J. Sangerman et al., alpha-Spectrins are major ubiquitinated proteins in rat hippocampal neurons and components of ubiquitinated inclusions in neurodegenerative disorders, BRAIN RES B, 54(4), 2001, pp. 405-411
We have demonstrated that rr-spectrins (alpha Spl Sigma* and alpha Spll Sig
ma1) are major ubiquitinated proteins in terminally differentiated hippocam
pal neurons in culture. Western blotting experiments, using alpha Spl Sigma
1, alpha Spll Sigma1, and ubiquitin antibodies and lysates of 11-day-old cu
ltured rat hippocampal neurons, have demonstrated that a single band comigr
ating with rYSplI* and alpha Spll Sigma1 in a 5% polyacrylamide sodium dode
cyl sulfate gel is recognized by ubiquitin antibodies when I-125-protein A
is used for detection, Immunofluorescence staining of the 7- and 12 -day-ol
d rat hippocampal neuron cultures using ubiquitin, alpha Spl Sigma1, and al
pha Spll Sigma1 antibodies demonstrated that all of these antibodies label
neurons but not the astrocytes in the cultures. Immunoprecipitation of spec
trin subunits in lysates of 12-day-old rat hippocampal neurons under string
ent conditions (9.5 M urea) using alpha Spl Sigma1 and alpha Spll Sigma1 an
tibodies followed by Western blot experiments of the immunoprecipitated spe
ctrin subunits using alpha Spl Sigma1, alpha Spll Sigma1 and ubiquitin anti
bodies confirmed that both alpha Spl Sigma* and alpha Spll Sigma1 are ubiqu
itinated in rat hippocampal neurons. Furthermore, we demonstrated by immuno
histochemistry that cu-spectrins are components of the cytoplasmic ubiquiti
nated inclusions in hippocampal neurons in Alzheimers and Parkinson's disea
se patients. (C) 2001 Elsevier Science Inc.