Glyceraldehyde-3-phosphate dehydrogenase activity and F-actin associations in synaptosomes and postsynaptic densities of porcine cerebral cortex.
1. Glyceraldehyde-3-phosphate dehydrogenase (G3PD) is a glycolytic enzyme that has also been implicated in a wide variety of functions within neurons. Because of the well-documented role of G3PD as an actin-binding protein, we sought evidence for a G3PD-actin complex in synaptosomes and postsynaptic densities (PSDs). 2. We have shown G3PD association with 0.5-microgram synaptosomal particles by immunofluorescence as similarly demonstrated for actin (Toh et al., Nature 264:648-650, 1976). An immunoblot analysis also showed G3PD and actin to be enriched in synaptosomes. Further analysis of subcellular fractions from synaptosomes showed the PSD but not the synaptosomal plasma membranes to be enriched in G3PD and actin. 3. Highest levels of G3PD catalytic activity were found in synaptosomes and PSDs. Although synaptosomes showed significant activity for phosphoglycerate kinase (PGK), an enzyme in sequence with G3PD for ATP production in the glycolytic pathway, no such activity was detected in the PSD fraction. 4. Our studies indicate that a G3PD-actin complex may exist at the synapse. A physical association of G3PD with endogenous F-actin in synaptosomes and PSDs was demonstrated by combined phalloidin shift velocity sedimentation/immunoblot studies. By this approach, synaptosomal G3PD-actin complexes were also found to be significantly less dense than the PSD G3PD-actin complexes. 5. G3PD and PGK catalytic activity in synaptosomes suggests a role in glycolysis, as well as actin binding, in the presynaptic terminals. On the other hand, the high levels of G3PD activity in PSDs but lack of PGK activity suggests that G3PD is involved in nonglycolytic functions, such as actin binding and actin filament network organization.
Rogalski-Wilk
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Department of Anatomy and Cell Biology, University of Illinois at Chicago 60612, USA.
Cohen
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D000199:Actins / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D002540:Cerebral Cortex / Q000378:metabolism*; D005455:Fluorescent Antibody Technique; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism* / Q000502:physiology; D015151:Immunoblotting; D010735:Phosphoglycerate Kinase / Q000378:metabolism; D011485:Protein Binding; D013347:Subcellular Fractions; D013552:Swine; D013569:Synapses / Q000378:metabolism*; D013570:Synaptic Membranes / Q000378:metabolism; D013572:Synaptic Vesicles / Q000378:metabolism; D013574:Synaptosomes / Q000378:metabolism*
D000199:Actins; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010735:Phosphoglycerate Kinase