Two 68‐kDa Proteins in Slow Axonal Transport Belong to the 70‐kDa Heat Shock Protein Family and the Annexin Family

Abstract

The major 68-kDa protein found selectively in the faster of the two subcomponents of slow axonal transport [group IV or slow component b (SCb)] in the rat sciatic nerve has been characterized. It was found to contain two distinct classes of proteins, S1 and S2, both of which have isoelectric points of 5.7, but differ in their solubility in the presence of calcium. The S1 protein, which contributes up to 70% of the 68-kDa component, was soluble in the presence or absence of calcium, whereas the S2 protein was bound to the cytoskeleton in a calcium-dependent manner. Further characterization of the two proteins by peptide mapping and immunological methods revealed that the S1 protein belonged to a family of proteins related to the 70-kDa heat shock protein, whereas the S2 protein was identical to 68-kDa calelectrin (annexin VI). Selective occurrence in SCb of these proteins with potential abilities to regulate protein-protein or protein-membrane interactions suggests that they may play important roles in the control of cytoskeletal organization in the axon, because SCb contains mainly cytoskeletal proteins in a more dynamic form compared with the slowest rate component, slow component a, which is enriched in the stably polymerized form of these proteins.

Interactions between brain mitochondria and cytoskeleton: Evidence for specialized outer membrane domains involved in the association of cytoskeleton‐associated proteins to mitochondria in situ and in vitro

“…In support of the possibility that proteins of the HSP 70 family might be similar or identical to a-IFA-labelled proteins associated to MT or mitochondrial OM described in the present work, is the homology of the MT-associated HSP 70 with a-internexin (Green and Liem, 1989;Napolitano et al, 19851, a NF-associated protein which belongs to the IF family (Fliegner et al, 1990). Evidence for ubiquitous distribution of the HSP 70 is further illustrated by its involvement in ATP-dependent disassembly of clathrin-coated vesicles (Chappel et al, 1986;Ungewickell, 19851, its translocation into the nucleole after heat shock (Welch and Feramisco, 1984), and its axonal transport in the slow component B (actin and MT polymers, Sekimoto et al, 1991). The latter report (Sekimoto et al, 1991) further suggests the involvement of both HSP 70 and annexin VI (a C a + + -dependent phospholipid binding protein cross-linking membranous organelles and cytoskeleton) in the dynamic interactions between membranous organelles and cytoskeleton polymers in axons, thus opening new perspectives in the regulation of mitochondrial motion.…”

“…Evidence for ubiquitous distribution of the HSP 70 is further illustrated by its involvement in ATP-dependent disassembly of clathrin-coated vesicles (Chappel et al, 1986;Ungewickell, 19851, its translocation into the nucleole after heat shock (Welch and Feramisco, 1984), and its axonal transport in the slow component B (actin and MT polymers, Sekimoto et al, 1991). The latter report (Sekimoto et al, 1991) further suggests the involvement of both HSP 70 and annexin VI (a C a + + -dependent phospholipid binding protein cross-linking membranous organelles and cytoskeleton) in the dynamic interactions between membranous organelles and cytoskeleton polymers in axons, thus opening new perspectives in the regulation of mitochondrial motion.…”

Slow axonal transport of the cytosolic chaperonin CCT with Hsc73 and actin in motor neurons

“…Observed differences are due in part to macromolecular crowding, it follows that in vivo studies may offer particularly valuable information about the function of molecular chaperones in axonal transport. It is well established that Hsc73, as clathrin-uncoating ATPase, is present in mature (connected) axons and transported in SCb (de Waegh and Brady, 1989;Black et al, 1991;Sekimoto et al, 1991). In particular, the transport of a member of the constitutive 70 kDa heat shock protein family in SCb of rat sciatic nerve has only been reported by Sekimoto et al (1991).…”

“…Because these proteins can take weeks or even months to reach their destinations, they may require chaperones to maintain their native form. One possible candidate is Hsc73, which has been shown to be transported in SCb (de Waegh and Brady, 1989;Black et al, 1991;Sekimoto et al, 1991). Another is CCT, the ␣-subunit of which has been shown in neuritic processes of neuronal phenotype ND7/23 cells to colocalize with unassembled actin at the leading edge of growth cones (Roobol et al, 1995).…”

Heat-shock proteins in axoplasm: High constitutive levels and transfer of inducible isoforms from glia

“…For example, the expression of constitutive HSP 70 isoforms or mRNA is much higher in the cell bodies of cerebellar, hippocampal, and spinal cord motor neurons than in glia (Manzerra and Brown, 1992;Brown, 1994). Although it is well documented that axons receive HSPs from the cell body at a slow transport rate of 0.5-3.0 mm/day (Clark and Brown, 1985;Tytell and Barbe, 1987;deWaegh and Brady, 1989;Black et al, 1991;Sekimoto et al, 1991), the levels of constitutive HSP 70s in axoplasm have not been measured previously. High levels of constitutive HSP 70s in axoplasm could help axons to buffer the deleterious effects of acute heat shock (Manzerra and Brown, 1992).…”