dementia ͉ motor neuron disease ͉ neurodegeneration ͉ protein aggregation F TLD is a relatively common cause of dementia among patients with onset before 65, typically manifesting with behavioral changes or language impairment due to degeneration of subpopulations of cortical neurons in the frontal, temporal and insular regions (1). By contrast, ALS presents with muscle weakness and spasticity due to degeneration of motor neurons in both layer 5 of cortex and in the spinal cord, resulting in death from respiratory failure in 3-5 years (2, 3). Interestingly, approximately 20% of patients with ALS also develop FTLD, and approximately 15% of FLTD patients also develop ALS (4, 5).The discovery that TDP-43 is present in cytoplasmic aggregates in both ALS and FTLD-U provided evidence that the two disorders may share a common underlying mechanism (6). TDP-43 is an RNA/DNA binding protein, implicated in regulation of alternative splicing of messenger RNA, RNA stability, and transcriptional control (7). The concept that TDP-43 can play a direct role in neurodegeneration was strengthened by recent reports that dominantly inherited missense mutations in TDP-43 are found in patients with familial ALS (8-12). Mutations in TDP-43 associated with ALS cluster in the C-terminal glycine-rich region, which is involved in protein-protein interactions between TDP-43 and other heterogeneous nuclear ribonuclear proteins (hnRNPs) (13). Furthermore, C-terminal fragments of TDP-43 are observed selectively in ALS and FTLD-U tissues, suggesting that proteolytic cleavage of TDP-43 leads to protein aggregation or another toxic property (6). Therefore, several putative mechanisms of TDP-43 induced neurodegeneration are currently under investigation, including toxic protein aggregation, and/or disruption of normal TDP-43 RNA/DNA binding protein function. Here we report a mouse model of TDP-43 induced neurodegeneration which recapitulates key features of ALS and FTLD-U, including ubiquitin aggregate pathology with selective vulnerability of cortical projection neurons and spinal motor neurons, but without the presence of TDP-43 aggregates. Together with recent reports of mutations in another RNA binding protein (FUS/TLS) in familial ALS (14, 15), this supports that altered RNA-binding protein function (rather than toxic aggregation of TDP-43) likely plays a central and unexpected role in ALS pathogenesis.

Mitofusins (Mfn1 and Mfn2) are outer mitochondrial membrane proteins involved in regulating mitochondrial dynamics. Mutations in Mfn2 cause Charcot-Marie-Tooth disease (CMT) type 2A, an inherited disease characterized by degeneration of long peripheral axons, but the nature of this tissue selectivity remains unknown. Here, we present evidence that Mfn2 is directly involved in and required for axonal mitochondrial transport, distinct from its role in mitochondrial fusion. Live imaging of neurons cultured from Mfn2 knock-out mice or neurons expressing Mfn2 disease mutants shows that axonal mitochondria spend more time paused and undergo slower anterograde and retrograde movements, indicating an alteration in attachment to microtubule-based transport systems. Furthermore, Mfn2 disruption altered mitochondrial movement selectively, leaving transport of other organelles intact. Importantly, both Mfn1 and Mfn2 interact with mammalian Miro (Miro1/Miro2) and Milton (OIP106/GRIF1) proteins, members of the molecular complex that links mitochondria to kinesin motors. Knockdown of Miro2 in cultured neurons produced transport deficits identical to loss of Mfn2, indicating that both proteins must be present at the outer membrane to mediate axonal mitochondrial transport. In contrast, disruption of mitochondrial fusion via knockdown of the inner mitochondrial membrane protein Opa1 had no effect on mitochondrial motility, indicating that loss of fusion does not inherently alter mitochondrial transport. These experiments identify a role for mitofusins in directly regulating mitochondrial transport and offer important insight into the cell type specificity and molecular mechanisms of axonal degeneration in CMT2A and dominant optic atrophy.

The identification of pathologic TDP-43 aggregates in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration, followed by the discovery of dominantly inherited point mutations in TDP-43 in familial ALS, have been critical insights into the mechanism of these untreatable neurodegenerative diseases. However, the biochemical basis of TDP-43 aggregation and the mechanism of how mutations in TDP-43 lead to disease remain enigmatic. In efforts to understand how TDP-43 alters its cellular localization in response to proteotoxic stress, we found that TDP-43 is sequestered into polyglutamine aggregates. Furthermore, we found that binding to polyglutamine aggregates requires a previously uncharacterized glutamine/asparagine (Q/N)-rich region in the C-terminal domain of TDP-43. Sequestration into polyglutamine aggregates causes TDP-43 to be cleared from the nucleus and become detergent-insoluble. Finally, we observed that sequestration into polyglutamine aggregates led to loss of TDP-43-mediated splicing in the nucleus and that polyglutamine toxicity could be partially rescued by increasing expression of TDP-43. These data indicate pathologic sequestration into polyglutamine aggregates, and loss of nuclear TDP-43 function may play an unexpected role in polyglutamine disease pathogenesis. Furthermore, as Q/N domains have a strong tendency to self-aggregate and in some cases can function as prions, the identification of a Q/N domain in TDP-43 has important implications for the mechanism of pathologic aggregation of TDP-43 in ALS and other neurodegenerative diseases.Abnormal protein aggregation is a hallmark of most inherited and acquired neurodegenerative diseases. Recently, TDP-43 was identified as a component of ubiquitinated aggregates in amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) 4 (1). The subsequent finding of mutations in TDP-43 in cases of inherited ALS indicates that TDP-43 can be directly involved in the pathogenesis of at least the familial forms of this disease (2-6). It is notable that although aggregates of a particular protein are initially associated with a specific clinical and pathologic syndrome, they are often observed in multiple other neurodegenerative disorders. For example, although cytoplasmic inclusions of TDP-43 were initially described in ALS and FTLD, they have also been observed in Alzheimer disease, diffuse Lewy body disease, dementia pugilistica, Huntington disease, and even inclusion body myopathies (7-11). Whether TDP-43 translocation to the cytosol and aggregation plays a direct role in the pathogenesis of these disorders, or instead is part of a more general cellular stress response, remains to be elucidated (12).Polyglutamine diseases are a family of neurodegenerative disorders caused by expansion of a CAG trinucleotide repeat in the coding regions of certain genes. Examples include Huntington disease, X-linked spinal-bulbar muscular atrophy, and several of the spinocerebellar ataxias (13-15). Expanded polyglutamine proteins have a ...

Glutamate carboxypeptidase II (GCPII, EC 3.14.17.21) is a membrane-bound enzyme found on the extracellular face of glia. The gene for this enzyme is designated FOLH1 in humans and Folh1 in mice. This enzyme has been proposed to be responsible for inactivation of the neurotransmitter N-acetylaspartylglutamate (NAAG) following synaptic release. Mice harboring a disruption of the gene for GCPII/Folh1 were generated by inserting into the genome a targeting cassette in which the intron-exon boundary sequences of exons 1 and 2 were removed and stop codons were inserted in exons 1 and 2. Messenger RNA for GCPII was not detected by northern blotting or RT-PCR analysis of RNA from the brains of -/-mutant mice nor was GCPII protein detected on western blots of this tissue. These GCPII null mutant mice developed normally to adulthood and exhibited a normal range of neurologic responses and behaviors including mating, open field activity and retention of position in rotorod tests. No significant differences were observed among responses of wild type, heterozygous mutant and homozygous mutant mice on tail flick and hot plate latency tests. Glutamate, NAAG and mRNA for metabotropic glutamate receptor type 3 levels were not significantly altered in response to the deletion of glutamate carboxypeptidase II. A novel membrane-bound NAAG peptidase activity was discovered in brain, spinal cord and kidney of the GCPII knock out mice. The kinetic values for brain NAAG peptidase activity in the wild type and GCPII null mutant were V max ¼ 45 and 3 pmol/mg/min and K m ¼ 2650 nM and 2494 nM, respectively. With the exception of magnesium and copper, this novel peptidase activity had a similar requirement for metal ions as GCPII. Two potent inhibitors of GCPII, 4,4¢-phosphinicobis-(butane-1,3 dicarboxilic acid) (FN6) and 2-(phosphonomethyl)pentanedioic acid (2-PMPA) 2 inhibited the residual activity. The IC 50 value for 2-PMPA was about 1 nM for wild-type brain membrane NAAG peptidase activity consistent with its activity against cloned rat and human GCPII, and 88 nM for the activity in brain membranes of the null mutants. Keywords: N-acetylaspartylglutamate (NAAG), N-acetylaspartylglutamate-peptidase (NAAG-peptidase), glutamate carboxypeptidase II metabotropic glutamate receptor. 3

The clinical and pathological overlap between amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD) suggests these diseases share common underlying mechanisms, a suggestion underscored by the discovery that TDP-43 inclusions are a key pathologic feature in both ALS and FTLD. This finding, combined with the identification of TDP-43 mutations in ALS, directly implicates this DNA/RNA binding protein in disease pathogenesis in ALS and FTLD. However, many key questions remain, including what is the normal function of TDP-43, and whether disease-associated mutations produce toxicity in the nucleus, cytoplasm or both. Furthermore, although pathologic TDP-43 inclusions are clearly associated with many forms of neurodegeneration, whether TDP-43 aggregation is a key step in the pathogenesis in ALS, FTLD and other disorders remains to be proven. This review will compare the features of numerous recently developed animal models of TDP-43-related neurodegeneration, and discuss how they contribute to our understanding of the pathogenesis of human ALS and FTLD.