Axons and their synapses distal to an injury undergo rapid Wallerian degeneration, but axons in the C57BL/WldS mouse are protected. The degenerative and protective mechanisms are unknown. We identified the protective gene, which encodes an N-terminal fragment of ubiquitination factor E4B (Ube4b) fused to nicotinamide mononucleotide adenylyltransferase (Nmnat), and showed that it confers a dose-dependent block of Wallerian degeneration. Transected distal axons survived for two weeks, and neuromuscular junctions were also protected. Surprisingly, the Wld protein was located predominantly in the nucleus, indicating an indirect protective mechanism. Nmnat enzyme activity, but not NAD+ content, was increased fourfold in WldS tissues. Thus, axon protection is likely to be mediated by altered ubiquitination or pyridine nucleotide metabolism.

Exons of three genes were identified within the 85-kilobase tandem triplication unit of the slow Wallerian degeneration mutant mouse, C57BL͞Wld S . Ubiquitin fusion degradation protein 2 (Ufd2) and a previously undescribed gene, D4Cole1e, span the proximal and distal boundaries of the repeat unit, respectively. They have the same chromosomal orientation and form a chimeric gene when brought together at the boundaries between adjacent repeat units in Wld S . The chimeric mRNA is abundantly expressed in the nervous system and encodes an in-frame fusion protein consisting of the N-terminal 70 amino acids of Ufd2, the C-terminal 302 amino acids of D4Cole1e, and an aspartic acid formed at the junction. Antisera raised against synthetic peptides detect the expected 43-kDa protein specifically in Wld S brain. This expression pattern, together with the previously established role of ubiquitination in axon degeneration, makes the chimeric gene a promising candidate for Wld. The third gene altered by the triplication, Rbp7, is a novel member of the cellular retinoid-binding protein family and is highly expressed in white adipose tissue and mammary gland. The whole gene lies within the repeat unit leading to overexpression of the normal transcript in Wld S mice. However, it is undetectable on Northern blots of Wld S brain and seems unlikely to be the Wld gene. These data reveal both a candidate gene for Wld and the potential of the Wld S mutant for studies of ubiquitin and retinoid metabolism.

Axons in WldS mutant mice are protected from Wallerian degeneration by overexpression of a chimeric Ube4b/Nmnat (Wld) gene. Expression of Wld protein was independent of age in these mice. However we identified two distinct neuromuscular synaptic responses to axotomy. In young adult Wlds mice, axotomy induced progressive, asynchronous synapse withdrawal from motor endplates, strongly resembling neonatal synapse elimination. Thus, five days after axotomy, 50–90 % of endplates were still partially or fully occupied and expressed endplate potentials (EPPs). By 10 days, fewer than 20 % of endplates still showed evidence of synaptic activity. Recordings from partially occupied junctions indicated a progressive decrease in quantal content in inverse proportion to endplate occupancy. In Wlds mice aged > 7 months, axons were still protected from axotomy but synapses degenerated rapidly, in wild‐type fashion: within three days less than 5 % of endplates contained vestiges of nerve terminals. The axotomy‐induced synaptic withdrawal phenotype decayed with a time constant of ∼30 days. Regenerated synapses in mature Wlds mice recapitulated the juvenile phenotype. Within 4–6 days of axotomy 30–50 % of regenerated nerve terminals still occupied motor endplates. Age‐dependent synapse withdrawal was also seen in transgenic mice expressing the Wld gene. Co‐expression of Wld protein and cyan fluorescent protein (CFP) in axons and neuromuscular synapses did not interfere with the protection from axotomy conferred by the Wld gene. Thus, Wld expression unmasks age‐dependent, compartmentally organised programmes of synapse withdrawal and degeneration.

In vitro evidence has suggested a change in the ability of tau bearing mutations associated with fronto-temporal dementia to promote microtubule assembly. We have used a cellular assay to quantitate the effect of both isoform differences and mutations on the physiological function of tau. Whilst all variants of tau bind to microtubules, microtubule extension is reduced in cells transfected with 3-relative to 4-repeat tau. Mutations reduce microtubule extension with the P301L mutation having a greater effect than the V337M mutation. The R406W mutation had a small effect on microtubule extension but, surprisingly, tau with this mutation was less phosphorylated in intact cells than the other variants.z 1999 Federation of European Biochemical Societies.

No abstract

No abstract

Defects in actin dynamics affect activity-dependent modulation of synaptic transmission and neuronal plasticity, and can cause cognitive impairment. A salient candidate actin-binding protein linking synaptic dysfunction to cognitive deficits is Drebrin (DBN). However, the specific mode of how DBN is regulated at the central synapse is largely unknown. In this study we identify and characterize the interaction of the PTEN tumor suppressor with DBN. Our results demonstrate that PTEN binds DBN and that this interaction results in the dephosphorylation of a site present in the DBN C-terminus - serine 647. PTEN and pS647-DBN segregate into distinct and complimentary compartments in neurons, supporting the idea that PTEN negatively regulates DBN phosphorylation at this site. We further demonstrate that neuronal activity increases phosphorylation of DBN at S647 in hippocampal neurons in vitro and in ex vivo hippocampus slices exhibiting seizure activity, potentially by inducing rapid dissociation of the PTEN:DBN complex. Our results identify a novel mechanism by which PTEN is required to maintain DBN phosphorylation at dynamic range and signifies an unusual regulation of an actin-binding protein linked to cognitive decline and degenerative conditions at the CNS synapse.

The microtubule-associated protein, tau, is the principal component of paired helical filaments (PHFs) in Alzheimer's disease. PHF-tau is highly phosphorylated and a total of 25 sites of phosphorylation have so far been identified. Many of these sites are serine or threonine residues that are immediately followed in the sequence by proline residues, and hence are candidate phosphorylation sites for proline-directed kinases. In vitro, glycogen synthase kinase-3 (GSK-3), extracellular signal-related kinase-1 and -2, and mitogen-activated protein kinases, p38 kinase and c-jun N-terminal kinase, all phosphorylate many of these sites, although with different efficiencies for particular sites. Phosphorylation studies in transfected cells and neurons show that GSK-3 phosphorylates tau more extensively than do these other proline-directed kinases. Mutations in tau have been shown to affect in vitro phosphorylation of tau by GSK-3. The Arg406-->Trp (R406W) tau mutation also affects tau phosphorylation in cells.