Biology Open最新文献

Reproduction in chondrichthyan fishes (sharks, rays, skates, and chimaeras) is generally assumed to be a long-term, energetically costly process, given their slow generation times. However, metabolic costs of reproduction remain poorly understood due to a lack of direct, non-lethal measurements. To address this, we investigated metabolic and physiological changes during oviparous reproduction in five female epaulette sharks (Hemiscyllium ocellatum). We tracked oxygen uptake rates - a proxy for metabolic rate - across a 3-week cycle, capturing data before, during, and after egg case encapsulation and oviposition. We also measured reproductive hormones (testosterone, 17β-estradiol, progesterone) and hematological parameters (hematocrit, hemoglobin concentration). Results revealed a positive but non-significant relationship between metabolic rate and body mass, and contrary to expectations, metabolic rate did not significantly change throughout the 19-day cycle. Hormone levels remained stable, except for a transient testosterone peak early in the cycle, and hematological parameters showed no significant variation. These findings tentatively suggest epaulette sharks maintain reproductive effort without marked increases in metabolic or physiological costs. Continued research under seasonal environmental variation could clarify reproductive energetics in chondrichthyans further. This study provides the first direct measurement of metabolic effects of oviparous reproduction in chondrichthyans, challenging assumptions about energetic demands in this taxon.

The size of the nucleus is tightly coordinated with cell size across eukaryotes, yet the physiological significance of maintaining proper nuclear dimensions remains poorly understood. Here, we investigate how nuclear size dysregulation resulting from perturbed nucleocytoplasmic transport affects mitotic fidelity in Schizosaccharomyces pombe. Overexpression of a GFP-tagged nuclear export signal (NES-GFP) induced nuclear expansion, leading to severe growth defects and frequent errors in chromosome segregation during mitosis. Live-cell imaging revealed that enlarged nuclei underwent delayed mitotic progression and abnormal nuclear division. Strikingly, genetic suppression of nuclear expansion alleviated these defects, whereas enhancement of nuclear size exacerbated them. Together, these findings suggest that maintaining proper nuclear dimensions contributes to accurate chromosome segregation, although additional effects of NES-GFP overproduction and other factors influencing nuclear size should be further examined.

Great reed warblers (Acrocephalus arundinaceus) have become an important species for understanding long-distance avian migration, yet the genetic basis of their migratory timing remains unknown. While previous studies have identified candidate genes influencing migration timing in other species, their role in great reed warblers remains unexplored. Additionally, it is unclear whether the genetic basis of migratory timing differs between spring and autumn migrations. This study aims to uncover genetic factors influencing migration timing, providing insights into the evolutionary and ecological processes shaping long-distance migration. We conducted pooled whole-genome sequencing representing four great reed warbler migratory chronotypes: early spring, late spring, early autumn, and late autumn. By comparing FST and allele frequency differences, we determined that the spring migration had a larger genetic contribution than the autumn migration; however, the effect sizes were small (0.03 and 0.001, respectively). When comparing the early and late spring pools, we identified 93 candidate genes enriched for functions related to lipid hydrolysis that putatively influence great reed warbler migratory behavior. Our results provide insight into the genetic differentiation underlying migratory timing in great reed warblers, which is crucial for predicting how they will adapt to shifting environmental conditions due to climate change and habitat loss.

Creating hypomorphic mutations is crucial to study gene function in vivo, especially when null mutations result in (embryonic) lethality. This applies to enzymes involved in glycosylation that, when mutated in human patients, cause the disease congenital disorders of glycosylation (CDG). In order to resemble patient condition, it would be ideal to acutely modulate the proteins in question to directly interfere with protein levels of such essential enzymes. These methods offer to establish pathogenic enzyme levels resembling net enzyme activity reported in patients suffering from CDG, with phosphomannomutase 2-CDG (PMM2-CDG) as the most common form. We established an auxin-inducible acute protein knockdown system for the use in the teleost fish medaka (Oryzias latipes) by combining an improved degron (AID2) technology with an mAID-nanobody targeting endogenously GFP-tagged Pmm2 protein. We generated a fishline expressing a functional Pmm2-GFP fusion protein, by single copy integration of GFP into the pmm2 locus. Upon induction, the degron system efficiently reduced Pmm2-GFP levels and enzyme activity, recapitulating the activity level of the hypomorphic mutations associated with PMM2-CDG in patients. This broadly applicable approach enables the investigation of CDG disease mechanisms during early embryonic development through reduction of protein abundance, mimicking hypomorphic mutations and thus substantially expanding the range of the genetic toolbox.

Transposon silencing is essential for germline development. In Drosophila oogenesis, DNA damage caused by transposon activation affects microtubule-dependent mRNA localisation in the oocyte and impairs embryonic axes formation. Our previous EMS mutagenesis screen for gurken mRNA mislocalisation on chromosome 3L of Drosophila melanogaster identified several piRNA pathway mutants ( Hayashi et al., 2014). Here, we report the screen for chromosome 3R. We identified ten mutation groups disrupting gurken mRNA localisation and other mutations affecting different aspects of oogenesis. We found that mutations in karyopherin-β3 affect localisation and translation of gurken mRNA in a transposon silencing-independent manner. Characterisation of the new mutation in vreteno revealed that the piRNA pathway is essential for the basal stalk development, the process of holding the ovariole and encapsulating the first egg chambers. Females transheterozygous for vreteno and armitage mutations both showed abnormal basal stalks, defective egg chamber formation and loss of germline cells. We also found that the mutation in the Zinc Finger motif of Spindle-E shows a hypomorphic transposon activation phenotype, consistent with the previous study ( Ott et al., 2014). Further characterisation showed that the Zinc Finger is required for robust ping-pong piRNA biogenesis and the nuage localisation of AGO3, but not of Aubergine, suggesting that it is involved in a specific step of ping-pong biogenesis.

Genomic studies of neurodevelopmental disorders (NDDs) have identified several relevant genomic variants. EBF3 is a gene with an excess of protein-coding de novo variants and underlies Hypotonia, Ataxia, and Delayed Development Syndrome. We previously identified noncoding de novo variants in an enhancer of EBF3 and further found enrichment of deletions of this enhancer in NDDs. In this study, we generated a novel mouse line that deletes the highly conserved, orthologous mouse region within the Rr169617 regulatory region, and characterized the molecular and phenotypic aspects of this mouse model. We found a deviation from Mendelian expectation (P=0.02) with significant depletion of the deletion allele (P=5.8×10-4). Rr169617+/- mice had a reduction of Ebf3 expression by 10% and Rr169617-/- mice had a reduction by 20%. Differential expression analyses in E12.5 forebrain, midbrain, and hindbrain in Rr169617+/+ versus Rr169617-/- mice identified dysregulated genes including histone and brain development related genes. A priori phenotyping analysis (open field, hole board and light/dark transition) identified sex-specific differences in mobility only for Rr169617-/- mice across multiple behavioral assays with Rr169617-/- males less mobile than Rr169617-/- females. Furthermore, both sexes when homozygous for the enhancer deletion displayed body composition differences when compared to wildtype mice. Overall, we show that deletion within Rr169617 reduces expression of Ebf3 and results in phenotypic outcomes consistent with potential sex specific behavioral differences.

Transcription factors (TFs) regulate cell differentiation in multicellular organisms and were mostly identified by forward genetics in model organisms. However, genomes contain several-fold more TFs without known roles. To classify these orphans, we investigated conservation, cell-type specificity and temporal expression of the ∼290 TFs of Dictyostelia amoebas, which aggregate when starved to form migrating slugs and fruiting bodies consisting of spores and three somatic cell types. Here we deleted seven somatically expressed TF genes and found that four knock-outs were developmentally defective. ariA- lost slug migration and robust fruiting body formation. gtaJ- skipped slug migration and directly developed aggregates into robust fruiting bodies. mybAA- formed multi-tipped aggregates, defective slugs and fruiting bodies with few spores. Hierarchical clustering of the expression profiles of mybAA and 45 other multi-tip suppressing genes grouped mybAA with seven autophagy genes, with similar developmental defects as mybAA-, suggesting that mybAA induces autophagy gene expression. mybM- slugs poorly migrated and fruiting bodies had kinked, rough stalks, but normally expressed cell-type marker genes, indicating defective morphogenesis. Overall, transcriptomics informed TF selection proved useful for gene function discovery.

Rod photoreceptor stability is critical for retinal health and lifelong vision. Rhodopsin (Rho) trafficking is essential for rod homeostasis, as its mislocalization precedes rod cell death in inherited retinal disorders such as retinitis pigmentosa. Despite its importance, the molecular mechanisms of Rho trafficking in mammalian rods remain largely undefined. We investigated Rho's subcellular organization in the mammalian rod Golgi complex. We utilized STORM and structured illumination microscopy super-resolution imaging to map Golgi proteins with Rho in mouse and macaque rods. Our analysis found that a large proportion of Rho in this subcellular region colocalizes with Rab6a in the trans-Golgi. To functionally test this interaction, we utilized a dominant-negative Rab6a mutant in HEK293T cells and mouse rods. The mutant significantly inhibits Rho secretion in cell culture, causing intracellular retention. In mouse rods, the mutant similarly causes significant trans-Golgi Rho retention; however, a majority of Rho protein still escaped the Golgi and reached the outer segment. Together, these findings uncover critical new subcellular details about Rho organization at the Golgi and establish a role for Rab6a as a regulator of Rho protein release from the trans-Golgi in mammalian rods. Our results provide critical insight into the protein trafficking mechanisms essential for long-term photoreceptor health.

Collagen is the most abundant protein in the human body, providing structural stability to connective tissues. It organises and interacts with other proteins to form a complex extracellular matrix (ECM), with loss of collagen in the ECM seen in diseases such as osteoarthritis and osteoporosis. As collagen, and other ECM components, are atypically large proteins, they require specific endoplasmic reticulum (ER) export machinery. A key player in the export of procollagen from the ER is the MIA3 gene product, TANGO1. We introduced mutations to both tango1 isoforms in zebrafish independently to understand the importance of the previously unexplored short isoform in zebrafish development and tissue homeostasis. We show that the long isoform of tango1 (tango1L) is mostly able to compensate for loss of the short isoform (tango1S) in larvae. However, non-collagenous components of the ECM (such as proteoglycans) were disrupted during development, leading to abnormal matrix patterning, visible by electron microscopy. Adult tango1S zebrafish show altered spinal morphology and changes to intervertebral discs, suggesting that tango1S plays a role in skeletal patterning and homeostasis that is independent of the long isoform.

Juvenile pikeperch (Sander lucioperca) undergo several ontogenetic shifts, the timing of which determines the survival of their first winter. The shift from planktivory to a more active piscivorous phenotype involves moving from pelagic to demersal habitat with more stimuli and hence potential brain functional reorganizations. During two consecutive years, we collected planktivores and piscivores with different body sizes between the years, recording distinct stages relative to the shift, and analyzed their whole-brain transcriptomes in an ecological context. We identified a distinct non-overlapping group of transcription factors (TFs) significantly upregulated in each phenotype: TFs upregulated in planktivores correspond to initial establishment of brain regions and overall architecture; TFs upregulated in piscivores correspond to the refinement of neurons and the formation of specific neuronal circuits. The planktivores independently of body size were characterized by interconnected activity of two TFs, fosab and junba. Gene set enrichment revealed extracellular matrix and collagen-related transcripts in piscivores from both years. A high activity of solute carrier (Slc) transporters was identified in the smaller-bodied piscivores. The neurotranscriptomics results reflected differences in body size and matched with ecological data and survival rates. The brain regulome indicated that body size differences translate into the specific gene activity of juvenile pikeperch.