The Ca 2؉ -independent immunoglobulin-like molecule nectin first forms cell-cell adhesion and then assembles cadherin at nectinbased cell-cell adhesion sites, resulting in the formation of adherens junctions (AJs). Afadin is a nectin-and actin filament-binding protein that connects nectin to the actin cytoskeleton. Here, we studied the roles and modes of action of nectin and afadin in the formation of AJs in cultured MDCK cells. The trans-interaction of nectin assembled E-cadherin, which associated with p120 ctn , ␤-catenin, and ␣-catenin, at the nectin-based cell-cell adhesion sites in an afadin-independent manner. However, the assembled E-cadherin showed weak cell-cell adhesion activity and might be the non-transinteracting form. This assembly was mediated by the IQGAP1-dependent actin cytoskeleton, which was organized by Cdc42 and Rac small G proteins that were activated by the action of trans-interacting nectin through c-Src and Rap1 small G protein in an afadinindependent manner. However, Rap1 bound to afadin, and this Rap1-afadin complex then interacted with p120 ctn associated with non-trans-interacting E-cadherin, thereby causing the trans-interaction of E-cadherin. Thus, nectin regulates the assembly and cellcell adhesion activity of E-cadherin through afadin, nectin signaling, and p120 ctn for the formation of AJs in Madin-Darby canine kidney cells.

Inactivation of a range of viruses, such as adeno-, mumps, rota-, polio- (types 1 and 3), coxsackie-, rhino-, herpes simplex, rubella, measles, influenza and human immunodeficiency viruses, by povidone-iodine (PVP-I) and other commercially available antiseptics in Japan was studied in accordance with the standardized protocol in vitro. In these experiments, antiseptics such as PVP-I solution, PVP-I gargle, PVP-I cream, chlorhexidine gluconate, alkyldiamino-ethyl-glycine hydrochloride, benzalkonium chloride (BAC) and benzethonium chloride (BEC) were used. PVP-I was effective against all the virus species tested. PVP-I drug products, which were examined in these experiments, inactivated all the viruses within a short period of time. Rubella, measles, mumps viruses and HIV were sensitive to all of the antiseptics, and rotavirus was inactivated by BAC and BEC, while adeno-, polio- and rhinoviruses did not respond to the other antiseptics. PVP-I had a wider virucidal spectrum, covering both enveloped and nonenveloped viruses, than the other commercially available antiseptics.

Junctional adhesion molecule (JAM) is a Ca2+ -independent immunoglobulin-like cell -cell adhesion molecule which localizes at tight junctions (TJs). Claudin is a key cell -cell adhesion molecule that forms TJ strands at TJs. JAM is associated with claudin through their cytoplasmic tail-binding protein, ZO-1. JAM is furthermore associated with Par-3, a cell polarity protein which forms a ternary complex with Par-6 and atypical protein kinase C. Nectin is another Ca 2+ -independent immunoglobulin-like cell -cell adhesion molecule which localizes at adherens junctions (AJs). Nectin is associated with E-cadherin through their respective cytoplasmic tail-binding proteins, afadin and catenins, and involved in the formation of AJs cooperatively with E-cadherin. We show here that nectin is furthermore involved in the localization of JAM at TJs. During the formation of the junctional complex consisting of AJs and TJs in Madin-Darby canine kidney (MDCK) cells, JAM was recruited to the nectin-based cell -cell adhesion sites. This recruitment of JAM was inhibited by nectin inhibitors, which inhibited the trans-interaction of nectin. Microbeads coated with the extracellular fragment of nectin, that interacted with cellular nectin, also recruited JAM to the bead -MDCK cell contact sites. Furthermore, when cadherin-deficient L fibroblasts stably expressing both exogenous JAM and nectin (nectin-JAM-L cells) were co-cultured with L fibroblasts expressing only nectin (nectin-L cells), JAM was concentrated at the cell -cell adhesion sites between nectin-JAM-L and nectin-L cells without the transinteraction of JAM. Analyses of the localization and immunoprecipitation of JAM revealed that it was associated with nectin through afadin and ZO-1. These results suggest that nectin has a role in the localization of JAM at TJs in the process of the formation of the junctional complex in epithelial cells.

Necl-5/Tage4/poliovirus receptor/CD155 has been shown to be the poliovirus receptor and to be up-regulated in rodent and human carcinoma. We have found previously that mouse Necl-5 regulates cell motility. We show here that mouse Necl-5 is furthermore involved in the regulation of cell proliferation. Studies using a specific antibody against Necl-5 and a dominant negative mutant of Necl-5 revealed that Necl-5 enhanced the serum-induced proliferation of NIH3T3, Swiss3T3, and mouse embryonic fibroblast cells. Necl-5 enhanced the serum-induced activation of the Ras-Raf-MEK-ERK signaling, up-regulated cyclins D2 and E, and down-regulated p27 Kip1 Necl-5/Tage4/poliovirus receptor (PVR) 1 /CD155 is an immunoglobulin (Ig)-like molecule having a domain structure consisting of one extracellular region with three Ig-like loops, one transmembrane region, and one cytoplasmic region (1-4). Human PVR/CD155 was originally identified as the human PVR (1, 2), whereas rodent Tage4 was originally identified as the product of a gene overexpressed in rat and mouse colon carcinoma (3, 4). PVR/CD155 has also been shown to be overexpressed in human colorectal carcinoma and malignant glioma (5, 6). The PVR/CD155 gene has thus far been found only in the primates, and the Tage4 gene has thus far been found only in the rodent, but these genes are likely to be derived from the common ancestor gene (7,8) and tentatively renamed nectinlike molecule-5, Necl-5 (for a review, see Ref. 9). Nectin-like molecules (Necls) have been named for a group of Ig-like molecules whose domain structures are similar to, but slightly different from, those of nectins (9). Nectins are Ca 2ϩ -independent Ig-like cell-cell adhesion molecules that constitute a family of four members, nectin-1, -2, -3, and -4 (for reviews, see Refs. 9 and 10). Nectins form cis-dimers followed by formation of trans-dimers (trans-interaction), eventually causing cell-cell adhesion. Nectins first form cell-cell adhesion where cadherins are recruited, resulting in formation of adherens junctions in epithelial cells and fibroblasts. Nectins are associated with the actin cytoskeleton through afadin, a nectin-and actin filamentbinding protein, as cadherins are associated with the actin cytoskeleton through ␣-and ␤-catenins (for a review, see Ref. 11). Necl-5 is one member of the Necl family consisting of five members, Necl-1, -2, -3, -4, and -5 (9). Although they have domain structures similar to those of nectins, they do not directly bind afadin.Nectin-3 forms not only homo-trans-dimers but also heterotrans-dimers (heterophilic trans-interaction) with either nectin-1 or -2 (9, 10). Nectin-4 forms hetero-trans-dimers with nectin-1, but nectin-1 does not form hetero-trans-dimers with nectin-2. These hetero-trans-dimers show much higher cell-cell adhesion activity than the homo-trans-dimers. In contrast to nectins, Necl-5 does not show homophilic cell-cell adhesion activity (12, 13). Thus, the role of Necl-5 as the PVR has been established, but its physiological role remained unknown for a...

The sequence data (H. Okamoto et al., Hepatol. Res. 10:1–16, 1998) of a newly discovered single-stranded DNA virus, TT virus (TTV), showed that it did not have the terminal structure typical of a parvovirus. Elucidation of the complete genome structure was necessary to understand the nature of TTV. We obtained a 1.0-kb amplified product from serum samples of four TTV carriers by an inverted, nested long PCR targeted for nucleotides (nt) 3025 to 3739 and 1 to 216 of TTV. The sequence of a clone obtained from serum sample TA278 was compared with those registered in GenBank. The complete circular TTV genome contained a novel sequence of 113 nt (nt 3740 to 3852 [=0]) in between the known 3′- and 5′-end arms, forming a 117-nt GC-rich stretch (GC content, 90.6% at nt 3736 to 3852). We found a 36-nt stretch (nt 3816 to 3851) with an 80.6% similarity to chicken anemia virus (CAV) (nt 2237 to 2272 of M55918), a vertebrate circovirus. A putative SP-1 site was located at nt 3834 to 3839, followed by a TATA box at nt 85 to 90, the first initiation codon of a putative VP2 at nt 107 to 109, the termination codon of a putative VP1 at nt 2899 to 2901, and a poly(A) signal at nt 3073 to 3078. The arrangement was similar to that of CAV. Furthermore, several AP-2 and ATF/CREB binding sites and an NF-κB site were arranged around the GC-rich region in both TTV and CAV. The data suggested that TTV is circular and similar to CAV in its genomic organization, implying that TTV is the first human circovirus.

Afadin is an actin filament (F-actin)-binding protein that is associated with the cytoplasmic tail of nectin, a Ca 2؉ -independent immunoglobulin-like cell-cell adhesion molecule. Nectin and afadin strictly localize at cellcell adherens junctions (AJs) undercoated with F-actin bundles and are involved in the formation of AJs in cooperation with E-cadherin in epithelial cells. In epithelial cells of afadin (؊/؊) mice and (؊/؊) embryoid bodies, the proper organization of AJs is markedly impaired. However, the molecular mechanism of how the nectin-afadin system is associated with the E-cadherincatenin system or functions in the formation of AJs has not yet been fully understood. Here we identified a novel afadin-binding protein, named ADIP (afadin DIL domain-interacting protein). ADIP consists of 615 amino acids with a calculated M r of 70,954 and has three coiledcoil domains. Northern and Western blot analyses in mouse tissues indicated that ADIP was widely distributed. Immunofluorescence and immunoelectron microscopy revealed that ADIP strictly localized at cell-cell AJs undercoated with F-actin bundles in small intestine absorptive epithelial cells. This localization pattern was the same as those of afadin and nectin. ADIP was undetectable at cell-matrix AJs. ADIP furthermore bound ␣-actinin, an F-actin-bundling protein known to be indirectly associated with E-cadherin through its direct binding to ␣-catenin. These results indicate that ADIP is an afadin-and ␣-actinin-binding protein that localizes at cell-cell AJs and may have two functions. ADIP may connect the nectin-afadin and E-cadherin-catenin systems through ␣-actinin, and ADIP may be involved in organization of the actin cytoskeleton at AJs through afadin and ␣-actinin.

Par-3 is a cell-polarity protein that regulates the formation of tight junctions (TJs) in epithelial cells, where claudin is a major cell-cell adhesion molecule (CAM). TJs are formed at the apical side of adherens junctions (AJs), where Ecadherin and nectin are major CAMs. We have revealed that nectin first forms cell-cell adhesions, and then recruits cadherin to nectin-based cell-cell adhesion sites to form AJs and subsequently recruits claudin to the apical side of AJs to form TJs. The cytoplasmic tail of nectin binds afadin and Par-3. Afadin regulates the formation of AJs and TJs cooperatively with nectin. Here, we studied the role of Par-3 in the formation of these junctions by using Par-3-knockdown MDCK cells. Par-3 was necessary for the formation of AJs and TJs but was not necessary for nectinbased cell-cell adhesion. Par-3 promoted the association of afadin with nectin, whereas afadin was not necessary for the association of Par-3 with nectin. However, the association of afadin with nectin alone was not sufficient for the formation of AJs or TJs, and Par-3 and afadin cooperatively regulated it. We describe here these novel roles of Par-3 in the formation of junctional complexes.

No abstract