Presenilins are part of a protease complex that is responsible for the intramembraneous cleavage of the amyloid precursor protein involved in Alzheimer's disease and of Notch receptors. In Caenorhabditis elegans, mutations in the presenilin sel-12 result in a highly penetrant egg-laying defect. spr-5 was identi®ed as an extragenic suppressor of the sel-12 mutant phenotype. The SPR-5 protein has similarity to the human polyamine oxidase-like protein encoded by KIAA0601 that is part of the HDAC±CoREST corepressor complex. Suppression of sel-12 by spr-5 requires the activity of HOP-1, the second somatic presenilin in C.elegans. spr-5 mutants derepress hop-1 expression 20-to 30-fold in the early larval stages when hop-1 normally is almost undetectable. SPR-1, a C.elegans homologue of CoREST, physically interacts with SPR-5. Moreover, down-regulation of SPR-1 by mutation or RNA interference also bypasses the need for sel-12. These data strongly suggest that SPR-5 and SPR-1 are part of a CoREST-like co-repressor complex in C.elegans. This complex might be recruited to the hop-1 locus controlling its expression during development. Keywords: Alzheimer's disease/C.elegans/Notch/ presenilins/suppressor genetics IntroductionMutations in the presenilins PS1 and PS2 account for the majority of early-onset familial Alzheimer's disease (Selkoe, 2001). These mutations lead to the aberrant processing of the amyloid precursor protein (APP) and the generation of increasing amounts of the highly amyloidogenic form of Ab, the amyloid b-peptide. Ab is the predominant component of the plaques found in the brains of Alzheimer's patients. Presenilins are polytopic transmembrane proteins that are part of the proteolytic gsecretase complex that liberates Ab. Apart from their role in APP processing, presenilins are also required for proteolytic processing of the Notch receptors in their transmembrane domain. Ligand-induced cleavage and release of the intracellular domain of the Notch receptor (NICD) are crucial for nuclear Notch signalling (Struhl and Adachi, 1998). In agreement with these results, in all organisms tested so far, the loss of presenilin activity leads to phenotypes that resemble that of Notch loss-of-function mutants (Fortini, 2001).Similarly to humans, Caenorhabditis elegans has two somatically expressed presenilin genes, hop-1 and sel-12 (Levitan and Greenwald, 1995;Li and Greenwald, 1997). hop-1 and sel-12, like PS1 and PS2, show redundant activities since only double mutants display phenotypes associated with a complete loss of Notch signalling in C.elegans (Li and Greenwald, 1997;Westlund et al., 1999). A sel-12 null mutant can be rescued by transgenic expression of hop-1 (as well as either of the human PS1 or PS2). sel-12 is expressed rather uniformly at all developmental stages, while hop-1 expression is very weak and could not be detected by reporter gene fusions (Westlund et al., 1999). Probably as a consequence of these different levels of expression, hop-1 mutants are viable with no obvious phenotype, whereas sel...

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Mutations in presenilin genes impair Notch signalling and, in humans, have been implicated in the development of familial Alzheimer's disease. We show here that a reduction of the activity of the Caenorhabditis elegans presenilin sel-12 results in a late defect during sex muscle development. The morphological abnormalities and functional deficits in the sex muscles contribute to the egg-laying defects seen in sel-12 hermaphrodites and to the severely reduced mating efficiency of sel-12 males. Both defects can be rescued by expressing sel-12 from the hlh-8 promoter that is active during the development of the sex muscle-specific M lineage, but not by expressing sel-12 from late muscle-specific promoters. Both weak and strong sel-12 mutations cause defects in the sex muscles that resemble the defects we found in lin-12 hypomorphic alleles, suggesting a previously uncharacterised LIN-12 signalling event late in postembryonic mesoderm development. Together with a previous study indicating a role of lin-12 and sel-12 during the specification of the pi cell lineage required for proper vulva-uterine connection, our data suggest that the failure of sel-12 animals to lay eggs properly is caused by defects in at least two independent signalling events in different tissues during development.

The POU homeodomain protein UNC-86 and the LIM homeodomain protein MEC-3 are essential for the differentiation of the six mechanoreceptor neurons in the nematode Caenorhabditis elegans. Previous studies have indicated that UNC-86 and MEC-3 bind cooperatively to at least three sites in the mec-3 promoter and synergistically activate transcription. However, the molecular details of the interactions of UNC-86 with MEC-3 and DNA have not been investigated so far. Here we used a yeast system to identify the functional domains in UNC-86 required for transcriptional activation and to characterize the interaction of UNC-86 with MEC-3 in vivo. Our results suggest that transcriptional activation is mediated by the amino terminus of UNC-86, whereas amino acids in the POU domain mediate DNA binding and interaction with MEC-3. By random mutagenesis, we identified mutations that only affect the DNA binding properties of UNC-86, as well as mutations that prevent coactivation by MEC-3. We demonstrated that both the POU-specific domain and the homeodomain of UNC-86, as well as DNA bases adjacent to the proposed UNC-86 binding site, are involved in the formation of a transcriptionally active complex with MEC-3. These data suggest that some residues involved in the contact of UNC-86 with MEC-3 also contribute to the interaction of the functionally nonrelated POU protein Oct-1 with Oca-B, whereas other positions have different roles.POU domain transcription factors are characterized by their bipartite DNA binding domain, consisting of a helix-turn-helix POU-specific domain (POU S ) and an adjacent POU homeodomain (POU HD ) (for reviews, see references 15 and 28). Both protein domains contact DNA and are necessary for high-affinity DNA binding (15). POU class IV is comprised of the mammalian Brn-3-encoding genes, the Drosophila I-POU/ acj6 gene, and the Caenorhabditis elegans unc-86 gene (28). These are all expressed exclusively in the nervous system, but their expression is not limited to one specific neuronal cell type. In vitro studies have shown that POU class IV proteins bind very similar DNA sequences (12, 24). Therefore, it has been proposed that any differences in target gene activation are due to modulatory protein interactions (28). Most of the data for determination of the promoter specificity of POU proteins are derived from the nonrelated human POU class III protein Oct-1 (13, 41).POU homeobox gene unc-86 is expressed in 57 neurons in adult C. elegans. These neurons comprise one-fifth of the animal's nervous system and represent 27 different functional classes (10). Consequently, null alleles of unc-86 result in several behavioral defects affecting mechanosensation, egg laying, chemosensation, and thermosensation and cause an uncoordinated phenotype (5,10,18,26,37). The diversity of neuron types that require UNC-86 raises the possibility that in different cell types, UNC-86 targets the promoters of different genes. This is supported by the fact that in at least two unc-86 mutants, only a subset of the unc-86-mediated be...

The Caenorhabditis elegans POU protein UNC-86 specifies the HSN motor neurons, which are required for egg-laying, and six mechanosensory neurons. To investigate how UNC-86 controls neuronal specification, we characterized two unc-86 mutants that do not respond to touch but show wild-type egg-laying behavior. Residues P145 and L195, which are altered by these mutations, are located in the POU-specific domain and abolish the physical interaction of UNC-86 with the LIM homeodomain protein, MEC-3. This results in a failure to maintain mec-3 expression and in loss of expression of the mechanosensory neuron-specific gene, mec-2. unc-86-dependent expression of genes in other neurons is not impaired. We conclude that distinct residues in the POU domain of UNC-86 are involved in modulating UNC-86 activity during its specification of different neurons. A structural model of the UNC-86 POU domain, including base pairs and amino acid residues required for MEC-3 interaction, revealed that P145 and L195 are part of a hydrophobic pocket which is similar to the OCA-B-binding domain of the mammalian POU protein, Oct-1.