scholarly journals Expansion microscopy provides new insights into the cytoskeleton of malaria parasites including the conservation of a conoid

PLoS Biology ◽  
2021 ◽  
Vol 19 (3) ◽  
pp. e3001020
Author(s):  
Eloïse Bertiaux ◽  
Aurélia C. Balestra ◽  
Lorène Bournonville ◽  
Vincent Louvel ◽  
Bohumil Maco ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Crucial Role ◽  
Reduced Form ◽  
Malaria Parasites ◽  
Whole Cells ◽  
Apicomplexan Parasites ◽  
Dynamic Assembly

Malaria is caused by unicellular Plasmodium parasites. Plasmodium relies on diverse microtubule cytoskeletal structures for its reproduction, multiplication, and dissemination. Due to the small size of this parasite, its cytoskeleton has been primarily observable by electron microscopy (EM). Here, we demonstrate that the nanoscale cytoskeleton organisation is within reach using ultrastructure expansion microscopy (U-ExM). In developing microgametocytes, U-ExM allows monitoring the dynamic assembly of axonemes and concomitant tubulin polyglutamylation in whole cells. In the invasive merozoite and ookinete forms, U-ExM unveils the diversity across Plasmodium stages and species of the subpellicular microtubule arrays that confer cell rigidity. In ookinetes, we additionally identify an apical tubulin ring (ATR) that colocalises with markers of the conoid in related apicomplexan parasites. This tubulin-containing structure was presumed to be lost in Plasmodium despite its crucial role in motility and invasion in other apicomplexans. Here, U-ExM reveals that a divergent and considerably reduced form of the conoid is actually conserved in Plasmodium species.

scholarly journals Expansion Microscopy provides new insights into the cytoskeleton of malaria parasites including the conservation of a conoid

2020 ◽  
Author(s):  
Eloïse Bertiaux ◽  
Aurélia C Balestra ◽  
Lorène Bournonville ◽  
Mathieu Brochet ◽  
Paul Guichard ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Crucial Role ◽  
Reduced Form ◽  
Malaria Parasites ◽  
Whole Cells ◽  
Cytoskeleton Organization ◽  
Dynamic Assembly ◽  
Microtubule Structure

AbstractMalaria is caused by unicellular Plasmodium parasites. Plasmodium relies on diverse microtubule cytoskeletal structures for its reproduction, multiplication or dissemination. Due to the small size of this parasite, its cytoskeleton has been primarily observable by electron microscopy. Here, we demonstrate that the nanoscale cytoskeleton organization is within reach using ultrastructure expansion microscopy (U-ExM). In developing microgametocytes, U-ExM allows to monitor the dynamic assembly of axonemes and concomitant tubulin polyglutamylation in whole cells. In the invasive merozoite and ookinete forms, U-ExM unveils the subpellicular microtubule arrays that confer cell rigidity. In ookinete, we additionally identify an apical tubulin ring above the subpellicular microtubules that colocalises with markers of the conoid in related Apicomplexa parasites. This microtubule structure was presumed to be lost in Plasmodium despite its crucial role in both motility and invasion in most apicomplexans. Here, U-ExM reveals that a divergent and reduced form of the conoid is actually conserved in the Plasmodium genus.

Progressive Mucinous Histiocytosis: Importance of Electron Microscopy to Confirm Diagnosis

2010 ◽  
Vol 14 (5) ◽  
pp. 245-248 ◽  
Author(s):  
Iman Hemmati ◽  
W. Alastair McLeod ◽  
Richard I. Crawford
Keyword(s):  
Electron Microscopy ◽  
Light Microscopy ◽  
Clinical Examination ◽  
Crucial Role ◽  
Langerhans Cell Histiocytosis ◽  
Langerhans Cell ◽  
History Of ◽  
Characteristic Features ◽  
Immunohistochemical Studies

Background: Progressive mucinous histiocytosis (PMH) is a benign, non-Langerhans cell histiocytosis with characteristic ultrastructural features that can be used for diagnosis. Once an important tool in dermatologic diagnosis, electron microscopy has been largely replaced by immunohistochemistry and immunofluorescence techniques today. However, electron microscopy occasionally still plays a crucial role in the diagnosis of dermatologic conditions. We report a case of PMH as an example of a dermatologic disorder that requires electron microscopy for its diagnosis. Methods: A 60-year-old woman presented to our clinic with a history of small, sharply demarcated, skin-colored papules ranging from 2 to 5 mm in diameter distributed over the arms, forearms, and dorsal hands. The results of light microscopy, immunohistochemical studies, and clinical examination were inconclusive. Another biopsy for electron microscopy showed the characteristic features of PMH. Conclusion: This case demonstrates that a dermatopathology service still needs to have access to electron microscopy for diagnostic purposes to successfully diagnose a small number of rare conditions.

scholarly journals Electron microscopy snapshots of single particles from single cells

2018 ◽  
Vol 294 (5) ◽  
pp. 1602-1608 ◽  
Author(s):  
Xiunan Yi ◽  
Eric J. Verbeke ◽  
Yiran Chang ◽  
Daniel J. Dickinson ◽  
David W. Taylor
Keyword(s):  
Electron Microscopy ◽  
Single Particle ◽  
Protein Complexes ◽  
Signal To Noise Ratio ◽  
Single Cells ◽  
Particle Analysis ◽  
Biological Macromolecules ◽  
Single Particle Analysis ◽  
Whole Cells ◽  
Potential Applications

Cryo-electron microscopy (cryo-EM) has become an indispensable tool for structural studies of biological macromolecules. Two additional predominant methods are available for studying the architectures of multiprotein complexes: 1) single-particle analysis of purified samples and 2) tomography of whole cells or cell sections. The former can produce high-resolution structures but is limited to highly purified samples, whereas the latter can capture proteins in their native state but has a low signal-to-noise ratio and yields lower-resolution structures. Here, we present a simple, adaptable method combining microfluidic single-cell extraction with single-particle analysis by EM to characterize protein complexes from individual Caenorhabditis elegans embryos. Using this approach, we uncover 3D structures of ribosomes directly from single embryo extracts. Moreover, we investigated structural dynamics during development by counting the number of ribosomes per polysome in early and late embryos. This approach has significant potential applications for counting protein complexes and studying protein architectures from single cells in developmental, evolutionary, and disease contexts.

An open-access volume electron microscopy atlas of whole cells and tissues

Nature ◽  
2021 ◽  
Author(s):  
C. Shan Xu ◽  
Song Pang ◽  
Gleb Shtengel ◽  
Andreas Müller ◽  
Alex T. Ritter ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Open Access ◽  
Whole Cells ◽  
Volume Electron Microscopy

Phase Contrast Enhancement with Phase Plates in Biological Electron Microscopy

2010 ◽  
Vol 18 (4) ◽  
pp. 10-13 ◽  
Author(s):  
Kuniaki Nagayama ◽  
Radostin Danev ◽  
Hideki Shigematsu ◽  
Naoki Hosogi ◽  
Yoshiyuki Fukuda ◽  
...  
Keyword(s):  
Thin Film ◽  
Electron Microscopy ◽  
Phase Contrast ◽  
Phase Plate ◽  
Electron Loss ◽  
Whole Cells ◽  
Transmission Electron ◽  
Different Types ◽  
Transparent Objects ◽  
Phase Plates

Theoretically, transmission electron microscopy (TEM) is compatible with three different types of phase plate: thin-film, electrostatic, and magnetic. However, designing functional phase plates has been an arduous process that has suffered from unavoidable technical obstacles such as phase-plate charging and difficulties associated with micro-fabrication of electrostatic and magnetic phase plates. This review discusses phase-contrast schemes that allow visualization of transparent objects with high contrast. Next it deals with recent studies on biological applications ranging from proteins and viruses to whole cells. Finally, future prospects for overcoming the problem of phase-plate charging and for designing the next generation of phase-plates to solve the problem of electron loss inherent in thin-film phase plates are discussed.

scholarly journals ATG8 Is Essential Specifically for an Autophagy-Independent Function in Apicoplast Biogenesis in Blood-Stage Malaria Parasites

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Marta Walczak ◽  
Suresh M. Ganesan ◽  
Jacquin C. Niles ◽  
Ellen Yeh
Keyword(s):  
Protein Import ◽  
Blood Stage ◽  
Model Organisms ◽  
Secondary Endosymbiosis ◽  
Organelle Biogenesis ◽  
Independent Function ◽  
Malaria Parasites ◽  
Apicomplexan Parasites ◽  
Independent Functions ◽  
Parasite Replication

ABSTRACT Plasmodium parasites and related pathogens contain an essential nonphotosynthetic plastid organelle, the apicoplast, derived from secondary endosymbiosis. Intriguingly, a highly conserved eukaryotic protein, autophagy-related protein 8 (ATG8), has an autophagy-independent function in the apicoplast. Little is known about the novel apicoplast function of ATG8 and its importance in blood-stage Plasmodium falciparum. Using a P. falciparum strain in which ATG8 expression was conditionally regulated, we showed that P. falciparum ATG8 (PfATG8) is essential for parasite replication. Significantly, growth inhibition caused by the loss of PfATG8 was reversed by addition of isopentenyl pyrophosphate (IPP), which was previously shown to rescue apicoplast defects in P. falciparum. Parasites deficient in PfATG8, but whose growth was rescued by IPP, had lost their apicoplast. We designed a suite of functional assays, including a new fluorescence in situ hybridization (FISH) method for detection of the low-copy-number apicoplast genome, to interrogate specific steps in apicoplast biogenesis and detect apicoplast defects which preceded the block in parasite replication. Though protein import and membrane expansion of the apicoplast were unaffected, the apicoplast was not inherited by daughter parasites. Our findings demonstrate that, though multiple autophagy-dependent and independent functions have been proposed for PfATG8, only its role in apicoplast biogenesis is essential in blood-stage parasites. We propose that PfATG8 is required for fission or segregation of the apicoplast during parasite replication. IMPORTANCE Plasmodium parasites, which cause malaria, and related apicomplexan parasites are important human and veterinary pathogens. They are evolutionarily distant from traditional model organisms and possess a unique plastid organelle, the apicoplast, acquired by an unusual eukaryote-eukaryote endosymbiosis which established novel protein/lipid import and organelle inheritance pathways in the parasite cell. Though the apicoplast is essential for parasite survival in all stages of its life cycle, little is known about these novel biogenesis pathways. We show that malaria parasites have adapted a highly conserved protein required for macroautophagy in yeast and mammals to function specifically in apicoplast inheritance. Our finding elucidates a novel mechanism of organelle biogenesis, essential for pathogenesis, in this divergent branch of pathogenic eukaryotes.

scholarly journals Publisher Correction: An open-access volume electron microscopy atlas of whole cells and tissues

Nature ◽  
2021 ◽  
Author(s):  
C. Shan Xu ◽  
Song Pang ◽  
Gleb Shtengel ◽  
Andreas Müller ◽  
Alex T. Ritter ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Open Access ◽  
Whole Cells ◽  
Volume Electron Microscopy

scholarly journals Quantitative Seq-LGS – Genome-Wide Identification of Genetic Drivers of Multiple Phenotypes in Malaria Parasites

2016 ◽  
Author(s):  
Hussein M. Abkallo ◽  
Axel Martinelli ◽  
Megumi Inoue ◽  
Abhinay Ramaprasad ◽  
Phonepadith Xangsayarath ◽  
...  
Keyword(s):  
Mathematical Modeling ◽  
Genetic Determinants ◽  
Malaria Parasites ◽  
Apicomplexan Parasites ◽  
Mendelian Genetics ◽  
Sequencing Technologies ◽  
Multiple Phenotypes ◽  
Genome Wide ◽  
Sexual Recombination ◽  
Novel Alleles

ABSTRACTIdentifying the genetic determinants of phenotypes that impact on disease severity is of fundamental importance for the design of new interventions against malaria. Traditionally, such discovery has relied on labor-intensive approaches that require significant investments of time and resources. By combining Linkage Group Selection (LGS), quantitative whole genome population sequencing and a novel mathematical modeling approach (qSeq-LGS), we simultaneously identified multiple genes underlying two distinct phenotypes, identifying novel alleles for growth rate and strain specific immunity (SSI), while removing the need for traditionally required steps such as cloning, individual progeny phenotyping and marker generation. The detection of novel variants, verified by experimental phenotyping methods, demonstrates the remarkable potential of this approach for the identification of genes controlling selectable phenotypes in malaria and other apicomplexan parasites for which experimental genetic crosses are amenable.Significance StatementThis paper describes a powerful and rapid approach to the discovery of genes underlying medically important phenotypes in malaria parasites. This is crucial for the design of new drug and vaccine interventions. The approach bypasses the most time-consuming steps required by traditional genetic linkage studies and combines Mendelian genetics, quantitative deep sequencing technologies, genome analysis and mathematical modeling. We demonstrate that the approach can simultaneously identify multigenic drivers of multiple phenotypes, thus allowing complex genotyping studies to be conducted concomitantly. This methodology will be particularly useful for discovering the genetic basis of medically important phenotypes such as drug resistance and virulence in malaria and other apicomplexan parasites, as well as potentially in any organism undergoing sexual recombination.

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