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Trehalose accumulation is commonly observed in extremophile organisms capable of surviving extended periods of complete desiccation and freezing. However, direct evidence linking the sugar to cryptobiotic survival in metazoans is generally lacking. Here, using the extremophile crustacean Artemia franciscana as a model, we test the anhydrobiotic and cryobiotic roles of trehalose through RNAi-mediated knockdown of its biosynthetic and transport pathways. Transcriptomic and genomic screening uncovered a major expansion of trehalose transporter (tret) genes in branchiopods with up to 27 copies in Artemia sp., but only a single trehalase enzyme (treh), and three trehalose-6-phosphate synthase (tps) genes containing fused synthase and phosphatase domains. Gene expression analysis determined that the tps and selected tret genes are expressed within the diapause-programmed oocytes and embryos, with independent RNAi-mediated knockdown of each reducing cyst trehalose accumulation by approximately 90% and 30%, respectively. Trehalose depletion in diapause cysts critically affects the anhydrobiotic and cryobiotic survival of embryos, a feature confirmed through diapause-termination experiments using either desiccation or H2O2 exposure. These latter data further uncovered the developmental reprogramming of carbohydrate metabolism to cope with low amounts of stored trehalose in the cysts. The findings demonstrate that diapause-induced biosynthesis and transport of trehalose are essential for A. franciscana anhydrobiosis.

The glial cells missing (GCM) genes were first discovered in Drosophila and encode transcription factors important for gliogenesis. In placental mammals, GCM1 regulates several genes that are important for early placenta development, while its paralogue GCM2 is important for parathyroid gland development. The egg-laying monotremes, which represent the most diverged extant mammalian lineage, undergo a short period of intrauterine development and form a simple placenta. To gain more insight into the evolution of GCM genes, we analysed the sequence, expression and genomic localization of GCM1 and GCM2 genes in the platypus and echidna. We found that the chromosomal localization of GCM1 changed after the divergence of therian mammals, coinciding with the evolution of a complex placenta. Expression analysis revealed the presence of GCM transcripts in male and female monotreme gonads, as well as expression of GCM1 in the female reproductive tract. GCM-binding sites in target genes associated with placental development in therian mammals were also present in the monotremes and the chicken. Together, this suggests that the role of GCM1 in reproduction emerged early in mammalian evolution.

Neuronal morphogenesis is regulated by intracellular transport and cytoskeletal dynamics. Kinesin superfamily proteins (KIFs), or kinesins, function as molecular motors for intracellular transport and as regulators of the microtubule cytoskeleton, making them essential for neuronal development. Caenorhabditis elegans has been widely used as a model organism to study neuronal morphogenesis. Due to the critical roles of kinesins in neuronal functions, numerous kinesin mutants, including unique gain-of-function mutants and temperature-sensitive mutants, have been identified through forward genetic screens in C. elegans. The availability of whole-genome knockout resources and CRISPR/Cas9 genome editing has further enabled precise genetic analysis, facilitating the modelling of human kinesin-related diseases in C. elegans. In this review, we discuss the functions of C. elegans kinesins in neuronal morphogenesis, focusing on their roles in neuronal transport and cytoskeletal regulations.

The specification of abdominal segments A5 to A9 depends on the expression of Abdominal-B (Abd-B), which is regulated by four infraabdominal domains: iab-5 through iab-8,9. Each iab domain contains an initiation element that determines its active state, along with enhancers responsible for tissue-specific activation of Abd-B. These iab domains function autonomously due to their flanking boundaries, Fab-6, Fab-7 and Fab-8, which both block crosstalk between adjacent iab domains (insulator function) and facilitate long-range interactions with the Abd-B promoter (bypass function). In inactive iab domains, enhancers are repressed by Polycomb group (PcG) proteins. Activation of the iab domains is driven by initiators, which are stimulated in a segment-specific manner by products of the gap and pair-rule genes during early embryogenesis. By creating truncations of the Fab-6 boundary, we identified that the bypass module is adjacent to CTCF binding sites and overlaps with sequences responsible for recruiting PcG proteins. Using genome editing, we demonstrated that the iab-5 and iab-6 initiators enhance the activity of the Fab-6 bypass module. Therefore, initiators represent a novel class of regulatory elements that control long-distance interactions between iab enhancers and Abd-B promoters.

Amphibia-infecting members of the genus Trypanosoma represent its most divergent lineage and exhibit remarkable morphological and genetic diversity. However, their detailed study has been hindered by pleomorphism, morphological convergence, multiple infections and scarcity of molecular data. By combining light microscopy with molecular phylogenetics, we investigated trypanosome diversity in a broad sample of frogs from Panama, a climatically stable tropical biodiversity hotspot. The uncovered diversity of trypanosomes parasitizing amphibians was exceptional, exceeding the host species richness twofold. Phylogenetic analyses revealed conspicuous ecological partitioning: distinct lineages were primarily associated with the arboreal hylids, with only rare host switches to understorey species, suggesting that vector feeding preferences structure the parasites' community. Notably, none of the identified haplotypes matched those from South America, underscoring geographic isolation as a driver of diversification. This study revealed a vast, previously undescribed diversity of trypanosomes that reflects the ecological and taxonomic breadth of their amphibian hosts. Given the ongoing global amphibian decline, documenting these parasitic communities is urgent. Our findings highlight how host ecology and biogeography shape parasite evolution, and offer a framework for future research in threatened tropical ecosystems.

Extracellular vesicles are small particles released by all cell types. Different extracellular vesicle isolation methods are widely used, yet none achieve an optimal balance between yield, purity and structural integrity. This study aimed to establish a comparative approach for evaluating different extracellular vesicle preparations using nanoparticle tracking analysis. A simple one-step assay relying on fluorescence-based nanoparticle tracking analysis was used to evaluate lectin binding to extracellular vesicles as a measure of possible changes in their surface glycosylation during various isolation methods. Seminal extracellular vesicles were isolated from normozoospermic men using ultracentrifugation alone-UC-sEVs-or combined with size exclusion chromatography-UC-SEC-sEVs-or microfiltration-UC-MF-sEVs. They were analysed based on their size and lectin-binding properties using wheat germ agglutinin and Ricinus communis agglutinin I. While total seminal extracellular vesicles and tetraspanin-positive seminal extracellular vesicles maintained similar size distributions across all isolates, lectin-positive seminal extracellular vesicles displayed a shift towards larger than 200 nm seminal extracellular vesicles in UC-SEC-sEVs and UC-MF-sEVs, as compared to UC-sEVs. The ratio of larger (>200 nm) to smaller (30-200 nm) lectin-positive sEVs was increased, particularly for wheat germ agglutinin in UC-MF-sEVs and Ricinus communis agglutinin I in UC-SEC-sEVs. These findings demonstrate that size exclusion chromatography and microfiltration combined with ultracentrifugation influence seminal extracellular vesicle surface glycosylation and alter lectin binding across extracellular vesicles of different sizes.

The morphogenetic protein DivIVA exhibits diverse functions across bacterial phyla. In Bacillota, DivIVA is primarily involved in cell division, whereas in Actinomycetota, it plays a central role in coordinating polar growth. Due to its essentiality in Actinomycetota, gaining insight into its structural functions is challenging. We studied truncated DivIVA proteins using a unique divIVA deletion mutant in cell wall-deficient Kitasatospora viridifaciens L-forms. DivIVA comprises an N-terminal domain consisting of a coiled-coil segment bearing a membrane-targeting structure at the N-terminal end, followed by an intercoil region, a larger coiled-coil and a C-terminal domain. Deleting either the intercoil or C-terminal region affected branching. We also created a minimized variant in which both were deleted simultaneously, retaining the N-terminal domain and the second coiled-coil. Expression of this variant caused severe growth defects. Cells showed increased hyphal width, thicker cell walls and frequent tip bursting. Finally, we successfully introduced chimeric DivIVA from the unicellular actinobacterium Mycolicibacterium smegmatis with the membrane targeting domain of K. viridifaciens DivIVA, demonstrating functional conservation within the phylum. By contrast, chimeric DivIVA proteins from Bacillus subtilis could not support growth, underscoring that polar growth is encoded in Actinomycetota-specific amino acid motifs in the first and second coiled-coils. These findings enhance our understanding of the structure-function relationship for DivIVA and present new opportunities to study polar growth.

Several types of cancer and some auto-immune diseases are linked to Epstein-Barr virus (EBV) infection. In addition to viral proteins, EBV expresses many functional RNAs to influence cell biology, avoiding the immune responses that would be targeted to viral proteins. Some of these RNAs are also exported from the EBV-infected cells into surrounding cells, including those involved in immune surveillance of the EBV infected cells. EBER1 is the most abundant of these EBV RNAs and recent progress in understanding EBER1 mechanisms might make it a drug target for some diseases associated with EBV.

Although the difference between the characteristics of fast and slow muscles has been extensively studied, it is still not fully understood. Here, we propose that nicotinic acetylcholine receptors (AChRs) expressed in slow muscles of zebrafish have high Ca²⁺ permeability compared to that of AChRs of fast muscles. To analyse the significance of the Ca²⁺ influx through AChRs in slow muscles, we generated a transgenic (Tg) zebrafish line that expresses Ca²⁺-impermeable AChRs in its slow muscles. The locomotor activities of the Tg zebrafish were markedly decreased at 1-3 days post-fertilization (dpf) compared to those of zebrafish expressing Ca²⁺-permeable AChRs in their slow muscles. Ca²⁺ imaging suggested that Ca²⁺ influx via AChRs is crucial for the Ca²⁺ response during muscle contraction in 2 dpf larvae, as slow muscle cells of the Tg line lacked a sustained Ca²⁺ response. Furthermore, we found that slow muscles of the Tg line became thinner compared to those expressing Ca²⁺-permeable AChRs. These short Ca²⁺ responses and thinner slow muscles may have induced locomotion impairment in the Tg line. These results suggested the physiological roles of the Ca²⁺ influx through AChRs in slow muscles and provided new insights into the characterization of fast and slow muscles.