The general secretion pathway (GSP) of Vibrio cholerae is required for secretion of proteins including chitinase, enterotoxin, and protease through the outer membrane. In this study, we report the cloning and sequencing of a DNA fragment from V. cholerae, containing 12 open reading frames, epsC to -N, which are similar to GSP genes of Aeromonas, Erwinia, Klebsiella, Pseudomonas, and Xanthomonas spp. In addition to the two previously described genes, epsE and epsM (M. Sandkvist, V. Morales, and M. Bagdasarian, Gene 123: 81-86, 1993; L. J. Overbye, M. Sandkvist, and M. Bagdasarian, Gene 132:101-106, 1993), it is shown here that epsC, epsF, epsG, and epsL also encode proteins essential for GSP function. Mutations in the eps genes result in aberrant outer membrane protein profiles, which indicates that the GSP, or at least some of its components, is required not only for secretion of soluble proteins but also for proper outer membrane assembly. Several of the Eps proteins have been identified by use of the T7 polymerase-promoter system in Escherichia coli. One of them, a pilin-like protein, EpsG, was analyzed also in V. cholerae and found to migrate as two bands on polyacrylamide gels, suggesting that in this organism it might be processed or otherwise modified by a prepilin peptidase. We believe that TcpJ prepilin peptidase, which processes the subunit of the toxin-coregulated pilus, TcpA, is not involved in this event. This is supported by the observations that apparent processing of EpsG occurs in a tcpJ mutant of V. cholerae and that, when coexpressed in E. coli, TcpJ cannot process EpsG although the PilD peptidase from Neisseria gonorrhoeae can.In Vibrio cholerae, the general secretion pathway (GSP) is required for extracellular secretion of several proteins including chitinase, cholera toxin (CT), and protease (51, 60). This pathway likely plays a significant role in the survival of Vibrio in different environments as well as in its pathogenicity. For instance, secreted chitinase and protease may assist in the detachment of V. cholerae from chitinous waterborne particles and from host epithelial cells, respectively, thus facilitating dissemination (14, 71). CT is the major component responsible for inducing the diarrheal disease cholera and is therefore also involved in the dissemination process (for a review, see reference 30).Escherichia coli heat-labile enterotoxin (LT) is very similar to CT in its structure, biochemical function, and ability to translocate across the outer membrane of V. cholerae (66). It is a multimeric protein which consists of a single A subunit and a pentamer of identical B-subunit polypeptides (18,64,65). Its secretion from V. cholerae proceeds in two steps, similar to the secretion of CT. The first step is translocation of the individual subunit polypeptides across the cytoplasmic membrane. This is mediated by a pathway which is presumably similar to the sec system of E. coli (63). During this step, the signal peptides of the A and B precursor polypeptides are cleaved off (8,25,52) a...

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Linking specific ecological factors to the evolution of parental care pattern and mating system is a difficult task of key importance. We provide evidence from comparative analyses that an ecological factor (breeding pool size) is associated with the evolution of parental care across all frogs. We further show that the most intensive form of parental care (trophic egg feeding) evolved in concert with the use of small pools for tadpole deposition and that egg feeding was associated with the evolution of biparental care. Previous research on two Peruvian poison frogs (Ranitomeya imitator and Ranitomeya variabilis) revealed similar life histories, with the exception of breeding pool size. This key ecological difference led to divergence in parental care patterns and mating systems. We present ecological field experiments that demonstrate that biparental care is essential to tadpole survival in small (but not large) pools. Field observations demonstrate social monogamy in R. imitator, the species that uses small pools. Molecular analyses demonstrate genetic monogamy in R. imitator, the first example of genetic monogamy in an amphibian. In total, this evidence constitutes the most complete documentation to date that a single ecological factor drove the evolution of biparental care and genetic and social monogamy in an animal.

Carcinoembryonic Ag-related cellular adhesion molecule 1 (CEACAM1) represents a group of transmembrane protein isoforms that consist of variable numbers of extracellular Ig-like domains together with either a long cytoplasmic (cyt) tail containing two immunoreceptor tyrosine-based inhibitory motifs or a unique short cyt tail. Although CEACAM1 has been reported to be expressed on the surface of T lymphocytes upon activation, its roles in T cell regulation are controversial due to the lack of functional characterization of each individual CEACAM1 isoform. We thus cotransfected Jurkat T cells with CEACAM1 isoform-encoding constructs and an IL-2 promoter-bearing plasmid or a small interference RNA targeting src homology domain 2 containing phosphatase 1. In a luciferase reporter assay and through measurements of cytokine secretion (IL-2, IL-4, and IFN-γ), CEACAM1 containing either a long or a short cyt tail inhibited or costimulated, respectively, TCR/CD3 complex plus CD28 mediated activation with the inhibitory functions of the long cyt tail dominating. The inhibitory function of CEACAM1, was dependent upon src homology domain 2 containing phosphatase 1 activity, required both tyrosine residues within the immunoreceptor tyrosine-based inhibitory motif domains of the cyt tail and was mediated through the mitogen-activated protein kinase pathway. CEACAM1-mediated inhibition could be functionally reconstituted by incubation of PBMC with either a CEACAM1-specific mAb or CEACAM1-Fc fusion protein in the presence of an allogeneic or mitogenic stimulus, respectively. These studies indicate that the long and short cyt tails of CEACAM1 serve as inhibitory and costimulatory receptors, respectively, in T cell regulation.