Tyrosination state of alpha-tubulin in regenerating peripheral nerve

Certain modifications of the neuronal cytoskeleton that are associated with development also occur during regeneration of adult mammalian peripheral nerve. The aim of the present study was to examine one such modification, the tyrosination of alpha-tubulin. Adult rats were anaesthetized and the left or right sciatic nerve randomly selected and crushed to induce regeneration. In certain instances nerves were crushed then ligatured about the crush, to prevent regeneration. Five days later the rats were killed and the regenerating (or ligatured) and the contralateral (control) nerves were removed. Quantitative immunoblotting of nerve homogenates with antibodies that recognize tyrosinated alpha-tubulin and total alpha-tubulin revealed a significant increase (p < 0.01) in the proportion of alpha-tubulin that was tyrosinated in nerve pieces distal (peripheral) to a nerve crush and to uncrushed nerve. No such difference occurred in ligatured (crushed but nonregenerating) nerve, implying that the increase was related to the presence of regenerating fibres; nor was there any gradient in tyrosination of alpha-tubulin in control nerves. This effect was confirmed by cytofluorimetric scanning and fluorescence confocal laser scanning microscopy of fixed sections of control and regenerating nerve, stained with antibodies directed against tyrosinated alpha-tubulin. When nerves were separated into fractions containing assembled and nonassembled tubulin, a significant (p < 0.01) increase was found in the proportion of tyrosinated alpha-tubulin in the nonassembled tubulin fraction in nerve pieces containing regenerating fibres. This occurred in the absence of a change in the proportion of assembled and nonassembled tubulin.(ABSTRACT TRUNCATED AT 250 WORDS)