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Blastema formation is a crucial process that provides a cellular source for regenerating tissues and organs. While bilaterians have diversified blastema formation methods, its mechanisms in non-bilaterians remain poorly understood. Cnidarian jellyfish, or medusae, represent early-branching metazoans that exhibit complex morphology and possess defined appendage structures highlighted by tentacles with stinging cells (nematocytes). Here, we investigate the mechanisms of tentacle regeneration, using the hydrozoan jellyfish Cladonema pacificum. We show that proliferative cells accumulate at the tentacle amputation site and form a blastema composed of cells with stem cell morphology. Nucleoside pulse-chase experiments indicate that most repair-specific proliferative cells (RSPCs) in the blastema are distinct from resident stem cells. We further demonstrate that resident stem cells control nematogenesis and tentacle elongation during both homeostasis and regeneration as homeostatic stem cells, while RSPCs preferentially differentiate into epithelial cells in the newly formed tentacle, analogous to lineage-restricted stem/progenitor cells observed in salamander limbs. Taken together, our findings propose a regeneration mechanism that utilizes both resident homeostatic stem cells (RHSCs) and RSPCs, which in conjunction efficiently enable functional appendage regeneration, and provide novel insight into the diversification of blastema formation across animal evolution. Jellyfish are known for their remarkable regenerative abilities. This study shows that in the jellyfish Cladonema, following tentacle amputation, repair-specific proliferating cells robustly form a blastema with stem-like properties; these are distinct from resident stem cells and have a preference for differentiating into epithelial cells. [ABSTRACT FROM AUTHOR]

Caspases cleave over 1500 substrates in the human proteome in both lethal and non‐lethal scenarios. However, reports of the physiological consequences of substrate cleavage are limited. Additionally, the manner in which caspase cleaves only a subset of substrates in the non‐lethal scenario remains to be elucidated. BubR1, a spindle assembly checkpoint component, is a caspase substrate in humans, the physiological function of which remains unclear. Here, we found that caspases, especially Drice, cleave Drosophila BubR1 between the N‐terminal KEN box motif and C‐terminal kinase domain. By using proximity labelling, we found that Drice, but not Dcp‐1, is in proximity to BubR1, suggesting that protein proximity facilitates substrate preference. The cleaved fragments displayed altered subcellular localization and protein–protein interactions. Flies that harboured cleavage‐resistant BubR1 showed longer duration of BubR1 localization to the kinetochore upon colchicine treatment. Furthermore, these flies showed extended lifespan. Thus, we propose that the caspase‐mediated cleavage of BubR1 limits spindle assembly checkpoint and organismal lifespan. Our results highlight the importance of the individual analysis of substrates in vivo to determine the biological significance of caspase‐dependent non‐lethal cellular processes. [ABSTRACT FROM AUTHOR]

The mechanisms by which the innate immune system senses damage have been extensively explored in multicellular organisms. In Drosophila, various types of tissue damage, including epidermal injury, tumor formation, cell competition, and apoptosis deficiency, induce sterile activation of the Toll pathway, a process that requires the use of extracellular serine protease (SP) cascades. Upon infection, the SP Spätzle (Spz)-processing enzyme (SPE) cleaves and activates the Toll ligand Spz downstream of two paralogous SPs, Hayan and Persephone (Psh). However, upon tissue damage, it is not fully understood which SPs establish Spz activation cascades nor what damage-associated molecules can activate SPs. In this study, using newly generated uncleavable spz mutant flies, we revealed that Spz cleavage is required for the sterile activation of the Toll pathway, which is induced by apoptosis-deficient damage of wing epidermal cells in adult Drosophila. Proteomic analysis of hemolymph, followed by experiments with Drosophila Schneider 2 (S2) cells, revealed that among hemolymph SPs, both SPE and Melanization Protease 1 (MP1) have high capacities to cleave Spz. Additionally, in S2 cells, MP1 acts downstream of Hayan and Psh in a similar manner to SPE. Using genetic analysis, we found that the upstream SPs Hayan and Psh contributes to the sterile activation of the Toll pathway. While SPE/MP1 double mutants show more impairment of Toll activation upon infection than SPE single mutants, Toll activation is not eliminated in these apoptosis-deficient flies. This suggests that Hayan and Psh sense necrotic damage, inducing Spz cleavage by SPs other than SPE and MP1. Furthermore, hydrogen peroxide, a representative damage-associated molecule, activates the Psh-Spz cascade in S2 cells overexpressing Psh. Considering that reactive oxygen species (ROS) were detected in apoptosis-deficient wings, our findings highlight the importance of ROS as signaling molecules that induce the activation of SPs such as Psh in response to damage. Author summary: The innate immune system is conserved among multicellular organisms and has developed the mechanisms to recognize various dangers including infection and non-infectious damage. In Drosophila, the Toll pathway, an innate immune signaling pathway, is activated in response to tissue damage. Toll activation requires the use of serine protease (SP) cascades to activate the Toll ligand Spätzle. In this study, we explored SPs and damage-associated molecules involved in the sterile activation of the Toll pathway using a fly model, where apoptosis deficiency of wing epidermal cells causes damage. Genetic analyses demonstrated that paralogous SPs Hayan and Persephone contribute to Toll activation in apoptosis-deficient flies. Furthermore, we found that hydrogen peroxide, a representative damage-associated molecule, activates Persephone in cells and that apoptosis-deficient wings produce reactive oxygen species (ROS). Our findings highlight the importance of ROS in initiating the SP cascade for Toll activation in response to tissue damage. [ABSTRACT FROM AUTHOR]

Commensal microbes in animals have a profound impact on tissue homeostasis, stress resistance, and ageing. We previously showed in Drosophila melanogaster that Acetobacter persici is a member of the gut microbiota that promotes ageing and shortens fly lifespan. However, the molecular mechanism by which this specific bacterial species changes lifespan and physiology remains unclear. The difficulty in studying longevity using gnotobiotic flies is the high risk of contamination during ageing. To overcome this technical challenge, we used a bacteria-conditioned diet enriched with bacterial products and cell wall components. Here, we demonstrate that an A. persici-conditioned diet shortens lifespan and increases intestinal stem cell (ISC) proliferation. Feeding adult flies a diet conditioned with A. persici, but not with Lactiplantibacillus plantarum, can decrease lifespan but increase resistance to paraquat or oral infection of Pseudomonas entomophila, indicating that the bacterium alters the trade-off between lifespan and host defence. A transcriptomic analysis using fly intestine revealed that A. persici preferably induces antimicrobial peptides (AMPs), while L. plantarum upregulates amidase peptidoglycan recognition proteins (PGRPs). The specific induction of these Imd target genes by peptidoglycans from two bacterial species is due to the stimulation of the receptor PGRP-LC in the anterior midgut for AMPs or PGRP-LE from the posterior midgut for amidase PGRPs. Heat-killed A. persici also shortens lifespan and increases ISC proliferation via PGRP-LC, but it is not sufficient to alter the stress resistance. Our study emphasizes the significance of peptidoglycan specificity in determining the gut bacterial impact on healthspan. It also unveils the postbiotic effect of specific gut bacterial species, which turns flies into a "live fast, die young" lifestyle. Author summary: Microbiota plays a vital role in our health, but it can also have a negative impact on the lifespan of certain model organisms, such as the fruit fly Drosophila melanogaster. Despite its impact, the molecular mechanism behind how gut bacteria limits host lifespan remains unclear. In this study, we investigated the mechanism that one specific species of microbiota shortens lifespan and disrupts gut homeostasis of the aged flies, using a "fermented" fly diet. We found that the specific effects of this bacterium on fly healthspan were due to its capability of stimulating a specific receptor in the gut that recognizes peptidoglycan, a component of bacterial cell wall. Paradoxically, the same bacterium also increases stress resistance and defence against oral infection of a pathogen. Our study provides insight into the mechanisms underlying how certain members of the microbiota can lead to a "life fast, die young" lifestyle. [ABSTRACT FROM AUTHOR]