Single-cell transcriptomics identifies new blood cell populations in Drosophila released at the onset of metamorphosis
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ABSTRACT:
PROVIDER: PRJNA893004 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR22007812.fastq.gz | Fastqsanger.gz | |||
| SRR22007813.fastq.gz | Fastqsanger.gz | |||
| SRR22007814.fastq.gz | Fastqsanger.gz | |||
| SRR22007815.fastq.gz | Fastqsanger.gz |
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Project description:Drosophila blood cells called hemocytes form an efficient barrier against infections and tissue damage. During metamorphosis, hemocytes undergo tremendous changes in their shape and behavior, preparing them for tissue clearance. Yet, the diversity and functional plasticity of pupal blood cells have not been explored. Here, we combine single-cell transcriptomics and high-resolution microscopy to dissect the heterogeneity and plasticity of pupal hemocytes. We identified undifferentiated and specified hemocytes with different molecular signatures associated with distinct functions such as antimicrobial, antifungal immune defense, cell adhesion or secretion. Strikingly, we identified a highly migratory and immuneresponsive pupal cell population expressing typical markers of the posterior signaling center (PSC), which is known to be an important niche in the larval lymph gland. PSC-like cells become restricted to the abdominal segments and are morphologically very distinct from typical Hemolectin (Hml)-positive plasmatocytes. G-TRACE lineage experiments further suggest that PSC-like cells can transdifferentiate to lamellocytes triggered by parasitoid wasp infestation. In summary, we present the first molecular description of pupal Drosophila blood cells, providing insights into blood cell functional diversification and plasticity during pupal metamorphosis.
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Project description:Drosophila blood cells called hemocytes form an efficient barrier against infections and tissue damage. During metamorphosis, hemocytes undergo tremendous changes in their shape and behavior, preparing them for tissue clearance. Yet, the diversity and functional plasticity of pupal blood cells have not been explored. Here, we combine single-cell transcriptomics and high-resolution microscopy to dissect the heterogeneity and plasticity of pupal hemocytes. We identified undifferentiated and specified hemocytes with different molecular signatures associated with distinct functions such as antimicrobial, antifungal immune defense, cell adhesion or secretion. Strikingly, we identified a highly migratory and immune-responsive pupal cell population expressing typical markers of the posterior signaling center (PSC), which is known to be an important niche in the larval lymph gland. PSC-like cells become restricted to the abdominal segments and are morphologically very distinct from typical Hemolectin (Hml)-positive plasmatocytes. G-TRACE lineage experiments further suggest that PSC-like cells can transdifferentiate to lamellocytes triggered by parasitoid wasp infestation. In summary, we present the first molecular description of pupal Drosophila blood cells, providing insights into blood cell functional diversification and plasticity during pupal metamorphosis.
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Project description:We used microarray analysis of RNA obtained from w1118 and w1118;sox14L1/sox14L1 animals staged at pupariation to identify genes regulated by Drosophila Sox14 at the onset of metamorphosis. RNA isolated from whole Drosophila w1118 and w1118; sox14L1/sox14L1 animals selected immediately after pupariation was hybridized on Affymetrix Drosophila Genome 2.0 Arrays. Each genotype was assayed independently three times with samples obtained from 10 prepupae.
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Project description:MicroRNAs (miRNAs) are endogenous RNA molecules that contribute to the regulation of gene expression post-transcriptionally. Recently, patterns of sequence evolution and evolutionary emergence of miRNAs have been analyzed in detail. However, the extent to which miRNA expression levels vary within and between species, the role of stabilizing selection versus other evolutionary mechanisms in shaping the temporal dynamics of these changes, and how this variation might unveil the interplay between miRNAs and mRNAs remain poorly understood. This is especially true during key developmental transitions involving miRNA regulation. Combining deep-sequencing with multi-species microarrays, we assayed miRNA expression levels immediately before (≥18BPF) and after (PF) the increase in steroid hormone ecdysone that triggers metamorphosis. We did so both at the intra- and interspecific levels in the Drosophila melanogaster species group. In contrast to their sequence conservation, ~25% of the analyzed miRNAs differed significantly in expression levels at ≥18BPF and/or PF across D. melanogaster strains. Additionally, ~33% showed modifications in their pattern of expression bias between developmental timepoints. Relative to the patterns of evolution, changes in miRNA abundance at PF accumulate linearly over evolutionary time while similar changes at ≥18BPF do not. Importantly, expression levels of ≥18BPF-enriched miRNAs are the most variable within and between species and therefore the least likely to evolve under stabilizing selection, especially as compared to non-developmentally enriched miRNAs. Overall, functional attributes such as expression ubiquity are more tightly associated with lower levels of miRNA expression polymorphism at PF than at ≥18BPF. Furthermore, ≥18BPF- and PF-enriched miRNAs showed opposite patterns of covariation in expression with mRNAs, which was found to denote the type of regulatory relationship between miRNA and mRNA. By focusing on the miRNA expression levels at the onset of metamorphosis, our results have unveiled contrasting functional patterns of evolutionary divergence during ontogeny.
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Project description:Coupling immunity and development is essential to ensure survival despite changing internal conditions in the organism. The metamorphosis of the fruit fly represents a striking example of drastic and systemic physiological changes that need to be integrated with the innate immune system. However, the mechanisms that coordinate development and immune cell activity in the transition from larva to adult in Drosophila remain to elucidate. The steroid hormone ecdysone is known to act as a key coordinator of metamorphosis. This hormone activates a nuclear receptor, the Ecdysone Receptor (EcR), which acts as a heterodimer with its partner Ultraspiracle (USP). Together, they activate the transcription of primary response genes, which in turn activate the transcription of a battery of late response genes. We have revealed that regulation of macrophage-like cells (hemocytes) by the steroid hormone ecdysone is essential for an effective innate immune response over metamorphosis. We have shown that in response to ecdysone signalling, hemocytes rapidly up regulate actin dynamics, motility and phagocytosis of apoptotic corpses, and acquire the ability to chemotax to damaged epithelia. Most importantly, individuals lacking ecdysone-activated hemocytes are defective in bacterial phagocytosis and are fatally susceptible to infection by bacteria ingested at larval stages, despite the normal systemic production of antimicrobial peptides. This decrease in survival is comparable to the one observed in pupae lacking immune cells altogether, indicating that ecdysone-regulation is essential to hemocyte immune functions and survival after infection. To better understand the ecdysone regulation of hemocyte activities, we have performed gene expression analysis. In order to identify the genes which expression change at the onset of metamorphosis, we have sorted hemocytes from 3rd instar larvae and from young prepupae and compared their gene expression. Moreover, and in order to identify which genes are regulated by the ecdysone signalling, we have used individuals expressing a dominant negative form of the Ecdysone Receptor specifically in their hemocytes. We have sorted hemocytes from 3rd instar and young prepupae of this genotype to compare their gene expression to the gene expression in larvae and prepupae from the control individuals.
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