Probing extracellular Sonic hedgehog in neurons
The bioactivity of Sonic hedgehog (Shh) depends on specific lipid modifications; a palmitate at its N-terminus and a cholesterol at its C-terminus. This dual-lipid modification makes Shh molecules lipophilic, which prevents them from diffusing freely in extracellular space. Multiple lines of evidenc...
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2016-08-01
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doaj-c9508cd94ee7445f916a2ebe217bafb32021-06-02T19:22:37ZengThe Company of BiologistsBiology Open2046-63902016-08-01581086109210.1242/bio.019422019422Probing extracellular Sonic hedgehog in neuronsErez Eitan0Ronald S. Petralia1Ya-Xian Wang2Fred E. Indig3Mark P. Mattson4Pamela J. Yao5 Laboratory of Neurosciences, National Institute on Aging Intramural Research Program, Baltimore, MD 21224, USA Advanced Imaging Core, NIDCD/NIH, Bethesda, MD 20892, USA Advanced Imaging Core, NIDCD/NIH, Bethesda, MD 20892, USA Confocal Imaging Facility, Laboratory of Clinical Investigation, National Institute on Aging Intramural Research Program, Baltimore, MD 21224, USA Laboratory of Neurosciences, National Institute on Aging Intramural Research Program, Baltimore, MD 21224, USA Laboratory of Neurosciences, National Institute on Aging Intramural Research Program, Baltimore, MD 21224, USA The bioactivity of Sonic hedgehog (Shh) depends on specific lipid modifications; a palmitate at its N-terminus and a cholesterol at its C-terminus. This dual-lipid modification makes Shh molecules lipophilic, which prevents them from diffusing freely in extracellular space. Multiple lines of evidence indicate that Shh proteins are carried by various forms of extracellular vesicles (EVs). It also has been shown, for instance, that in some tissues Shh proteins are transported to neighboring cells directly via filopodia. We have previously reported that Shh proteins are expressed in hippocampal neurons. In this study we show that, in the hippocampus and cerebellum of postnatal day (P)2 rats, Shh is mostly found near or on the membrane surface of small neurites or filopodia. We also examined cultured hippocampal neurons where we observed noticeable and widespread Shh-immunolabeled vesicles located outside neurons. Through immunoelectron microscopy and biochemical analysis, we find Shh-containing EVs with a wide range of sizes. Unlike robust Shh activity in EVs isolated from cells overexpressing an N-terminal Shh fragment construct, we did not detect measurable Shh activity in EVs purified from the medium of cultured hippocampal neurons. These results suggest the complexity of the transcellular Shh signaling mechanisms in neurons.http://bio.biologists.org/content/5/8/1086Sonic hedgehogHippocampal neuronsExtracellular vesicleFilopodia |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Erez Eitan Ronald S. Petralia Ya-Xian Wang Fred E. Indig Mark P. Mattson Pamela J. Yao |
spellingShingle |
Erez Eitan Ronald S. Petralia Ya-Xian Wang Fred E. Indig Mark P. Mattson Pamela J. Yao Probing extracellular Sonic hedgehog in neurons Biology Open Sonic hedgehog Hippocampal neurons Extracellular vesicle Filopodia |
author_facet |
Erez Eitan Ronald S. Petralia Ya-Xian Wang Fred E. Indig Mark P. Mattson Pamela J. Yao |
author_sort |
Erez Eitan |
title |
Probing extracellular Sonic hedgehog in neurons |
title_short |
Probing extracellular Sonic hedgehog in neurons |
title_full |
Probing extracellular Sonic hedgehog in neurons |
title_fullStr |
Probing extracellular Sonic hedgehog in neurons |
title_full_unstemmed |
Probing extracellular Sonic hedgehog in neurons |
title_sort |
probing extracellular sonic hedgehog in neurons |
publisher |
The Company of Biologists |
series |
Biology Open |
issn |
2046-6390 |
publishDate |
2016-08-01 |
description |
The bioactivity of Sonic hedgehog (Shh) depends on specific lipid modifications; a palmitate at its N-terminus and a cholesterol at its C-terminus. This dual-lipid modification makes Shh molecules lipophilic, which prevents them from diffusing freely in extracellular space. Multiple lines of evidence indicate that Shh proteins are carried by various forms of extracellular vesicles (EVs). It also has been shown, for instance, that in some tissues Shh proteins are transported to neighboring cells directly via filopodia. We have previously reported that Shh proteins are expressed in hippocampal neurons. In this study we show that, in the hippocampus and cerebellum of postnatal day (P)2 rats, Shh is mostly found near or on the membrane surface of small neurites or filopodia. We also examined cultured hippocampal neurons where we observed noticeable and widespread Shh-immunolabeled vesicles located outside neurons. Through immunoelectron microscopy and biochemical analysis, we find Shh-containing EVs with a wide range of sizes. Unlike robust Shh activity in EVs isolated from cells overexpressing an N-terminal Shh fragment construct, we did not detect measurable Shh activity in EVs purified from the medium of cultured hippocampal neurons. These results suggest the complexity of the transcellular Shh signaling mechanisms in neurons. |
topic |
Sonic hedgehog Hippocampal neurons Extracellular vesicle Filopodia |
url |
http://bio.biologists.org/content/5/8/1086 |
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