Cross-modality synthesis of EM time series and live fluorescence imaging

  1. Anthony Santella
  2. Irina Kolotuev  Is a corresponding author
  3. Caroline Kizilyaprak
  4. Zhirong Bao  Is a corresponding author
  1. Memorial Sloan Kettering Cancer Center, United States
  2. University of Lausanne, Switzerland

Abstract

Analyses across imaging modalities allow the integration of complementary spatiotemporal information about brain development, structure and function. However, systematic atlasing across modalities is limited by challenges to effective image alignment. We combine highly spatially resolved electron microscopy (EM) and highly temporally resolved time-lapse fluorescence microscopy (FM) to examine the emergence of a complex nervous system in C. elegans embryogenesis. We generate an EM time series at four classic developmental stages and create a landmark-based co-optimization algorithm for cross-modality image alignment, which handles developmental heterochrony among datasets to achieve accurate single-cell level alignment. Synthesis based on the EM series and time-lapse FM series carrying different cell-specific markers reveals critical dynamic behaviors across scales of identifiable individual cells in the emergence of the primary neuropil, the nerve ring, as well as a major sensory organ, the amphid. Our study paves the way for systematic cross-modality data synthesis in C. elegans and demonstrates a powerful approach that may be applied broadly.

Data availability

EM data has been made available on WebKnossos, and source code made available on Github. Links are provided in MS and on project website.

The following data sets were generated

Article and author information

Author details

  1. Anthony Santella

    Molecular Cytology Core, Memorial Sloan Kettering Cancer Center, New York, United States
    Competing interests
    The authors declare that no competing interests exist.
  2. Irina Kolotuev

    Electron Microscopy Facility, University of Lausanne, Lausanne, Switzerland
    For correspondence
    irina.kolotueva@unil.ch
    Competing interests
    The authors declare that no competing interests exist.
  3. Caroline Kizilyaprak

    Electron Microscopy Facility, University of Lausanne, Lausanne, Switzerland
    Competing interests
    The authors declare that no competing interests exist.
  4. Zhirong Bao

    Developmental Biology Program, Memorial Sloan Kettering Cancer Center, New York, United States
    For correspondence
    baoz@mskcc.org
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2201-2745

Funding

National Institutes of Health (R01GM097576)

  • Zhirong Bao

National Institutes of Health (R24OD016474)

  • Zhirong Bao

National Institutes of Health (P30CA008748)

  • Zhirong Bao

Chan Zuckerberg Initiative (2019-198110 (5022))

  • Anthony Santella

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Oliver Hobert, Columbia University, Howard Hughes Medical Institute, United States

Publication history

  1. Preprint posted: February 13, 2022 (view preprint)
  2. Received: February 15, 2022
  3. Accepted: June 5, 2022
  4. Accepted Manuscript published: June 6, 2022 (version 1)
  5. Version of Record published: June 21, 2022 (version 2)

Copyright

© 2022, Santella et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 498
    Page views
  • 149
    Downloads
  • 0
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Anthony Santella
  2. Irina Kolotuev
  3. Caroline Kizilyaprak
  4. Zhirong Bao
(2022)
Cross-modality synthesis of EM time series and live fluorescence imaging
eLife 11:e77918.
https://doi.org/10.7554/eLife.77918

Further reading

    1. Developmental Biology
    Vincent Loreau, Renate Rees ... Dirk Görlich
    Tools and Resources Updated

    Measuring the positions and dynamics of proteins in intact tissues or whole animals is key to understanding protein function. However, to date, this is challenging, as the accessibility of large antibodies to dense tissues is often limited, and fluorescent proteins inserted close to a domain of interest may affect protein function. These complications apply in particular to muscle sarcomeres, arguably one of the most protein-dense assemblies in nature, which complicates studying sarcomere morphogenesis at molecular resolution. Here, we introduce a toolbox of nanobodies recognising various domains of the two Drosophila titin homologs, Sallimus and Projectin, as well as the key sarcomeric proteins Obscurin, α-Actinin, and Zasp52. We verified the superior labelling qualities of our nanobodies in muscle tissue as compared to antibodies. By applying our toolbox to larval muscles, we found a gigantic Sallimus isoform stretching more than 2 µm to bridge the sarcomeric I-band, while Projectin covers almost the entire myosin filaments in a polar orientation. Transgenic expression of tagged nanobodies confirmed their high affinity-binding without affecting target protein function. Finally, adding a degradation signal to anti-Sallimus nanobodies suggested that it is difficult to fully degrade Sallimus in mature sarcomeres; however, expression of these nanobodies caused developmental lethality. These results may inspire the generation of similar toolboxes for other large protein complexes in Drosophila or mammals.

    1. Developmental Biology
    2. Evolutionary Biology
    James W Truman, Jacquelyn Price ... Tzumin Lee
    Research Article

    We have focused on the mushroom bodies (MB) of Drosophila to determine how the larval circuits are formed and then transformed into those of the adult at metamorphosis. The adult MB has a core of thousands of Kenyon neurons; axons of the early-born g class form a medial lobe and those from later-born a'b' and ab classes form both medial and vertical lobes. The larva, however, hatches with only g neurons and forms a vertical lobe 'facsimile' using larval-specific axon branches from its g neurons. Computations by the MB involves MB input (MBINs) and output (MBONs) neurons that divide the lobes into discrete compartments. The larva has 10 such compartments while the adult MB has 16. We determined the fates of 28 of the 32 types of MBONs and MBINs that define the 10 larval compartments. Seven larval compartments are eventually incorporated into the adult MB; four of their larval MBINs die, while 12 MBINs/MBONs continue into the adult MB although with some compartment shifting. The remaining three larval compartments are larval specific, and their MBIN/MBONs trans-differentiate at metamorphosis, leaving the MB and joining other adult brain circuits. With the loss of the larval vertical lobe facsimile, the adult vertical lobes, are made de novo at metamorphosis, and their MBONs/MBINs are recruited from the pool of adult-specific cells. The combination of cell death, compartment shifting, trans-differentiation, and recruitment of new neurons result in no larval MBIN-MBON connections persisting through metamorphosis. At this simple level, then, we find no anatomical substrate for a memory trace persisting from larva to adult. For the neurons that trans-differentiate, our data suggest that their adult phenotypes are in line with their evolutionarily ancestral roles while their larval phenotypes are derived adaptations for the larval stage. These cells arise primarily within lineages that also produce permanent MBINs and MBONs, suggesting that larval specifying factors may allow information related to birth-order or sibling identity to be interpreted in a modified manner in these neurons to cause them to adopt a modified, larval phenotype. The loss of such factors at metamorphosis, though, would then allow these cells to adopt their ancestral phenotype in the adult system.