{"title":"Application of STEM tomography to investigate smooth ER morphology under stress conditions","authors":"V. Heinz, R. Rachel, C. Ziegler","doi":"10.1111/jmi.70020","DOIUrl":null,"url":null,"abstract":"<p>The endoplasmic reticulum (ER) is a highly dynamic organelle that undergoes significant morphological alterations in response to cellular stress. While conventional transmission electron microscopy (TEM) has provided valuable insights into these changes, such as the formation of crystalloid-ER and ER whorls, obtaining comprehensive three-dimensional (3D) information on these large structures within their cellular context has remained a challenge. To overcome these limitations, this study introduces an innovative application of dual-axis scanning transmission electron microscopy (STEM) tomography to investigate ER morphology under stress conditions in human embryonic kidney (HEK) cells overexpressing the cation channel polycystin-2 (PC-2). Benefitting from high-resolution, increased depth-of-focus, and reduced aberrations, STEM tomography enabled the detailed 3D reconstruction of large cellular subvolumes, providing unprecedented views of stress-induced ER structures. Our findings reveal distinct ultrastructural details of both crystalloid-ER and ER whorls. Crystalloid-ER exhibited a tubular architecture with potential interconnectedness, while ER whorls displayed a lamellar organisation and distinct membrane curvature. We observed the co-occurrence of these distinct smooth ER (sER) morphotypes within the same cell, yet they remained spatially separated, suggesting potential functional specialisation. Furthermore, we identified direct membrane contacts in mixed morphotypes, hinting at a shared origin or dynamic relationship between these structures. The study also elucidated the interactions of these organised smooth ER (OSER) structures with other organelles, such as mitochondria (MAM sites) and vesicles. In summary, the presented ultra-structural insights have a significant impact on our understanding of stress-related ER morphology changes. The ability to visualise the intricate 3D architecture and spatial relationships of these structures provides novel perspectives on the ER's adaptive responses to stress, including potential roles in lipid and protein biosynthesis and intracellular communication. These findings underscore the power of dual-axis STEM tomography for elucidating complex organellar organisation and dynamics in their native cellular context.</p>","PeriodicalId":16484,"journal":{"name":"Journal of microscopy","volume":"299 3","pages":"228-241"},"PeriodicalIF":1.9000,"publicationDate":"2025-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1111/jmi.70020","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Journal of microscopy","FirstCategoryId":"5","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1111/jmi.70020","RegionNum":4,"RegionCategory":"工程技术","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"MICROSCOPY","Score":null,"Total":0}
引用次数: 0
Abstract
The endoplasmic reticulum (ER) is a highly dynamic organelle that undergoes significant morphological alterations in response to cellular stress. While conventional transmission electron microscopy (TEM) has provided valuable insights into these changes, such as the formation of crystalloid-ER and ER whorls, obtaining comprehensive three-dimensional (3D) information on these large structures within their cellular context has remained a challenge. To overcome these limitations, this study introduces an innovative application of dual-axis scanning transmission electron microscopy (STEM) tomography to investigate ER morphology under stress conditions in human embryonic kidney (HEK) cells overexpressing the cation channel polycystin-2 (PC-2). Benefitting from high-resolution, increased depth-of-focus, and reduced aberrations, STEM tomography enabled the detailed 3D reconstruction of large cellular subvolumes, providing unprecedented views of stress-induced ER structures. Our findings reveal distinct ultrastructural details of both crystalloid-ER and ER whorls. Crystalloid-ER exhibited a tubular architecture with potential interconnectedness, while ER whorls displayed a lamellar organisation and distinct membrane curvature. We observed the co-occurrence of these distinct smooth ER (sER) morphotypes within the same cell, yet they remained spatially separated, suggesting potential functional specialisation. Furthermore, we identified direct membrane contacts in mixed morphotypes, hinting at a shared origin or dynamic relationship between these structures. The study also elucidated the interactions of these organised smooth ER (OSER) structures with other organelles, such as mitochondria (MAM sites) and vesicles. In summary, the presented ultra-structural insights have a significant impact on our understanding of stress-related ER morphology changes. The ability to visualise the intricate 3D architecture and spatial relationships of these structures provides novel perspectives on the ER's adaptive responses to stress, including potential roles in lipid and protein biosynthesis and intracellular communication. These findings underscore the power of dual-axis STEM tomography for elucidating complex organellar organisation and dynamics in their native cellular context.
期刊介绍:
The Journal of Microscopy is the oldest journal dedicated to the science of microscopy and the only peer-reviewed publication of the Royal Microscopical Society. It publishes papers that report on the very latest developments in microscopy such as advances in microscopy techniques or novel areas of application. The Journal does not seek to publish routine applications of microscopy or specimen preparation even though the submission may otherwise have a high scientific merit.
The scope covers research in the physical and biological sciences and covers imaging methods using light, electrons, X-rays and other radiations as well as atomic force and near field techniques. Interdisciplinary research is welcome. Papers pertaining to microscopy are also welcomed on optical theory, spectroscopy, novel specimen preparation and manipulation methods and image recording, processing and analysis including dynamic analysis of living specimens.
Publication types include full papers, hot topic fast tracked communications and review articles. Authors considering submitting a review article should contact the editorial office first.