Assembly and turnover of neurofilaments in growing axonal neurites
Neurofilaments (NFs) are thought to provide stability to the axon. We examined NF dynamics within axonal neurites of NB2a/d1 neuroblastoma by transient transfection with green fluorescent protein-tagged NF-heavy (GFP-H) under the control of a tetracycline-inducible promoter. Immunofluorescent and bi...
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doaj-a6601704d68a4eb3a8df900f60642ad42021-06-02T10:51:49ZengThe Company of BiologistsBiology Open2046-63902018-01-017110.1242/bio.028795028795Assembly and turnover of neurofilaments in growing axonal neuritesEdward F. Boumil0Rishel Vohnoutka1Sangmook Lee2Harish Pant3Thomas B. Shea4 Laboratory for Neuroscience, University of Massachusetts Lowell, Lowell, MA 01854, USA Laboratory for Neuroscience, University of Massachusetts Lowell, Lowell, MA 01854, USA Laboratory for Neuroscience, University of Massachusetts Lowell, Lowell, MA 01854, USA Cytoskeletal Protein Regulation Section, National Institute of Neurological Disorders and Stroke, National Institutes of Health, Bethesda, MD 20892 , USA Laboratory for Neuroscience, University of Massachusetts Lowell, Lowell, MA 01854, USA Neurofilaments (NFs) are thought to provide stability to the axon. We examined NF dynamics within axonal neurites of NB2a/d1 neuroblastoma by transient transfection with green fluorescent protein-tagged NF-heavy (GFP-H) under the control of a tetracycline-inducible promoter. Immunofluorescent and biochemical analyses demonstrated that GFP-H expressed early during neurite outgrowth associated with a population of centrally-situated, highly-phosphorylated crosslinked NFs along the length of axonal neurites (‘bundled NFs’). By contrast, GFP-H expressed after considerable neurite outgrowth displayed markedly reduced association with bundled NFs and was instead more evenly distributed throughout the axon. This differential localization was maintained for up to 2 weeks in culture. Once considerable neurite outgrowth had progressed, GFP that had previously associated with the NF bundle during early expression was irreversibly depleted by photobleaching. Cessation of expression allowed monitoring of NF turnover. GFP-H associated bundled NFs underwent slower decay than GFP-H associated with surrounding, less-phosphorylated NFs. Notably, GFP associated with bundled NFs underwent similar decay rates within the core and edges of this bundle. These results are consistent with previous demonstration of a resident NF population within axonal neurites, but suggest that this population is more dynamic than previously considered.http://bio.biologists.org/content/7/1/bio028795NeurofilamentCytoskeletonAxonAxogenesisAxonal transport |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Edward F. Boumil Rishel Vohnoutka Sangmook Lee Harish Pant Thomas B. Shea |
spellingShingle |
Edward F. Boumil Rishel Vohnoutka Sangmook Lee Harish Pant Thomas B. Shea Assembly and turnover of neurofilaments in growing axonal neurites Biology Open Neurofilament Cytoskeleton Axon Axogenesis Axonal transport |
author_facet |
Edward F. Boumil Rishel Vohnoutka Sangmook Lee Harish Pant Thomas B. Shea |
author_sort |
Edward F. Boumil |
title |
Assembly and turnover of neurofilaments in growing axonal neurites |
title_short |
Assembly and turnover of neurofilaments in growing axonal neurites |
title_full |
Assembly and turnover of neurofilaments in growing axonal neurites |
title_fullStr |
Assembly and turnover of neurofilaments in growing axonal neurites |
title_full_unstemmed |
Assembly and turnover of neurofilaments in growing axonal neurites |
title_sort |
assembly and turnover of neurofilaments in growing axonal neurites |
publisher |
The Company of Biologists |
series |
Biology Open |
issn |
2046-6390 |
publishDate |
2018-01-01 |
description |
Neurofilaments (NFs) are thought to provide stability to the axon. We examined NF dynamics within axonal neurites of NB2a/d1 neuroblastoma by transient transfection with green fluorescent protein-tagged NF-heavy (GFP-H) under the control of a tetracycline-inducible promoter. Immunofluorescent and biochemical analyses demonstrated that GFP-H expressed early during neurite outgrowth associated with a population of centrally-situated, highly-phosphorylated crosslinked NFs along the length of axonal neurites (‘bundled NFs’). By contrast, GFP-H expressed after considerable neurite outgrowth displayed markedly reduced association with bundled NFs and was instead more evenly distributed throughout the axon. This differential localization was maintained for up to 2 weeks in culture. Once considerable neurite outgrowth had progressed, GFP that had previously associated with the NF bundle during early expression was irreversibly depleted by photobleaching. Cessation of expression allowed monitoring of NF turnover. GFP-H associated bundled NFs underwent slower decay than GFP-H associated with surrounding, less-phosphorylated NFs. Notably, GFP associated with bundled NFs underwent similar decay rates within the core and edges of this bundle. These results are consistent with previous demonstration of a resident NF population within axonal neurites, but suggest that this population is more dynamic than previously considered. |
topic |
Neurofilament Cytoskeleton Axon Axogenesis Axonal transport |
url |
http://bio.biologists.org/content/7/1/bio028795 |
work_keys_str_mv |
AT edwardfboumil assemblyandturnoverofneurofilamentsingrowingaxonalneurites AT rishelvohnoutka assemblyandturnoverofneurofilamentsingrowingaxonalneurites AT sangmooklee assemblyandturnoverofneurofilamentsingrowingaxonalneurites AT harishpant assemblyandturnoverofneurofilamentsingrowingaxonalneurites AT thomasbshea assemblyandturnoverofneurofilamentsingrowingaxonalneurites |
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1721404992453083136 |