Overexpression of neuronal intermediate filament protein α-internexin in PC12 cells

Chung Liang Chien, Tzu Chiang Liu, Chung-Liang Ho, Kuo Shyan Lu

Research output: Contribution to journalArticle

24 Citations (Scopus)

Abstract

The neuronal intermediate filaments include not only the neurofilament triplet proteins but also peripherin and α-internexin. To determine whether neurite outgrowth is enhanced by ́-internexin, the cDNA of rat α-internexin tagged with enhanced green fluorescent protein (EGFP) was transfected into a rat adrenal pheochromocytoma cell line PC12 that responds to nerve growth factor (NGF) by induction of the neuronal phenotype. Selected stable clones were induced by NGF and examined for expression patterns of neuronal intermediate filaments by Western blot and immunocytochemistry. Differentiating neurons were also collected after NGF induction for RT-PCR analysis. Overexpressed α-internexin-EGFPs were found mainly in cell bodies and the proximal part of neurites. It was also found that overexpression of α-internexin-EGFPs enhanced the neurite outgrowth of PC12 cells at the early stages of NGF induction. Meantime, NF-L and NF-M were upregulated by the overexpression of α-internexin-EGFPs. Interestingly, α-internexin- EGFP-transfected cells obviously detached from culture plates at the later stages of NGF induction. Massive IF accumulations, swelling mitochondria, and degenerating neurites with numerous electron-dense granules were observed ultrastructurally in the α-internexin-EGFP-transfected cells. In addition, neuronal death was also characterized positively by the TUNEL assay. These observations may imply that cell death was occurring in α-internexin-EGFP- transfected cells. From this study, it could be suggested that α-internexin plays an important role in neurite outgrowth and regulates the expression of other neurofilaments during neuronal development. Apoptosis-like cell death could also be induced by the overexpression of α-internexin-EGFP in PC12 cells after NGF induction.

Original languageEnglish
Pages (from-to)693-706
Number of pages14
JournalJournal of Neuroscience Research
Volume80
Issue number5
DOIs
Publication statusPublished - 2005 Jun 1

Fingerprint

Intermediate Filament Proteins
PC12 Cells
Nerve Growth Factor
Intermediate Filaments
Neurites
Cell Death
Peripherins
Neurofilament Proteins
In Situ Nick-End Labeling
Human Body
Mitochondria
Complementary DNA
Clone Cells
Western Blotting
Immunohistochemistry
enhanced green fluorescent protein
Electrons
Apoptosis
Phenotype
Neurons

All Science Journal Classification (ASJC) codes

  • Cellular and Molecular Neuroscience

Cite this

Chien, Chung Liang ; Liu, Tzu Chiang ; Ho, Chung-Liang ; Lu, Kuo Shyan. / Overexpression of neuronal intermediate filament protein α-internexin in PC12 cells. In: Journal of Neuroscience Research. 2005 ; Vol. 80, No. 5. pp. 693-706.
@article{a03515fb3ad2494788d6c594ddf8cdad,
title = "Overexpression of neuronal intermediate filament protein α-internexin in PC12 cells",
abstract = "The neuronal intermediate filaments include not only the neurofilament triplet proteins but also peripherin and α-internexin. To determine whether neurite outgrowth is enhanced by ́-internexin, the cDNA of rat α-internexin tagged with enhanced green fluorescent protein (EGFP) was transfected into a rat adrenal pheochromocytoma cell line PC12 that responds to nerve growth factor (NGF) by induction of the neuronal phenotype. Selected stable clones were induced by NGF and examined for expression patterns of neuronal intermediate filaments by Western blot and immunocytochemistry. Differentiating neurons were also collected after NGF induction for RT-PCR analysis. Overexpressed α-internexin-EGFPs were found mainly in cell bodies and the proximal part of neurites. It was also found that overexpression of α-internexin-EGFPs enhanced the neurite outgrowth of PC12 cells at the early stages of NGF induction. Meantime, NF-L and NF-M were upregulated by the overexpression of α-internexin-EGFPs. Interestingly, α-internexin- EGFP-transfected cells obviously detached from culture plates at the later stages of NGF induction. Massive IF accumulations, swelling mitochondria, and degenerating neurites with numerous electron-dense granules were observed ultrastructurally in the α-internexin-EGFP-transfected cells. In addition, neuronal death was also characterized positively by the TUNEL assay. These observations may imply that cell death was occurring in α-internexin-EGFP- transfected cells. From this study, it could be suggested that α-internexin plays an important role in neurite outgrowth and regulates the expression of other neurofilaments during neuronal development. Apoptosis-like cell death could also be induced by the overexpression of α-internexin-EGFP in PC12 cells after NGF induction.",
author = "Chien, {Chung Liang} and Liu, {Tzu Chiang} and Chung-Liang Ho and Lu, {Kuo Shyan}",
year = "2005",
month = "6",
day = "1",
doi = "10.1002/jnr.20506",
language = "English",
volume = "80",
pages = "693--706",
journal = "Journal of Neuroscience Research",
issn = "0360-4012",
publisher = "Wiley-Liss Inc.",
number = "5",

}

Overexpression of neuronal intermediate filament protein α-internexin in PC12 cells. / Chien, Chung Liang; Liu, Tzu Chiang; Ho, Chung-Liang; Lu, Kuo Shyan.

In: Journal of Neuroscience Research, Vol. 80, No. 5, 01.06.2005, p. 693-706.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Overexpression of neuronal intermediate filament protein α-internexin in PC12 cells

AU - Chien, Chung Liang

AU - Liu, Tzu Chiang

AU - Ho, Chung-Liang

AU - Lu, Kuo Shyan

PY - 2005/6/1

Y1 - 2005/6/1

N2 - The neuronal intermediate filaments include not only the neurofilament triplet proteins but also peripherin and α-internexin. To determine whether neurite outgrowth is enhanced by ́-internexin, the cDNA of rat α-internexin tagged with enhanced green fluorescent protein (EGFP) was transfected into a rat adrenal pheochromocytoma cell line PC12 that responds to nerve growth factor (NGF) by induction of the neuronal phenotype. Selected stable clones were induced by NGF and examined for expression patterns of neuronal intermediate filaments by Western blot and immunocytochemistry. Differentiating neurons were also collected after NGF induction for RT-PCR analysis. Overexpressed α-internexin-EGFPs were found mainly in cell bodies and the proximal part of neurites. It was also found that overexpression of α-internexin-EGFPs enhanced the neurite outgrowth of PC12 cells at the early stages of NGF induction. Meantime, NF-L and NF-M were upregulated by the overexpression of α-internexin-EGFPs. Interestingly, α-internexin- EGFP-transfected cells obviously detached from culture plates at the later stages of NGF induction. Massive IF accumulations, swelling mitochondria, and degenerating neurites with numerous electron-dense granules were observed ultrastructurally in the α-internexin-EGFP-transfected cells. In addition, neuronal death was also characterized positively by the TUNEL assay. These observations may imply that cell death was occurring in α-internexin-EGFP- transfected cells. From this study, it could be suggested that α-internexin plays an important role in neurite outgrowth and regulates the expression of other neurofilaments during neuronal development. Apoptosis-like cell death could also be induced by the overexpression of α-internexin-EGFP in PC12 cells after NGF induction.

AB - The neuronal intermediate filaments include not only the neurofilament triplet proteins but also peripherin and α-internexin. To determine whether neurite outgrowth is enhanced by ́-internexin, the cDNA of rat α-internexin tagged with enhanced green fluorescent protein (EGFP) was transfected into a rat adrenal pheochromocytoma cell line PC12 that responds to nerve growth factor (NGF) by induction of the neuronal phenotype. Selected stable clones were induced by NGF and examined for expression patterns of neuronal intermediate filaments by Western blot and immunocytochemistry. Differentiating neurons were also collected after NGF induction for RT-PCR analysis. Overexpressed α-internexin-EGFPs were found mainly in cell bodies and the proximal part of neurites. It was also found that overexpression of α-internexin-EGFPs enhanced the neurite outgrowth of PC12 cells at the early stages of NGF induction. Meantime, NF-L and NF-M were upregulated by the overexpression of α-internexin-EGFPs. Interestingly, α-internexin- EGFP-transfected cells obviously detached from culture plates at the later stages of NGF induction. Massive IF accumulations, swelling mitochondria, and degenerating neurites with numerous electron-dense granules were observed ultrastructurally in the α-internexin-EGFP-transfected cells. In addition, neuronal death was also characterized positively by the TUNEL assay. These observations may imply that cell death was occurring in α-internexin-EGFP- transfected cells. From this study, it could be suggested that α-internexin plays an important role in neurite outgrowth and regulates the expression of other neurofilaments during neuronal development. Apoptosis-like cell death could also be induced by the overexpression of α-internexin-EGFP in PC12 cells after NGF induction.

UR - http://www.scopus.com/inward/record.url?scp=21244491205&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=21244491205&partnerID=8YFLogxK

U2 - 10.1002/jnr.20506

DO - 10.1002/jnr.20506

M3 - Article

VL - 80

SP - 693

EP - 706

JO - Journal of Neuroscience Research

JF - Journal of Neuroscience Research

SN - 0360-4012

IS - 5

ER -