than in the direct regulation of immune response genes. The present study identified a novel Ciona hemocyte peptide, CiEMa, which paves the way for research on the biological roles of hemocyte peptides in chordates. [ABSTRACT FROM AUTHOR]

Cholinergic signaling, which modulates cell activities via nicotinic and muscarinic acetylcholine receptors (n- and mAChRs) in response to internal or external stimuli, has been demonstrated in mammalian non-neuronal cells that synthesize acetylcholine (ACh). One of the major pathways of excitatory transmission in the enteric nervous system (ENS) is mediated by cholinergic transmission, with the transmitter ACh producing excitatory potentials in postsynaptic effector cells. In addition to ACh-synthesizing and ACh-metabolizing elements in the ENS, the presence of non-neuronal ACh machinery has been reported in epithelial cells of the small and large intestines of rats and humans. However, little is known about how non-neuronal ACh controls physiological function in the intestine. Here, experiments using crypt-villus organoids that lack nerve and immune cells in culture suggest that endogenous ACh is synthesized in the intestinal epithelium to drive organoid growth and differentiation through activation of nAChRs. Treatment of organoids with nicotine enhanced cell growth and the expression of marker genes for stem and epithelial cells. On the other hand, the nAChR antagonist mecamylamine strongly inhibited the growth and differentiation of organoids, suggesting the involvement of nAChRs in the regulation of proliferation and differentiation of Lgr5-positive stem cells. More specifically, RNA sequencing analysis revealed that Wnt5a expression was dramatically upregulated after nicotine treatment, and Wnt5a rescued organoid growth and differentiation in response to mecamylamine. Taken together, our results indicate that coordinated activities of nAChR and Wnt signaling maintain Lgr5-positive stem cell activity and balanced differentiation. Furthermore, we could clearly separate the two groups, neuronal ACh in the ENS and non-neuronal ACh in the intestinal epithelium. Dysfunction of the non-neuronal cholinergic system is involved in the pathogenesis of disease. The data will increase our understanding of the cholinergic properties of non-neuronal cells and lead to optimization of drug therapy. [ABSTRACT FROM AUTHOR]

The ability of stem cells to divide and differentiate is necessary for tissue repair and homeostasis. Appropriate spatial and temporal mechanisms are needed. Local intercellular signaling increases expression of specific genes that mediate and maintain differentiation. Diffusible signaling molecules provide concentration-dependent induction of specific patterns of cell types or regions. Differentiation of adjacent cells, on the other hand, requires cell–cell contact and subsequent signaling. These two types of signals work together to allow stem cells to provide what organisms require. The ability to grow organoids has increased our understanding of the cellular and molecular features of small "niches" that modulate stem cell function in various organs, including the small intestine. [ABSTRACT FROM AUTHOR]

The Gly-Asp-Ser-Leu (GDSL) motif of esterase/lipase family proteins (GELPs) generally exhibit esterase activity, whereas transferase activity is markedly preferred in several GELPs, including the Tanacetum cinerariifolium GDSL lipase TciGLIP, which is responsible for the biosynthesis of the natural insecticide, pyrethrin I. This transferase activity is due to the substrate affinity regulated by the protein structure and these features are expected to be conserved in transferase activity-exhibiting GELPs (tr-GELPs). In this study, we identified two amino acid residues, [N/R]208 and D484, in GELP sequence alignments as candidate key residues for the transferase activity of tr-GELPs by two-entropy analysis. Molecular phylogenetic analysis demonstrated that each tr-GELP is located in the clusters for non-tr-GELPs, and most GELPs conserve at least one of the two residues. These results suggest that the two conserved residues are required for the acquisition of transferase activity in the GELP family. Furthermore, substrate docking analyses using ColabFold-generated structure models of both natives and each of the two amino acids-mutated TciGLIPs also revealed numerous docking models for the proper access of substrates to the active site, indicating crucial roles of these residues of TciGLIP in its transferase activity. This is the first report on essential residues in tr-GELPs for the transferase activity.

The plant Tanacetum coccineum (painted daisy) is closely related to Tanacetum cinerariifolium (pyrethrum daisy). However, T. cinerariifolium produces large amounts of pyrethrins, a class of natural insecticides, whereas T. coccineum produces much smaller amounts of these compounds. Thus, comparative genomic analysis is expected to contribute a great deal to investigating the differences in biological defense systems, including pyrethrin biosynthesis. Here, we elucidated the 9.4 Gb draft genome of T. coccineum , consisting of 2,836,647 scaffolds and 103,680 genes. Comparative analyses of the draft genome of T. coccineum and that of T. cinerariifolium , generated in our previous study, revealed distinct features of T. coccineum genes. While the T. coccineum genome contains more numerous ribosome-inactivating protein (RIP)-encoding genes, the number of higher-toxicity type-II RIP-encoding genes is larger in T. cinerariifolium . Furthermore, the number of histidine kinases encoded by the T. coccineum genome is smaller than that of T. cinerariifolium , suggesting a biological correlation with pyrethrin biosynthesis. Moreover, the flanking regions of pyrethrin biosynthesis-related genes are also distinct between these two plants. These results provide clues to the elucidation of species-specific biodefense systems, including the regulatory mechanisms underlying pyrethrin production.