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Polycystic lipomembranous osteodysplasia with sclerosing leukoencephalopathy (PLOSL; MIM 221770), also known as Nasu-Hakola disease, is a recessively inherited disease characterized by a combination of psychotic symptoms rapidly progressing to presenile dementia and bone cysts restricted to wrists and ankles. PLOSL has a global distribution, although most of the patients have been diagnosed in Finland and Japan, with an estimated population prevalence of 2x10-6 (ref. 2) in the Finns. We have previously identified a shared 153-kb ancestor haplotype in all Finnish disease alleles between markers D19S1175 and D19S608 on chromosome 19q13.1 (refs 5,6). Here we characterize the molecular defect in PLOSL by identifying one large deletion in all Finnish PLOSL alleles and another mutation in a Japanese patient, both representing loss-of-function mutations, in the gene encoding TYRO protein tyrosine kinase binding protein (TYROBP; formerly DAP12). TYROBP is a transmembrane protein that has been recognized as a key activating signal transduction element in natural killer (NK) cells. On the plasma membrane of NK cells, TYROBP associates with activating receptors recognizing major histocompatibility complex (MHC) class I molecules. No abnormalities in NK cell function were detected in PLOSL patients homozygous for a null allele of TYROBP.

Abstract. The envelope of the Semliki Forest virus (SFV) contains two transmembrane proteins, E2 and El, in a heterodimeric complex. The E2 subunit is initially synthesized as a precursor protein p62, which is proteolytically processed to the mature E2 form before virus budding at the plasma membrane. The p62 (E2) protein mediates binding of the heterodimer to the nucleocapsid during virus budding, whereas El carries the entry functions of the virus, that is, cell binding and low pH-mediated membrane fusion activity. We have investigated the significance of the cleavage event for the maturation and entry of the virus . To express SFV with an uncleaved p62 phenotype, BHK21 cells were transfected by electroporation with infectious viral RNA transcribed from a full-length SFV cDNA clone in which the p62 cleavage site had been changed . The uncleaved p62E1 heterodimer was found to be T HE spreading of enveloped animal viruses between cells is dependent on their ability to mature by budding at the membranes of the infected cells and to enter new cells by the process ofmembrane fusion. To perform these functions these viruses use different membrane proteins . Proteins located at the internal side of the membrane, like the M protein of orthomyxo-, paramyxo-, and rhabdoviruses, as well as the N11 2-terminal part of the gag precursor protein of retroviruses, seem to play a major role during virus assembly, whereas other proteins forming spike-like surface projections carry cell-binding and membrane fusion functions. A control mechanism ensures that assembly and entry functions do not interfere with each other. Typical for this regulation is that the functions required for virus entry are not activated before the final stage of maturation has been reached and the virus is released from the cell Dubois-Dalcq et al., 1984;Stegmann et al., 1989;Kielian and Jungerwirth, 1990;White, 1990;Pettersson, 1991) . For most enveloped viruses a host cell-mediated limited proteolysis of the viral fusion protein initiates this activation process . In some cases the cleavage generates a fusion protein that is activated upon Mario Lobigs7s present address is 36 Wattle St ., Lyneham, ACT 2602, Australia .

In addition, we have used this in vitro assay to examine the biochemical effects of several mutations in Sec9 that result in pronounced growth defects in vivo. As expected, a temperature-sensitive mutation in the region most highly conserved between Sec9 and SNAP-25 is severely diminished in its ability to bind Sso1 and Snc1 in vitro. In contrast, a temperature-sensitive mutation near the C terminus of Sec9 shows no defect in SNARE binding in vitro. Similarly, a deletion of the Cterminal 17 residues, which is lethal in vivo, also binds Sso1 and Snc1 normally in vitro. Interestingly, we find that these same two C-terminal mutants, but not mutants that show SNARE assembly defects in vitro, act as potent dominant negative alleles when expressed behind a strong regulated promoter. Taken together these results suggest that the C-terminal domain of Sec9 is specifically required for a novel interaction that is required at a step following SNARE assembly.In recent years, studies on neuronal exocytosis, biochemical analysis of in vitro transport systems, and yeast genetic analysis have converged on a set of structurally related proteins known as SNARE proteins, as critical for the process of vesicle targeting and fusion in eukaryotic cells (1). This has led to the SNARE hypothesis, which suggests that SNARE proteins on the surface of vesicles (v-SNAREs) 1 can interact specifically with SNARE proteins on the target membrane (t-SNAREs) to form a complex that recruits factors required for fusion of the two membrane bilayers (2). In neurons the vesicular protein, synaptobrevin, associates with the two plasma membrane proteins syntaxin and soluble SNAP-25 to form a complex that acts as a receptor for ␣-SNAP and NSF. Hydrolysis of ATP by NSF leads to SNARE complex disassembly and is thought to be linked to membrane fusion, although precisely how NSF is involved in the membrane fusion event is still unclear (3).Binding studies with recombinant neuronal proteins have shown that although syntaxin and SNAP-25 can bind to one another with high affinity, they can also individually bind to the vesicle protein, synaptobrevin, with lower affinities (4). The presence of both t-SNAREs, however, potentiates the interaction of syntaxin with synaptobrevin about 10-fold and that of SNAP-25 with synaptobrevin approximately 2-fold. This suggests that the formation of a highly stable ternary complex drives the interaction between these proteins and consequently aids in determining the overall specificity of synaptic vesicle docking (4).The regions of each neuronal SNARE protein that mediate these binding activities have been extensively characterized. Within the cytoplasmic domain of syntaxin 1, for example, a small 73-residue juxtamembrane region, predicted to form coiled-coils, can mediate both binding to synaptobrevin (5, 6) and 7,8). The amino-terminal region of syntaxin is also capable of binding the carboxyl-terminal domain of the protein, and this interaction appears to be somewhat inhibitory to synaptobrevin binding (5). The amino-termi...