Protective effect of supplemental superoxide dismutase on survival of neuronal cells during starvation. Requirement for cytosolic distribution.
There is evidence that raising cellular levels of Cu2+/Zn2+ superoxide dismutase (SOD1) can protect neurons from oxidative injury. We compared a novel method of elevating neuronal SOD activity using a recombinant hybrid protein composed of the atoxic neuronal binding domain of tetanus toxin (C fragment or TTC) and human SOD1 (hSOD1) with increasing cellular SOD levels through overexpression. Fetal murine cortical neurons or N18-RE-105 cells were incubated with the TTC-hSOD1 hybrid protein and compared to cells constitutively expressing hSOD1 for level of SOD activity, cellular localization of hSOD1, and capacity to survive glucose and pyruvate starvation. Cells incubated with TTC-hSOD1 showed a threefold increase in cellular SOD activity over control cells. This level of increase was comparable to fetal cortical neurons from transgenic mice constitutively expressing hSOD1 and transfected N18-RE-105 cells expressing a green fluorescent protein-hSOD1 fusion protein (GFP-hSOD1). Human SOD1 was distributed diffusely throughout the cytoplasm of the transgenic murine neurons and transfected N18-RE-105 cells. In contrast, cells incubated with TTC-hSOD1 showed hSOD1 localized to the cell surface and intra-cytoplasmic vesicles. The cells expressing hSOD1 showed enhanced survival in glucose- and pyruvate-free medium. Neither cortical neurons nor N18-RE-105 cells incubated in TTC-hSOD1 showed increased survival during starvation. Access to the site where toxic superoxides are generated or their targets may be necessary for the protective function of SOD1.
Matthews
C C
CC
Department of Neurology, University of Maryland, School of Medicine, and Veteran's Affairs Medical Center, Baltimore, MD, 21201, USA. ccmatthe@umaryland.edu
Figueiredo
D M
DM
Wollack
J B
JB
Fairweather
N F
NF
Dougan
G
G
Hallewell
R A
RA
Cadet
J L
JL
Fishman
P S
PS
D000818:Animals; D016923:Cell Death / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D002540:Cerebral Cortex / Q000166:cytology; D004734:Energy Metabolism / Q000502:physiology; D015971:Gene Expression Regulation, Enzymologic; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D007202:Indicators and Reagents / Q000378:metabolism; D008164:Luminescent Proteins / Q000235:genetics; D051379:Mice; D008815:Mice, Inbred Strains; D008822:Mice, Transgenic; D009447:Neuroblastoma; D009474:Neurons / Q000166:cytology* / Q000378:metabolism*; D018384:Oxidative Stress / Q000502:physiology*; D010446:Peptide Fragments / Q000235:genetics; D013217:Starvation / Q000378:metabolism; D013482:Superoxide Dismutase / Q000032:analysis / Q000235:genetics* / Q000378:metabolism*; D013744:Tetanus Toxin / Q000235:genetics; D014162:Transfection; D014407:Tumor Cells, Cultured
D007202:Indicators and Reagents; D008164:Luminescent Proteins; D010446:Peptide Fragments; D013744:Tetanus Toxin; C040635:tetanus toxin fragment C; D049452:Green Fluorescent Proteins; D013482:Superoxide Dismutase