Diversity in the axonal transport of structural proteins: major differences between optic and spinal axons in the rat

Abstract

Investigations of slow axonal transport reveal variation in both protein composition and the rate of movement. However, these studies involve a variety of nerve preparations in different species, and most lack the resolution needed to determine the kinetics of identified proteins. We have compared the axonal transport of slow-transported proteins in retinal ganglion cells and spinal motor neurons of young rats. Nine proteins that contribute to axonal structures were examined: the neurofilament triplet (NFT), alpha and beta tubulin, actin, fodrin, calmodulin, and clathrin. Axonally transported proteins were pulse-labeled by intraocular or intracord injections of 35S-methionine. After allowing sufficient time for labeled slow-component proteins to enter the spinal or optic nerves, consecutive 2-3 mm nerve segments were subjected to SDS-PAGE. Fluorographs were used as templates for locating the gel regions containing the above polypeptides, and the radioactivity in these regions was measured by liquid-scintillation spectrometry. In retinal ganglion cells, the peak of tubulin labeling advanced at 0.36 mm/d in association with the NFT and fodrin. The cotransport of tubulin and the NFT identified this complex as the slower subcomponent of slow transport, termed slow component a (SCa) and representing the movement of the microtubule-neurofilament network. The peaks of actin and calmodulin labeling were cotransported at 2.3 mm/d in near-register with peaks of fodrin and clathrin labeling. These 4 proteins, moving ahead of the NFT, identified this complex as SCb--the faster subcomponent of slow transport, which represents the movement of the cytoplasmic matrix and microtrabecular lattice. Both subcomponents had the same composition and rate as that reported for the optic axons of guinea pigs and rabbits, establishing a basic mammalian pattern. In spinal motor axons, the SCa tubulin peak advanced at 1.3 mm/d, and the SCb actin and calmodulin peaks were cotransported at 3.1 mm/d. Unlike optic axons, SCa in motor axons was more heavily labeled than SCb, and included labeled peaks of actin, clathrin, and calmodulin moving in register with the SCa tubulin peak. Actin was the most heavily labeled of these SCb proteins moving with SCa, and it left a higher plateau of radioactivity behind the advancing SCa peak. The SDS-PAGE labeling pattern for SCb did not differ from that seen in optic axons, except that some tubulin was found to form a peak that advanced in register with the actin and calmodulin peaks.(ABSTRACT TRUNCATED AT 400 WORDS)

Cytoplasmic dynein is associated with slow axonal transport.

“…Quantitation of the dynein polypeptides associated with the various rate components (Fig. IB) (7,23) (Fig. 2).…”

Application of Rho Antagonist to Neuronal Cell Bodies Promotes Neurite Growth in Compartmented Cultures and Regeneration of Retinal Ganglion Cell Axons in the Optic Nerve of Adult Rats

“…Axonal transport in normal and injured optic nerve of adult rat has been well described (McQuarrie et al, 1986(McQuarrie et al, , 1989McKerracher et al, 1990) and is detected by examining the movement of 35 Smethionine-labeled proteins along 2 mm optic nerve segments. In uninjured optic nerve, the rate of slow axonal transport of neurofilament, as detected by the 150 kDa middle neurofilament subunit (NF-M), is ϳ0.5 mm/d (McQuarrie et al, 1986), and the transport rate decreases significantly when the optic nerve is crushed intracranially near the optic chiasma (McKerracher et al, 1990).…”

“…In uninjured optic nerve, the rate of slow axonal transport of neurofilament, as detected by the 150 kDa middle neurofilament subunit (NF-M), is ϳ0.5 mm/d (McQuarrie et al, 1986), and the transport rate decreases significantly when the optic nerve is crushed intracranially near the optic chiasma (McKerracher et al, 1990). To determine whether C3-07 had an impact on slow axonal transport, the optic nerve was crushed 6 d after labeling, a time when tubulin and neurofilament proteins enter the optic nerve (Fig.…”

Fast axonal transport of kinesin in the rat visual system: functionality of kinesin heavy chain isoforms.

“…These values were added together and defined as the total amount of radiolabeled kinesin present in the homogenate. McQuarrie et al, 1986;Oblinger et al, 1987) The absolute mass of kinesin in the optic nerve of several animals was determined to insure that differences in the amount of radiolabeled kinesin at different time points post-labeling were due to differences in specific activity, not to differences in the amount of kinesin present in the axons of different animals. Optic nerves from two non-radiolabeled rats were harvested and homogenized in lysis buffer.…”

“…Rates of movement for different subclasses of membrane bounded organelles and cytoskeletal structures through the axon are known (Willard et al, 1974;McQuarrie et al, 1986;Oblinger et al, 1987). If rate(s) of movement for kinesin in the axon can be determined, then roles for this mechanochemical enzyme in the movement of specific types of axonal structures can be assessed.…”