Quantitative Super-Resolution Microscopy of the Mammalian Glycocalyx.
Leonhard Möckl1, Kayvon Pedram1, Anish R Roy1, Venkatesh Krishnan2, Anna-Karin Gustavsson3, Oliver Dorigo2, Carolyn R Bertozzi4, W E Moerner5
- Department of Chemistry, Stanford University, Stanford, CA 94305, USA.
- Department of Obstetrics and Gynecology, Division of Gynecologic Oncology, Stanford University School of Medicine, Stanford, CA 94305, USA.
- Department of Chemistry, Stanford University, Stanford, CA 94305, USA; Department of Biosciences and Nutrition, Karolinska Institutet, 171 77 Stockholm, Sweden.
- Department of Chemistry, Stanford University, Stanford, CA 94305, USA; Howard Hughes Medical Institute, Stanford, CA 94305, USA. Electronic address: bertozzi@stanford.edu.
- Department of Chemistry, Stanford University, Stanford, CA 94305, USA. Electronic address: wmoerner@stanford.edu.
Abstract
The mammalian glycocalyx is a heavily glycosylated extramembrane compartment found on nearly every cell. Despite its relevance in both health and disease, studies of the glycocalyx remain hampered by a paucity of methods to spatially classify its components. We combine metabolic labeling, bioorthogonal chemistry, and super-resolution localization microscopy to image two constituents of cell-surface glycans, N-acetylgalactosamine (GalNAc) and sialic acid, with 10-20 nm precision in 2D and 3D. This approach enables two measurements: glycocalyx height and the distribution of individual sugars distal from the membrane. These measurements show that the glycocalyx exhibits nanoscale organization on both cell lines and primary human tumor cells. Additionally, we observe enhanced glycocalyx height in response to epithelial-to-mesenchymal transition and to oncogenic KRAS activation. In the latter case, we trace increased height to an effector gene, GALNT7. These data highlight the power of advanced imaging methods to provide molecular and functional insights into glycocalyx biology.
Presented by Leonhard Möckl