The Drosophila melanogaster flightless I gene is required for normal cellularization of the syncytial blastoderm. Highly conserved homologues of flightless I are present in Caenorhabditis elegans, mouse, and human. We have disrupted the mouse homologue Fliih by homologous recombination in embryonic stem cells. Heterozygous Fliih mutant mice develop normally, although the level of Fliih protein is reduced. Cultured homozygous Fliih mutant blastocysts hatch, attach, and form an outgrowing trophoblast cell layer, but egg cylinder formation fails and the embryos degenerate. Similarly, Fliih mutant embryos initiate implantation in vivo but then rapidly degenerate. We have constructed a transgenic mouse carrying the complete human FLII gene and shown that the FLII transgene is capable of rescuing the embryonic lethality of the homozygous targeted Fliih mutation. These results confirm the specific inactivation of the Fliih gene and establish that the human FLII gene and its gene product are functional in the mouse. The Fliih mouse mutant phenotype is much more severe than in the case of the related gelsolin family members gelsolin, villin, and CapG, where the homozygous mutant mice are viable and fertile but display alterations in cytoskeletal actin regulation.We are studying the mammalian homologues of a number of Drosophila melanogaster genes concerned with development or behavior, as part of a program aimed at identifying novel mammalian developmental and neurobiological genes. The D. melanogaster flightless I (fliI) gene (4,15,23,24,33) is required for cellularization of the syncytial blastoderm. With severe mutations in fliI, when the contribution of maternal product is eliminated, cellularization is only partial and gastrulation fails (35,44).

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Homologues of the Drosophila melanogaster tweety (tty) gene are present in mammals and Caenorhabditis elegans. The encoded proteins have five predicted membrane-spanning regions and recent findings suggest that some family members may be chloride channels. Phylogenetic analysis of the tty family including novel members from slime mould Entamoeba and plants has revealed the occurrence of independent gene duplication events in different lineages. expressed sequence tag data indicate that expression of the mammalian Ttyh1 gene is restricted mainly to neural tissue and is up-regulated in astrocytoma, glioma and several other cancers. In this study, mammalian expression vectors were used to investigate the subcellular localization and the effect of over-expression of Ttyh1 in human epithelial kidney cells. The results confirm that Ttyh1 is a membrane protein and show that it is deposited on the substratum along the migration paths of motile cells above the a5b1-integrin complex. The ectopic expression of Ttyh1 also induced long filopodia, which were branched and dynamic in both stationary and migratory cells. The filopodia contained F-actin and occurred at the ends of microtubules which were polarized towards the membrane. Upon contact with nearby cells some filopodia stabilized and filled with F-actin, whereas Ttyh1 was highly concentrated at the cell-cell interface. Ttyh1 N-and C-terminal antipeptide antibodies detected Ttyh1 along the axons of neurones in primary rat hippocampal cell cultures, and in situ in whole rat brain slices around the hippocampus and occasionally between cells. These data suggest a role for Ttyh1 in process formation, cell adhesion and possibly as a transmembrane receptor.

The 70 kDa ribosomal S6 kinase (p70 S6K ) is activated through the phosphoinositide (PI) 3-kinase-regulated pathway (Fig. 1). This pathway is responsible for the generation of inositol lipids, which are key mediators of intracellular signalling. Multiple isoforms of PI 3-kinase have been identified and classed according to substrate specificity and structure. 1 Class I PI 3-kinase is involved in receptor-induced hormonal responses and the mechanism of activation of this enzyme differs depending on the particular extracellular stimuli. For tyrosine kinase-coupled receptor systems, p110 catalytic subunits α, β and δ are stimulated through interaction with the p85 adaptor subunit, which binds selectively through its SH2 domain to phosphorylated tyrosines on the activated receptor. 2 For G-protein-coupled receptor systems, the signalling for PI 3-kinase activation is less clear and several mechanisms have been suggested. A number of reports have identified a p110γ catalytic subunit, which is activated through binding either directly to the βγ subunits of activated trimeric G-proteins 3 or through interaction with a p101 adaptor protein. 4,5 Other studies have suggested alternative mechanisms involving G-protein-mediated activation of receptor tyrosine kinase activity 6 or through the p110β isoform, which can also be activated directly by G-protein βγ subunits. 7 For both receptor systems, PI 3-kinase catalyses the phosphorylation of phosphoinositides at the 3′-OH position. For hormone-induced responses in vivo, phosphatidylinositol 4,5-bisphosphate (PI-4,5P 2 ) is the primary substrate for PI 3-kinase, leading to the production of phosphatidylinositol 3,4,5-trisphosphate (PI-3,4,5P 3 ). Phosphatidylinositol 3,4,5P 3 is also rapidly dephosphorylated by 5′-phosphatases, giving rise to the production of phosphatidylinositol 3,4-bisphosphate (PI-3,4P 2 ). 8 Although the mechanism regulating the relative levels of these two phospholipids is complex, both are thought to act as second messengers in cellular responses such as mitosis, apoptosis, membrane trafficking, motility, differentiation and oncogenic transformation by targeting certain pleckstrin homology (PH) domain-containing proteins to the plasma membrane. 9 Two isoforms of p70 S6K have been identified: a 70 kDa cytoplasmic form and a 85 kDa nuclear form. The sequence of the longer form includes a 23 amino acid nuclear localization sequence at the amino terminus. The molecular sequences of these two isoforms are otherwise the same and can be jointly referred to as p70 S6K or S6K1. Recently, a second functional homologue, S6K2, has been identified. This second S6 kinase shows similar sensitivity to rapamycin and PI 3-kinase inhibitors and is upregulated in p70 S6K1 -deficient mice. 10,11 The identification of S6K2 has raised the possibility that some functional responses of S6K1 may be caused by activation of S6K2 or even other unidentified S6 kinases.Activation of p70 S6K occurs through a complex series of phosphorylation events on eight or more serine or threoni...

Interleukin-8 (IL-8