A central event in synapse development is formation of the presynaptic active zone in response to positional cues. Three active zone proteins, RIM, ELKS (also known as ERC or CAST) and Liprin-alpha, bind each other and are implicated in linking active zone formation to synaptic vesicle release. Loss of function in Caenorhabditis elegans syd-2 Liprin-alpha alters the size of presynaptic specializations and disrupts synaptic vesicle accumulation. Here we report that a missense mutation in the coiled-coil domain of SYD-2 causes a gain of function. In HSN synapses, the syd-2(gf) mutation promotes synapse formation in the absence of syd-1, which is essential for HSN synapse formation. syd-2(gf) also partially suppresses the synaptogenesis defects in syg-1 and syg-2 mutants. The activity of syd-2(gf) requires elks-1, an ELKS homolog; but not unc-10, a RIM homolog. The mutant SYD-2 shows increased association with ELKS. These results establish a functional dependency for assembly of the presynaptic active zone in which SYD-2 plays a key role.

Alcadeina (Alca) is an evolutionarily conserved type I membrane protein expressed in neurons. We show here that Alca strongly associates with kinesin light chain (K D E4-8 Â10 À9 M) through a novel tryptophan-and aspartic acid-containing sequence. Alca can induce kinesin-1 association with vesicles and functions as a novel cargo in axonal anterograde transport. JNK-interacting protein 1 (JIP1), an adaptor protein for kinesin-1, perturbs the transport of Alca, and the kinesin-1 motor complex dissociates from Alca-containing vesicles in a JIP1 concentration-dependent manner. Alca-containing vesicles were transported with a velocity different from that of amyloid b-protein precursor (APP)-containing vesicles, which are transported by the same kinesin-1 motor. Alca-and APP-containing vesicles comprised mostly separate populations in axons in vivo. Interactions of Alca with kinesin-1 blocked transport of APP-containing vesicles and increased b-amyloid generation. Inappropriate interactions of Alc-and APP-containing vesicles with kinesin-1 may promote aberrant APP metabolism in Alzheimer's disease.

Alzheimer's amyloid precursor protein (APP) 1 is an integral membrane protein with a receptor-like structure (1). A principal component of parenchymal amyloid deposits in Alzheimer's disease (AD) is ␤-amyloid (A␤) (2-4), which is derived from APP by proteolytic cleavage (1, 5-11). A␤ is thought to be generated through an intracellular protein secretory pathway of APP (for a review, see Ref. 12). The short APP cytoplasmic domain consisting of 47 amino acid residues is thought to be responsible for determination of APP metabolism (13-15) and possible signal transduction from a putative extracellular ligand that has yet to be identified (for a review, see Ref. 16). Because APP is a candidate pathogenic factor of AD, elucidation of the physiological function of APP as well as the determination of the metabolic mechanism of A␤ production should increase our understanding of the pathogenesis of AD. Using a yeast two-hybrid system, we isolated cDNA of proteins that interact with the cytoplasmic domain of APP (APP COOH ) in order to elucidate the molecular mechanisms of APP metabolism and function of APP. Previous efforts to identify and isolate proteins associating with APP COOH , utilizing the yeast two-hybrid system, have resulted in the isolation of proteins carrying phosphotyrosine binding/phosphotyrosine interaction (PI) domains such as Fe65 (17), 19), and X11 (20). The X11 gene, which is located on chromosome 9, was originally isolated as a gene candidate for Friedreich ataxia (21) and its partial cDNA was identified as a clone encoding a protein that associates with APP COOH (22). The PI domain of X11 interacts with the YENPTY motif of APP COOH (20). In the present study, we isolated a complete cDNA encoding an X11-like protein (X11L) from a human adult brain cDNA library, utilizing the yeast two-hybrid system and APP COOH as a bait. A partial short cDNA encoding approximately 190 amino acids in the PI domain of X11L has already been isolated using a similar procedure (22). However, detailed characterization of X11L binding to APP and identification of the role X11L plays in the physiological function of APP have not been performed. We found that human X11L requires a sequence containing the NPXY motif of APP COOH for APP binding and that association of the PI domain with APP was suppressed by a deletion of a amino-terminal domain fused to the PI domain (PI ϩ C construct) but enhanced by a deletion of a carboxyl-terminal domain fused to the PI domain (N ϩ PI construct). Co-transfection of full-length human X11L into cells that express stably transfected human APP695 cDNA resulted in decreased secretion of A␤40 but not A␤42. However, co-transfection into cells of the cDNA lacking the carboxyl-terminal domain (N ϩ PI construct) or a cDNA encoding only the PI domain (PI construct), whose protein products preserve the ability to bind to APP COOH , did not present the ability to modulate A␤ production. The present results suggest that the amino-terminal region of the PI domain is needed to regulate binding affinity...

We have isolated a novel protein based on its association with

SummaryLactic‐acid bacteria are widely recognized beneficial host associated groups of the microbiota of humans and animals. Some lactic‐acid bacteria have the ability to extend the lifespan of the model animals. The mechanisms behind the probiotic effects of bacteria are not entirely understood. Recently, we reported the benefit effects of Lactobacillus gasseri SBT2055 (LG2055) on animal and human health, such as preventing influenza A virus, and augmentation of IgA production. Therefore, it was preconceived that LG2055 has the beneficial effects on longevity and/or aging. We examined the effects of LG2055 on lifespan and aging of Caenorhabditis elegans and analyzed the mechanism of prolongevity. Our results demonstrated that LG2055 has the beneficial effects on longevity and anti‐aging of C. elegans. Feeding with LG2055 upregulated the expression of the skn‐1 gene and the target genes of SKN‐1, encoding the antioxidant proteins enhancing antioxidant defense responses. We found that feeding with LG2055 directly activated SKN‐1 activity via p38 MAPK pathway signaling. The oxidative stress response is elicited by mitochondrial dysfunction in aging, and we examined the influence of LG2055 feeding on the membrane potential of mitochondria. Here, the amounts of mitochondria were significantly increased by LG2055 feeding in comparison with the control. Our result suggests that feeding with LG2055 is effective to the extend lifespan in C. elegans by a strengthening of the resistance to oxidative stress and by stimulating the innate immune response signaling including p38MAPK signaling pathway and others.

APP associates with kinesin-1 via JIP1. In JIP1-decicient neurons, the fast velocity and high frequency of anterograde transport of APP cargo are impaired to reduced velocity and lower frequency, respectively. Interaction of JIP1 with KLC via two novel elements in JIP1 plays an important role in efficient APP axonal transport.

Liprin-α/SYD-2 activity promotes the polymerization of electron-dense projections in the presynaptic active zone through increased recruitment of ELKS-1/ELKS.

The ␤-amyloid peptide (A␤) 1 is a component of amyloid plaques, one of the major hallmarks of Alzheimer's disease (AD), and is implicated in the pathology of AD (reviewed in Ref. 1). A␤ is generated by proteolytic cleavage of the ␤-amyloid precursor protein (APP), an integral membrane protein with a short intracellular carboxyl terminus (2). Following translation, APP is subjected to N-glycosylation (immature APP) in the endoplasmic reticulum and additional O-glycosylation (mature APP) in the Golgi complex, transported to the plasma membrane, internalized by endocytosis, and delivered to endosomes and lysosomes (3-5). As it travels within the cell, APP is subject to proteolytic cleavage by ␣-or ␤-and ␥-secretases. In the minor, or amyloidogenic, processing pathway, the ␤-secretase, an aspartic protease BACE (6, 7), cleaves APP into two fragments, a large amino-terminal ectodomain (sAPP␤) and a truncated carboxyl-terminal fragment (CTF␤); cleavage takes place mainly in compartments of the secretory pathway (4, 8 -10). The CTF␤ is further cleaved by ␥-secretase, which results in the release of the A␤ peptides 40 (A␤40) or 42 (A␤42) (reviewed in Refs. 1 and 11). In contrast, the majority of APP is cleaved by ␣-secretase within the A␤ domain to produce sAPP␣ and CTF␣, either at the plasma membrane or in the secretary pathway (4, 12, 13). This non-amyloidogenic processing of APP results not in the secretion of A␤ peptides but rather of a small peptide (p3) that consists of the carboxyl-terminal half of A␤ generated by further cleavage of CTF␣ at the ␥-site. Like the full-length APP, the proteolytic fragments generated by either cleavage pathway are also proposed to play numerous roles in cell physiology. A␤ is widely known to be toxic for cells (14) and to be tightly associated with the progression of AD (reviewed in Ref. 1). The overexpression of human CTF␤ in mice causes neuronal and synaptic degeneration (15), the impairment of learning ability, and the suppression of long term potentiation (16). The secreted sAPP is thought to modulate neuronal function and cell survival (reviewed in Ref. 17). Recent reports suggest that the cytoplasmic tail fragment, which is generated from CTF by ␥-cleavage, may be implicated in transcriptional regulation (18). Therefore, an analysis of the intracellular mechanism of APP processing is important for understanding the pathogenesis of AD as well as the physiological function(s) of APP in normal tissues.The cytoplasmic domain of APP (APPcyt) has been suggested to be important for the regulation of intracellular trafficking and metabolism of APP (19). Substitutions of certain amino acids in APPcyt are known to affect the processing of APP. Substitution of Ala for Tyr-682 (with respect to the numbering convention for the APP 695 isoform), for Asn-684, and for Pro-685 in the 681 GYENPTY 687 motif results in the impaired internalization of APP from the plasma membrane and in a decrease in the secretion of sAPP and A␤ (20). Substitution of Ala for Arg-672 in the 667 VTPEER 672 motif inc...