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The neuronal ceroid lipofuscinoses (NCLs, Batten disease) are fatal inherited neurodegenerative diseases. Sheep affected with the CLN6 form provide a valuable model to investigate underlying disease mechanisms from preclinical stages. Excitatory neuron loss in these sheep is markedly regional, localized early reactive changes accurately predicting neuron loss and subsequent symptom development. This investigation of GABAergic interneuron loss revealed similar regional effects that correlate with symptoms. Loss of parvalbumin positive neurons from the affected cortex was apparent at four months and became profound by 19 months, as was somatostatin positive neuron loss to a lesser extent. Conversely calbindin and neuropeptide Y positive neurons were relatively preserved and calretinin staining temporarily increased. Staining of subcortical regions was more intense but subcortical architecture remained relatively intact. Discrete subcortical changes followed from cortical changes in interconnected regions. These data highlight cellular location and interconnectivity as the major determinants of neuron survival, rather than phenotype.

Many fungal pH responses depend upon conserved Rim101p/PacC transcription factors, which are activated by C-terminal proteolytic processing. The means by which environmental pH is sensed by this pathway are not known. Here, we report a screen of the Saccharomyces cerevisiae viable deletion mutant library that has yielded a new gene required for processed Rim101p accumulation, DFG16. An S. cerevisiae dfg16Delta mutant expresses Rim101p-repressed genes at elevated levels. In addition, Candida albicans dfg16Delta/dfg16Delta mutants are defective in alkaline pH-induced filamentation, and their defect is suppressed by expression of truncated Rim101-405p. Thus, Dfg16p is a functionally conserved Rim101p pathway member. Many proteins required for processed Rim101p accumulation are members of the ESCRT complex, which functions in the formation of multivesicular bodies (MVBs). Staining with the dye FM4-64 indicates that the S. cerevisiae dfg16Delta mutant does not have an MVB defect. We find that two transcripts, PRY1 and ASN1, respond to mutations that affect both the Rim101p and MVB pathways but not to mutations that affect only one pathway. The S. cerevisiae dfg16Delta mutation does not affect PRY1 and ASN1 expression, thus confirming that Dfg16p function is restricted to the Rim101p pathway. Dfg16p is homologous to Aspergillus nidulans PalH, a component of the well-characterized PacC processing pathway. We verify that the previously recognized PalH homolog, Rim21p, also functions in the S. cerevisiae Rim101p pathway. Dfg16p is predicted to have seven membrane-spanning segments and a long hydrophilic C-terminal region, as expected if Dfg16p were a G-protein-coupled receptor.

The MTG1 gene of Saccharomyces cerevisiae, corresponding to ORF YMR097c on chromosome XIII, codes for a mitochondrial protein essential for respiratory competence. A human homologue of Mtg1p capable of partially rescuing the respiratory deficiency of a yeast mtg1 mutant has also been localized in mitochondria. Mtg1p is a member of a family of GTPases with largely unknown functions. The respiratory deficiency of mtg1 mutants stems from a defect in mitochondrial protein synthesis. Mutations in the 21S rRNA locus are able to suppress the translation defect of mtg1 null mutants. This points to the 21S rRNA or the large ribosomal subunit as the most likely target of Mtg1p action. The presence of mature size 15S and 21S mitochondrial rRNAs in mtg1 mutants excludes Mtg1p from being involved in transcription or processing of these RNAs. More likely, Mtg1p functions in assembly of the large ribosomal subunit. This is consistent with the peripheral localization of Mtg1p on the matrix side of the inner membrane and the results of in vivo mitochondrial translation assays in a temperature-sensitive mtg1 mutant.

The gene involved in the classical juvenile form of Batten disease, CLN3 has been identified as being highly homologous to the Saccharomyces cerevisiae YHC3 gene. To provide a simple model for the biochemical events underlying this disease, several disruptions have been made in YHC3 in three different S. cerevisiae strains. No obvious growth differences were observed, and neither was the previously reported phenotypic difference between wild-type and yeast disrupted in YHC3.

The neuronal ceroid lipofuscinoses (NCLs) are a group of inherited degenerative neurological diseases affecting children. A number of non-allelic variants have been identified within the human population and the genes for some of these have recently been identified. The underlying mechanism for the neuropathology remains an enigma; however, pioneering studies with the naturally occurring ovine model (OCL) have led to the proposal that these diseases represent lesions in specific hydrophobic protein degradation pathways. In this study, we show linkage between OCL and microsatellite markers on OAR 7q13-15. Using interspecies chromosome painting we establish that OAR 7q13-15 is syntenic with human chromosome 15q21-23, the region which was recently defined as the location of a newly identified late infantile variant (CLN6). We propose that our ovine model represents a mutation in the gene orthologous to that mutated in the human late infantile variant CLN6. The ovine linkage flock, consisting of 56 families, represents a powerful resource for positional cloning of this NCL gene. The availability of such a large animal model will have important implications for experimentation in downstream corrective therapies.