RETROGRADE AXONAL-TRANSPORT OF LOCALLY SYNTHESIZED PROTEINS, EG, ACTIN AND HEAT-SHOCK PROTEIN-70, IN REGENERATING ADULT FROG SCIATIC SENSORY AXONS

The local synthesis and subsequent retrograde axonal transport of [S-3 5]methionine-labelled proteins was studied in the in vitro regeneratin g adult frog sciatic sensory axons. By the use of a three compartment culture system, proteins in the outgrowth region were selectively labe lled. After 2 days in culture a rise in TCA-insoluble radioactivity wa s detected in the dorsal root ganglia, which could be prevented by the addition of vinblastine or 2,4-dinitrophenol to the nerve proximal to the crush site. Two-dimensional polyacrylamide gel electrophoresis of ganglionic proteins revealed a pattern of 35 labelled polypeptides wi th apparent molecular masses (M(m)) ranging from < 15 to 95 kDa and wi th isoelectric points (pI) ranging from 4.5 to 6.5. The major ones, re presenting about 75% of the activity in a gel, were of M(m)/pI 47/5.4, 48/6.1, .57/6.0, 62/5.2, 65/4.9-5.0, 65/5.2, and 81/5.4 respectively. One of these polypeptides (47/5.4) was identified as actin and anothe r (81/5.4) as a member of the heat shock protein 70 family. The spots at 65/4.9-5.0 were tubulin isoforms. There was a striking similarity b etween transported proteins on one hand, and proteins synthesized in t he injured nerve on the other, with respect to the M(m)/pI of at least 14 protein species. The results suggest that a selected set of protei ns, synthesized by non-neuronal cells, e.g., Schwann cells, is transfe rred to the ganglionic cell bodies by retrograde axonal transport. (C) 1994 Wiley-Liss, Inc.