bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.02.18.638836

Complementary Volume Electron Microscopy-based approaches reveal ultrastructural changes in germline intercellular bridges of D. melanogaster

Kolotuev, I.·Williams, A.·Kizilyaprak, C.·Pellegrino, S.·Lewellyn, L.
View original source ↗Export metadata ↓

Abstract

Intercellular bridges are essential to connect developing germline cells. The Drosophila melanogaster egg chamber is a powerful model system to study germline intercellular bridges, or ring canals (RCs). RCs connect the developing oocyte to supporting nurse cells, and defects in their stability or growth lead to infertility. Despite their importance, it has been technically difficult to use electron microscopy-based approaches to monitor changes in RC structure during oogenesis. Here, we describe the application of a complementary set of volume EM-based approaches to visualize ultrastructural changes in the germline RCs. The combination of array tomography (AT) and focused ion beam (FIB) scanning electron microscopy (SEM) has allowed us to gain insight into previously unappreciated aspects of RC structure. We were able to quantify differences in RC size and thickness within and between germ cell clusters at different developmental stages. Within a cluster, RC size correlates with lineage; the largest RCs were formed during the first division, and the smallest RCs were formed during the fourth mitotic division. We observed the formation of membrane interdigitations in the vicinity of RCs much earlier than previously reported, and reconstruction of a RC from a mid-stage EC provided insight into the 3D orientation of these extensive cell-cell contacts. Our imaging also revealed a novel membrane structure that appeared to line the interior of the RC lumen. Although the focus was on ultrastructural changes in the germline RCs, our dataset contains valuable details of additional cell types and structures, including the fusome, the germline stem cells and their niche, and the migrating border cells. This imaging framework could be applied to other tissues or samples that face similar technical challenges, where the small structure of interest is located within a large sample volume.

Explore related subjects

developmental biologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Kolotuev, I., Williams, A., Kizilyaprak, C., Pellegrino, S., Lewellyn, L.. 2025-02-23. Complementary Volume Electron Microscopy-based approaches reveal ultrastructural changes in germline intercellular bridges of D. melanogaster. https://doi.org/10.1101/2025.02.18.638836

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Hyaluronic acid dynamics are regulated by neural crest cells and NEDD4 in the developing cardiac outflow tract valves

The extracellular matrix (ECM) is essential for cardiac valve development, with hyaluronic acid (HA) playing a critical role in cardiac cushion formation. While HA is required for atrioventricular valve development, its role in semilunar valve formation remains poorly understood. The ubiquitin ligase NEDD4 is essential for cardiac outflow tract development, particularly within cardiac neural crest cells, however its role in semilunar valve development has been unexplored. Here, we demonstrate that neural crest-specific deletion of NEDD4 in mice disrupts outflow tract valve development, resulting in abnormal leaflet number and morphology. Analysis of valve ECM revealed a specific reduction in HA within the neural crest-derived valve mesenchyme, while other ECM components such as fibronectin and versican remained unaffected. In contrast, deletion of NEDD4 in endocardium-derived valve mesenchyme did not alter HA abundance. Using NEDD4-deficient neural crest cells, we further demonstrate a cell-autonomous requirement for NEDD4 in regulating HA abundance, with in vitro evidence supporting impaired HA synthesis rather than increased turnover of HA in the ECM. Our findings identify a previously unrecognised role for NEDD4 in regulating HA synthesis in cardiac neural crest cells, revealing a cell-type-specific mechanism governing semilunar valve development and highlighting distinct cellular requirements for ECM regulation across cardiac valve populations.

developmental biology↗
bioRxiv2026

Endogenous Mechanical Reporters Encode the Global Mechanical Landscape of Morphogenesis

Developmental systems usually comprise multiple genetically programmed morphogenetic events within a mechanically continuous environment, yet how individual processes interact and contribute to the overall mechanical landscape remains unknown. Here, we introduce the concept of mechanical reporter: endogenous tissues function as in situ probes of the embedded mechanical landscape. We exemplified cephalic furrow (CF) migration as a mechanical reporter in gastrulating Drosophila. Using optogenetics and genetics, we revealed that CF displacement reflects the combined effect of both force and tissue mechanical properties. By incorporating condition-specific mechanical landscape into a continuum minimal-response model, we developed a framework to quantitatively predict the reporter dynamics. Using different genetic mutants, we successfully decoded changes in the mechanical landscape from CF dynamics in gastrulating Drosophila. Overall, we established a predictive framework that uses CF dynamics to infer changes in the global mechanical landscape. Beyond this, the concept of mechanical reporter provides a general strategy for systematically understanding and decoding complex mechanical environments in living systems.

developmental biology↗
bioRxiv2026

Dissecting human embryonic patterning during organogenesis reveals a rostral midline progenitor population for mesoendoderm induction

Understanding how human organogenesis is accomplished via intricate spatiotemporal patterning of organ primordia requires spatial atlases at cellular resolution. However, existing human embryo spatial atlases are built using sequencing-based platforms such as Stereo-seq, which have limited spatial resolution and blurred anatomical boundaries, leading to misclassification across adjacent cells. Here, we integrate in situ imaging-based transcriptomics (Xenium) with transcriptome-wide spatial profiling (Visium-HD) of Carnegie stages 9-12 embryos to delineate the patterning of core embryonic tissues at single-cell resolution, including the brain, spinal cord, somites, neural crest, notochord, heart, and gut tube. A new finding enabled by spatial single-cell resolution is the identification of rostral midline progenitors (RMPs), a cell population co-expressing SOX2, TBXT, and OTX2. Transcriptionally, RMPs are distinct from both neuromesodermal progenitors (NMPs) and prechordal mesoderm, and in vivo clonal barcoding in mice indicates that RMPs share a common developmental origin with rostral neural ectoderm. To interrogate their functional significance, we leveraged our spatial transcriptome data to recapitulate RMP-like cells from human pluripotent stem cells and showed that these cells can induce epiblast-like cells to acquire mesendodermal and endodermal fates, while themselves adopting an ectodermal fate consistent with their in vivo counterparts. This work provides a 3D, stage-resolved molecular cell atlas of tissue patterning in human embryo that serves as a reference for in vitro models.

developmental biology↗