Disruption of Cnp1 uncouples oligodendroglial functions in axonal support and myelination

Myelination of axons by oligodendrocytes enables rapid impulse propagation in the central nervous system. But long-term interactions between axons and their myelin sheaths are poorly understood. Here we show that Cnp1, which encodes 2',3'-cyclic nucleotide phosphodiesterase in oligodendrocytes, is essential for axonal survival but not for myelin assembly. In the absence of glial cyclic nucleotide phosphodiesterase, mice developed axonal swellings and neurodegeneration throughout the brain, leading to hydrocephalus and premature death. But, in contrast to previously studied myelin mutants, the ultrastructure, periodicity and physical stability of myelin were not altered in these mice. Genetically, the chief function of glia in supporting axonal integrity can thus be completely uncoupled from its function in maintaining compact myelin. Oligodendrocyte dysfunction, such as that in multiple sclerosis lesions, may suffice to cause secondary axonal loss.     

Disrupted function and axonal distribution of mutant tyrosyl-tRNA synthetase in dominant intermediate Charcot-Marie-Tooth neuropathy

Charcot-Marie-Tooth (CMT) neuropathies are common disorders of the peripheral nervous system caused by demyelination or axonal degeneration, or a combination of both features. We previously assigned the locus for autosomal dominant intermediate CMT neuropathy type C (DI-CMTC) to chromosome 1p34-p35. Here we identify two heterozygous missense mutations (G41R and E196K) and one de novo deletion (153-156delVKQV) in tyrosyl-tRNA synthetase (YARS) in three unrelated families affected with DI-CMTC. Biochemical experiments and genetic complementation in yeast show partial loss of aminoacylation activity of the mutant proteins, and mutations in YARS, or in its yeast ortholog TYS1, reduce yeast growth. YARS localizes to axonal termini in differentiating primary motor neuron and neuroblastoma cultures. This specific distribution is significantly reduced in cells expressing mutant YARS proteins. YARS is the second aminoacyl-tRNA synthetase found to be involved in CMT, thereby linking protein-synthesizing complexes with neurodegeneration.     

A critical role for the protein tyrosine phosphatase receptor type Z in functional recovery from demyelinating lesions

Several lines of evidence suggest that tyrosine phosphorylation is a key element in myelin formation, differentiation of oligodendrocytes and Schwann cells, and recovery from demyelinating lesions. Multiple sclerosis is a demyelinating disease of the human central nervous system, and studies of experimental demyelination indicate that remyelination in vivo requires the local generation, migration or maturation of new oligodendrocytes, or some combination of these. Failure of remyelination in multiple sclerosis could result from the failure of any of these processes or from the death of oligodendrocytes. Ptprz encodes protein tyrosine phosphatase receptor type Z (Ptpz, also designated Rptpbeta), which is expressed primarily in the nervous system but also in oligodendrocytes, astrocytes and neurons. Here we examine the susceptibility of mice deficient in Ptprz to experimental autoimmune encephalomyelitis (EAE), a model of multiple sclerosis. We observe that mice deficient in Ptprz show impaired recovery from EAE induced by myelin oligodendrocyte glycoprotein (MOG) peptide. This sustained paralysis is associated with increased apoptosis of mature oligodendrocytes in the spinal cords of mutant mice at the peak of inflammation. We further demonstrate that expression of PTPRZ1, the human homolog of Ptprz, is induced in multiple sclerosis lesions and that the gene is specifically expressed in remyelinating oligodendrocytes in these lesions. These results support a role for Ptprz in oligodendrocyte survival and in recovery from demyelinating disease.     

Neuroendocrine dysplasia in mice lacking protein tyrosine phosphatase sigma

Protein tyrosine phosphatase sigma (PTP-sigma, encoded by the Ptprs gene) is a member of the LAR subfamily of receptor-like protein tyrosine phosphatases that is highly expressed during mammalian embryonic development in the germinal cell layer lining the lateral ventricles of the developing brain, dorsal root ganglia, Rathke's pouch, olfactory epithelium, retina and developing lung and heart. On the basis of its expression and homology with the Drosophila melanogasterorthologues DPTP99 and DPTP100A (refs 5,6), which have roles in the targeting of axonal growth cones, we hypothesized that PTP-sigma may also have a modulating function in cell-cell interactions, as well as in axon guidance during mammalian embryogenesis. To investigate its function in vivo, we generated Ptprs-deficient mice. The resulting Ptprs-/-animals display retarded growth, increased neonatal mortality, hyposmia and hypofecundity. Anatomical and histological analyses showed a decrease in overall brain size with a severe depletion of luteinizing hormone-releasing hormone (LHRH)-immunoreactive cells in Ptprs-/- hypothalamus. Ptprs-/- mice have an enlarged intermediate pituitary lobe, but smaller anterior and posterior lobes. These results suggest that tyrosine phosphorylation-dependent signalling pathways regulated by PTP-sigma influence the proliferation and/or adhesiveness of various cell types in the developing hypothalamo-pituitary axis.     

Intragenic deletion in the gene encoding ubiquitin carboxy-terminal hydrolase in gad mice.

The gracile axonal dystrophy (gad) mouse is an autosomal recessive mutant that shows sensory ataxia at an early stage, followed by motor ataxia at a later stage. Pathologically, the mutant is characterized by 'dying-back' type axonal degeneration and formation of spheroid bodies in nerve terminals. Recent pathological observations have associated brain ageing and neurodegenerative diseases with progressive accumulation of ubiquitinated protein conjugates. In gad mice, accumulation of amyloid beta-protein and ubiquitin-positive deposits occur retrogradely along the sensory and motor nervous systems. We previously reported that the gad mutation was transmitted by a gene on chromosome 5 (refs 10,11). Here we find that the gad mutation is caused by an in-frame deletion including exons 7 and 8 of Uchl1, encoding the ubiquitin carboxy-terminal hydrolase (UCH) isozyme (Uch-l1) selectively expressed in the nervous system and testis. The gad allele encodes a truncated Uch-l1 lacking a segment of 42 amino acids containing a catalytic residue. As Uch-l1 is thought to stimulate protein degradation by generating free monomeric ubiquitin, the gad mutation appears to affect protein turnover. Our data suggest that altered function of the ubiquitin system directly causes neurodegeneration. The gad mouse provides a useful model for investigating human neurodegenerative disorders.     

Loss of BBS proteins causes anosmia in humans and defects in olfactory cilia structure and function in the mouse

Defects in cilia are associated with several human disorders, including Kartagener syndrome, polycystic kidney disease, nephronophthisis and hydrocephalus. We proposed that the pleiotropic phenotype of Bardet-Biedl syndrome (BBS), which encompasses retinal degeneration, truncal obesity, renal and limb malformations and developmental delay, is due to dysfunction of basal bodies and cilia. Here we show that individuals with BBS have partial or complete anosmia. To test whether this phenotype is caused by ciliary defects of olfactory sensory neurons, we examined mice with deletions of Bbs1 or Bbs4. Loss of function of either BBS protein affected the olfactory, but not the respiratory, epithelium, causing severe reduction of the ciliated border, disorganization of the dendritic microtubule network and trapping of olfactory ciliary proteins in dendrites and cell bodies. Our data indicate that BBS proteins have a role in the microtubule organization of mammalian ciliated cells and that anosmia might be a useful determinant of other pleiotropic disorders with a suspected ciliary involvement.     

Mutant dynactin in motor neuron disease

Impaired axonal transport in motor neurons has been proposed as a mechanism for neuronal degeneration in motor neuron disease. Here we show linkage of a lower motor neuron disease to a region of 4 Mb at chromosome 2p13. Mutation analysis of a gene in this interval that encodes the largest subunit of the axonal transport protein dynactin showed a single base-pair change resulting in an amino-acid substitution that is predicted to distort the folding of dynactin's microtubule-binding domain. Binding assays show decreased binding of the mutant protein to microtubules. Our results show that dysfunction of dynactin-mediated transport can lead to human motor neuron disease.     

Stochastic yet biased expression of multiple Dscam splice variants by individual cells

The Drosophila melanogaster gene Dscam is essential for axon guidance and has 38,016 possible alternative splice forms. This diversity can potentially be used to distinguish cells. We analyzed the Dscam mRNA isoforms expressed by different cell types and individual cells. The choice of splice variants expressed is regulated both spatially and temporally. Different subtypes of photoreceptors express broad yet distinctive spectra of Dscam isoforms. Single-cell RT-PCR documented that individual cells express several different Dscam isoforms and allowed an estimation of the diversity that is present. For example, we estimate that each R3/R4 photoreceptor cell expresses 14-50 distinct mRNAs chosen from the spectrum of thousands of splice variants distinctive of its cell type. Thus, the Dscam repertoire of each cell is different from those of its neighbors, providing a potential mechanism for generating unique cell identity in the nervous system and elsewhere.     

Identification and characterization of rod-derived cone viability factor

Retinitis pigmentosa is an untreatable, inherited retinal disease that leads to blindness. The disease initiates with the loss of night vision due to rod photoreceptor degeneration, followed by irreversible, progressive loss of cone photoreceptor. Cone loss is responsible for the main visual handicap, as cones are essential for day and high-acuity vision. Their loss is indirect, as most genes associated with retinitis pigmentosa are not expressed by these cells. We previously showed that factors secreted from rods are essential for cone viability. Here we identified one such trophic factor by expression cloning and named it rod-derived cone viability factor (RdCVF). RdCVF is a truncated thioredoxin-like protein specifically expressed by photoreceptors. The identification of this protein offers new treatment possibilities for retinitis pigmentosa.     

The mouse Ames waltzer hearing-loss mutant is caused by mutation of Pcdh15, a novel protocadherin gene

The neuroepithelia of the inner ear contain hair cells that function as mechanoreceptors to transduce sound and motion signals. Mutations affecting these neuroepithelia cause deafness and vestibular dysfuction in humans. Ames waltzer (av) is a recessive mutation found in mice that causes deafness and a balance disorder associated with the degeneration of inner ear neuroepithelia. Here we report that the gene that harbours the av mutation encodes a novel protocadherin. Cochlear hair cells in the av mutants show abnormal stereocilia by 10 days after birth (P10). This is the first evidence for the requirement of a protocadherin for normal function of the mammalian inner ear.     

Mutations in the RNA exosome component gene EXOSC3 cause pontocerebellar hypoplasia and spinal motor neuron degeneration

RNA exosomes are multi-subunit complexes conserved throughout evolution and are emerging as the major cellular machinery for processing, surveillance and turnover of a diverse spectrum of coding and noncoding RNA substrates essential for viability. By exome sequencing, we discovered recessive mutations in EXOSC3 (encoding exosome component 3) in four siblings with infantile spinal motor neuron disease, cerebellar atrophy, progressive microcephaly and profound global developmental delay, consistent with pontocerebellar hypoplasia type 1 (PCH1; MIM 607596). We identified mutations in EXOSC3 in an additional 8 of 12 families with PCH1. Morpholino knockdown of exosc3 in zebrafish embryos caused embryonic maldevelopment, resulting in small brain size and poor motility, reminiscent of human clinical features, and these defects were largely rescued by co-injection with wild-type but not mutant exosc3 mRNA. These findings represent the first example of an RNA exosome core component gene that is responsible for a human disease and further implicate dysregulation of RNA processing in cerebellar and spinal motor neuron maldevelopment and degeneration.     

Haploinsufficiency of protamine-1 or -2 causes infertility in mice

Protamines are the major DNA-binding proteins in the nucleus of sperm in most vertebrates and package the DNA in a volume less than 5% of a somatic cell nucleus. Many mammals have one protamine, but a few species, including humans and mice, have two. Here we use gene targeting to determine if the second protamine provides redundancy to an essential process, or if both protamines are necessary. We disrupted the coding sequence of one allele of either Prm1 or Prm2 in embryonic stem (ES) cells derived from 129-strain mice, and injected them into blastocysts from C57BL/6-strain mice. Male chimeras produced 129-genotype sperm with disrupted Prm1 or Prm2 alleles, but failed to sire offspring carrying the 129 genome. We also found that a decrease in the amount of either protamine disrupts nuclear formation, processing of protamine-2 and normal sperm function. Our studies show that both protamines are essential and that haploinsufficiency caused by a mutation in one allele of Prm1 or Prm2 prevents genetic transmission of both mutant and wild-type alleles.     

In vivo modulation of Hmgic reduces obesity

The HMGI family of proteins consists of three members, HMGIC, HMGI and HMGI(Y), that function as architectural factors and are essential components of the enhancesome. HMGIC is predominantly expressed in proliferating, undifferentiated mesenchymal cells and is not detected in adult tissues. It is disrupted and misexpressed in a number of mesenchymal tumour cell types, including fat-cell tumours (lipomas). In addition Hmgic-/- mice have a deficiency in fat tissue. To study its role in adipogenesis and obesity, we examined Hmgic expression in the adipose tissue of adult, obese mice. Mice with a partial or complete deficiency of Hmgic resisted diet-induced obesity. Disruption of Hmgic caused a reduction in the obesity induced by leptin deficiency (Lepob/Lepob) in a gene-dose-dependent manner. Our studies implicate a role for HMGIC in fat-cell proliferation, indicating that it may be an adipose-specific target for the treatment of obesity.     

Neurofibrillary tangles, amyotrophy and progressive motor disturbance in mice expressing mutant (P301L) tau protein

Neurofibrillary tangles (NFT) composed of the microtubule-associated protein tau are prominent in Alzheimer disease (AD), Pick disease, progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). Mutations in the gene (Mtapt) encoding tau protein cause frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), thereby proving that tau dysfunction can directly result in neurodegeneration. Expression of human tau containing the most common FTDP-17 mutation (P301L) results in motor and behavioural deficits in transgenic mice, with age- and gene-dose-dependent development of NFT. This phenotype occurred as early as 6.5 months in hemizygous and 4.5 months in homozygous animals. NFT and Pick-body-like neuronal lesions occurred in the amygdala, septal nuclei, pre-optic nuclei, hypothalamus, midbrain, pons, medulla, deep cerebellar nuclei and spinal cord, with tau-immunoreactive pre-tangles in the cortex, hippocampus and basal ganglia. Areas with the most NFT had reactive gliosis. Spinal cord had axonal spheroids, anterior horn cell loss and axonal degeneration in anterior spinal roots. We also saw peripheral neuropathy and skeletal muscle with neurogenic atrophy. Brain and spinal cord contained insoluble tau that co-migrated with insoluble tau from AD and FTDP-17 brains. The phenotype of mice expressing P301L mutant tau mimics features of human tauopathies and provides a model for investigating the pathogenesis of diseases with NFT.     

Amino-terminal fragments of mutant huntingtin show selective accumulation in striatal neurons and synaptic toxicity

Huntington disease (HD) is caused by expansion of a glutamine repeat in the amino-terminal region of huntingtin. Despite its widespread expression, mutant huntingtin induces selective neuronal loss in striatal neurons. Here we report that, in mutant mice expressing HD repeats, the production and aggregation of N-terminal huntingtin fragments preferentially occur in HD-affected neurons and their processes and axonal terminals. N-terminal fragments of mutant huntingtin form aggregates and induce neuritic degeneration in cultured striatal neurons. N-terminal mutant huntingtin also binds to synaptic vesicles and inhibits their glutamate uptake in vitro. The specific processing and accumulation of toxic fragments of N-terminal huntingtin in HD-affected striatal neurons, especially in their neuronal processes and axonal terminals, may contribute to the selective neuropathology of HD.