title: Selective accumulation of the high molecular weight neurofilament subunit within the distal region of growing axonal neurites.
abstract: Axonal maturation in situ is accompanied by the transition of neurofilaments (NFs) comprised of only NF-M and NF-L to those also containing NF-H. Since NF-H participates in interactions of NFs with each other and with other cytoskeletal constituents, its appearance represents a critical event in the stabilization of axons that accompanies their maturation. Whether this transition is effected by replacement of "doublet" NFs with "triplet" NFs, or by incorporation of NF-H into existing doublet NFs is unclear. To address this issue, we examined the distribution of NF subunit immunoreactivity within axonal cytoskeletons of differentiated NB2a/d1 cell and DRG neurons between days 3-7 of outgrowth. Endogenous immunoreactivity either declined in a proximal-distal gradient or was relatively uniform along axons. This distribution was paralleled by microinjected biotinylated NF-L. By contrast, biotinylated NF-H displayed a bipolar distribution, with immunoreactivity concentrated within the proximal- and distal-most axonal regions. Proximal biotinylated NF-H accumulation paralleled that of endogenous NF immunoreactivity; however, distal-most biotinylated NF-H accumulation dramatically exceeded that of endogenous NFs and microinjected NF-L. This phenomenon was not due to co-polymerization of biotin-H with vimentin or alpha-internexin. This phenomenon declined with continued time in culture. These data suggest that NF-H can incorporate into existing cytoskeletal structures, and therefore suggest that this mechanism accounts for at least a portion of the accumulation of triplet NFs during axonal maturation. Selective NF-H accumulation into existing cytoskeletal structures within the distal-most region may provide de novo cytoskeletal stability for continued axon extension and/or stabilization.
authors.affiliation: Center for Cellular Neurobiology and Neurodegeneration Research, Department of Biological Sciences, University of Massachusetts-Lowell, MA 01854, USA.
authors.forename: J T
authors.initials: JT
authors.lastname: Yabe
authors.forename: F S
authors.initials: FS
authors.lastname: Wang
authors.forename: T
authors.initials: T
authors.lastname: Chylinski
authors.forename: T
authors.initials: T
authors.lastname: Katchmar
authors.forename: T B
authors.initials: TB
authors.lastname: Shea
mesh_terms: D000818:Animals; D001369:Axons / Q000378:metabolism*; D019921:Biotinylation; D002352:Carrier Proteins / Q000378:metabolism; D002417:Cattle; D002454:Cell Differentiation; D002455:Cell Division; D003720:Densitometry; D050883:HSC70 Heat-Shock Proteins; D006801:Humans; D007381:Intermediate Filament Proteins; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D016900:Neurofilament Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D009474:Neurons / Q000378:metabolism*; D011506:Proteins / Q000378:metabolism; D013116:Spinal Cord / Q000378:metabolism; D013997:Time Factors; D014407:Tumor Cells, Cultured; D014746:Vimentin / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D050883:HSC70 Heat-Shock Proteins; C494371:HSPA8 protein, human; D007381:Intermediate Filament Proteins; D016900:Neurofilament Proteins; D011506:Proteins; D014746:Vimentin; C046643:alpha-internexin; C051774:neurofilament protein H